Aegilops L.

Abstract

This chapter discusses the classification of the genus Aegilops, and presents a detailed description of its sections and species. It shows the morphology, geographical distribution, ecological affinities, cytology, and cytogenetic aspects of the species. Similarly, the structure and distribution of repetitious DNA in the various species, evolution of the diploid and genome analyses of the allopolyploid species, and relationships between them and to species of Triticum, are presented. The occurrence of gametocidal (GC) genes in species of Aegilops, their mode of action, evolutionary significance, and use in the production of deletion and dissection bread wheat lines, are also being reviewed.

9.1 Classification of the Genus Aegilops L.

Aegilops is the name of a grass mentioned in Theophrastus’ botanical treatise “Enquiry into Plants”, that was a major source for botanical knowledge during antiquity and the Middle Ages. The name Aegilops comes from the Greek aegilos, which could mean “a herb liked by goats”, or “a goat-like herb”, and refers to the whiskery-awned spikelets of some of its species (Bor 1968; Watson and Dallwitz 1992). Since the taxonomic treatment of the Aegilops genus by Linnaeus (1753), various taxonomists provided different definitions of the species and sub-genus ranks of the genus. Zhukovsky (1928) described 20 species in the genus, which he classified into nine sections, while Eig (1929a) grouped 22 species into two sub-genera and six sections. In his review summarizing results from a genome analysis of the genus, Kihara (1954) recognized 21 species of which the tetraploid and hexaploid taxa of Ae. triaristata and Ae. crassa, were considered one species. He grouped the 21 species into six sections. Based on his karyomorphological study, Chennaveeraiah (1960) separated Ae. vavilovii as a new hexaploid species from Ae. crassa. Following Chennaveeraiah (1960), Kihara and Tanaka (1970) separated also the hexaploid taxon Ae. recta from the tetraploid Ae. neglecta, and accepted the separation of the hexaploid Ae. vavilovii from Ae. crassa. Consequently, Kihara and Tanaka (1970) grouped a total of 22 species into six sections. Later, Feldman and Kislev (1977) described Ae. searsii as a new diploid species belonging to section Sitopsis. While accepting Ae. searsii as a valid species, Hammer (1980) continue to consider Ae. vavilovii as a subspecies of Ae. crassa and grouped 22 species into three subgenera and four sections. All of the above classifications included Ae. mutica either as a separate section (Zhukovsky 1928; Kihara 1954; Kihara and Tanaka 1970) or as a sub-genus (Eig 1929a; Hammer 1980). Yet, Eig (1929b) removed Ae. mutica from the genus Aegilops and included it as a monotypic species in a new genus Amblyopyrum. Lastly, van Slageren (1994), in his recent comprehensive taxonomic classification of the genus Aegilops, recognized five sections containing 22 species. It included Ae. vavilovii and Ae. searsii as species, but kept Ae. recta in Ae. neglecta. This book recognizes Ae. recta as a species and, consequently, 23 Aegilops species are considered here: 10 diploids, 9 tetraploids, one species Ae. crassa containing two cytotypes, tetraploid and hexaploid, and 3 hexaploids (Table 9.1). A 23-species genus, instead of the usual 1–5, is exceptional in the sub-tribe Triticineae.

Table 9.1 Sections and species of Aegilops and their synonyms

An entirely different tendency exists regarding the sub-species level classification taxa (van Slageren 1994). Hammer (1980) suggested to regard taxa with different chromosome numbers, or differing in their morphology and/or geographical distribution, as subspecies. Van Slageren (1994), like Mac Key (1981), disagreed and suggested a drastic consolidation at the intra-specific level, i.e., to maintain only groups exhibiting obvious discontinuities in several characters. Kihara (1954) included Ae. kotschyi, Ae. heldreichii, Ae. aucheri and Ae. sharonensis in Ae. variabilis (now Ae. peregrina), Ae. comosa, Ae. speltoides and Ae. longissima, respectively, because they shared very similar genomes. However, like Eig (1929a), Hammer (1980) considered Ae. kotschyi a separate species, because it differs from Ae. peregrina in its exclusive geographical distribution, i.e., the two species are good vicariad species. Similarly, van Slageren (1994) accepted Eig’s (1929a) definition of Ae. sharonensis as a valid species due to its morphological and ecological distinction from Ae. longissima. On the other hand, van Slageren (1994) maintained the sub-specific rank of Ae. heldreichii and Ae. aucheri (Table 9.1).

The genus Aegilops L. [Syn.: Agicon Adans; Triticum L. Sect. Aegilops (L.) Godr. & Gren. in Grenier & Godron; Triticum L. Subg. Aegilops (L.) Schmahlh.; Frumentum E.H.L. Krause subg. Aegilops (L.) E.H.L. Krauase; Aegilops L. subg. Eu-Aegilops Eig; aegilopoides Á. Löve] consists of wild annual, mostly autogamous 15–100-cm-high grasses, with few to many tillers that are usually geniculate at the base then turning upright, and sparsely foliated in the lower parts. The leaf blades are flat, with short ligules. The leaves are linear and up to 15-cm long and one cm wide. The spikes are linear, or ovate to lanceolate, wholly or partly awned or awnless, with 2–20 solitary spikelets at each rachis node, with each spikelet up to 1.2-cm-long, sessile, or sub-sessile. Spikes contain 1–4 rudimentary spikelets at the base or the top. The spikes break off at maturity above the rudimentary spikelets and fall entire or disarticulate into single spikelets. Spikelets contain 2–8 florets, the upper often being staminate or sterile. The two glumes are more or less equal, shorter than the adjacent lemmas, or almost as long as the adjacent lemmas, with one or more teeth or awns, round on back, rarely keeled. The lemmas are papery or membranous with 1–3 teeth, or awns. The two palea are keeled. The caryopsis either adheres to the lemma and/or to the palea or is free from both lemma and palea. In some species, all the spikelets are fully awned, whereas in others, only the terminal spikelets are awned. Certain lines of several species (e.g., Ae. bicornis, Ae. sharonensis, Ae. peregrina) are awnless. Three types of dispersal units exist in the different species: wedge (the spikes disarticulate at maturity into spikelets with the rachis internode that belong to them), barrel (the spikes disarticulate at maturity into spikelets with the rachis internode that belong to the spikelet above them), and umbrella (the spikes fall entire at maturity) types.

The genus Aegilops is distinguished from the genus Triticum by the absence of well-developed keel on the glumes, causing the sharp angle in the glume outline of both the wild and domesticated Triticum taxa (van Slageren 1994). It also differs from Triticum by its glabrous rachis and a larger number of grains per spikelet.

9.2 Geographical Distribution and Ecological Affinities

The genus is a Mediterranean–western Asiatic element (Eig 1929a, 1936; Sakamoto 1973; Feinbrun-Dothan 1986a; van Slageren 1994; Hegde et al. 2002), containing species that are distributed from the Iberian Peninsula in the west, through the Mediterranean basin, southern Ukraine, Crimea, the Caucasus, the Middle East, and to central Asia and western China (Zhukovsky 1928; Eig 1929a, 1936; Kihara 1954; Miller 1987; Kimber and Feldman 1987; van Slageren 1994; Table 9.2), i.e., from about 10°W to 82°E and about 24°S to 47°N (van Slageren 1994). The area of distribution is rectangular in shape, with its width being about four times its length. In central, northern, and eastern Europe, the distribution of the genus is bordered by the cold climate, in North Africa by the Saharan desert, in the southwest by the deserts of Sinai and the Arabian peninsulas, in the central-northern Asia by the steppes of Turkmenistan and Uzbekistan, and in the east by the Tian Shan and Himalaya mountain ranges and in the south east by the banks of the Indus river (Zhukovsky 1928; Eig 1929a, 1936; Kihara 1954; Kimber and Feldman 1987; van Slageren 1994). Several allotetraploid species introduced in the USA, of which Ae. cylindrica (jointed goat grass) is widespread and reduces wheat yield due to its severe infestation of wheat fields. Two other allopolyploid species (Ae. geniculata, and Ae. triuncialis) are locally spread, and the others are adventive with a few locations only. Several species are adventive in Canada, in the Canary Islands, in northern and northwestern Europe and in China (van Slageren 1994).

Table 9.2 The occurrence of species of Aegilops in different countries

Most of the species grow in the central part of the genus distribution, i.e., in the Fertile Crescent arc (Israel, Jordan, Lebanon, Syria, southeastern Turkey, northern Iraq, and northwestern Iran) (Table 9.1). Countries like Iran, Iraq, Syria and Turkey contain many (13–17) of the 23 species in the genus, while in peripheral countries, like Afghanistan and Pakistan in the east and those of south-western Europe and North Africa in the west, contain few (2–6) species. The Fertile Crescent arc contains 11–17 species and can be considered as the center of origin and development of most of the species. From this primary center that is characterized by sub-Mediterranean ecological conditions, the various species spread westward to more typical Mediterranean conditions or southward and eastward to steppical areas comprising more extreme environments.

In many parts of the distribution area, the genus has a massive, broad and almost continuous distribution. Species of Aegilops are found in almost every place except for high mountains and deserts (Ae. bicornis, Ae. kotschyi, Ae. longissima, and Ae. crassa even penetrate into semi-deserts areas). All the islands of the Mediterranean Sea are also inhabited by some species. Several species grow only in the Mediterranean region (e.g., Ae. ventricosa, Ae. comosa, Ae. uniaristata, Ae. geniculata, Ae. biuncialis, Ae. sharonensis and Ae. peregrina), others are restricted to the Irano-Turanian region (west Asiatic-central Asiatic regions; Ae. crassa, Ae. kotschyi, Ae. vavilovii, and Ae. juvenalis) while still others (Ae. speltoides, Ae. caudata, Ae. cylindrica, Ae. umbellulata, Ae. triuncialis, and Ae. columnaris) grow in both regions.

The genus has very flexible adaptation capabilities and it occupies a large number of the habitats existing in its distribution area. The altitudinal distribution of the genus is from 400 m below sea level (Dead Sea area) to 2700 m above sea level (asl) with a great variation among species (van Slageren 1994). The climate of many parts of the genus distribution area, especially in the Fertile Crescent arc, where presumably the genus originated and developed, has a short, mild and rainy winter and a long, hot and dry summer. The genus has adapted itself to the conditions characterizing this climate in that all the species are annuals (grow in the winter and pass the dry, hot summer as dormant seeds) and the species are predominantly self-pollinated and have large, well-protected grains for the safe and rapid reestablishment of the stand (Sakamoto 1973; Feldman 1976, 2001). The self-pollination trait enables rapid colonization of newly disturbed habitats as well as maintenance of colonized sites by adaptive genotypes.

All of the species have a, more or less, continuous distribution and usually occupy open habitats in the edges and openings of Mediterranean plant formations, in herbaceous park-forest formations (in which some of the species are natural components), in pastures, abandoned fields, edges of cultivation and roadsides (Zohary and Feldman 1962; Feldman 1963). Some of the habitats are primary in well-defined and balanced ecological conditions, while many are secondary, i.e., in disturbed and degraded areas. Many of the species also grow as weeds in cultivated fields. In disturbed and newly opened habitats, some of the species (particularly the allotetraploids) can form massive and very dense stands, usually consisting of several species. Their genetic system provides them with the ability to colonize such newly opened areas quickly and efficiently.

The diversity in plant habitus and spike morphology enables adaptation of the various Aegilops species to a broad range of habitats. In addition, as has already been pointed out by Stebbins (1956), the adaptive specialization of the various Aegilops species is also reflected in their mode of seed dispersal: they have evolved complex and distinct fruiting spikes, which constitute highly efficient methods of fruit dissemination. Consequently, Aegilops species occupy a variety of primary and secondary habitats.

The distribution of the diploid species is as follows: three species (Ae. speltoides, Ae. caudata and Ae. umbellulata) are distributed in the central part of the genus distribution area. The remaining species of the Sitopsis section (Ae. longissima, Ae. searsii, Ae. sharonensis and Ae. bicornis) grow south of the center, and the species of section Comopyrum (Ae. comosa and Ae. uniaristata) are found west of the center, while Ae. tauschii is in the eastern part of the genus distribution (Table 9.1). Several diploid species have a relatively large distribution area (Ae. umbellulata, Ae. caudata and Ae. tauschii). The distribution of Ae. tauschii is very wide, due to its weediness and segetal growth habit (van Slageren 1994). Another diploid, Ae. speltoides, has a medium-sized distribution area, while others have smaller ones (Ae. bicornis, Ae. searsii, Ae. sharonensis, Ae. longissima, Ae. uniaristata and Ae. comosa). Ae. sharonensis is endemic to the coastal plain of Israel and south Lebanon. The pattern of the geographical distribution of the various diploid species indicates that the genus already underwent extensive differentiation in its early stages of development.

The allotetraploid species have, in general, a broader distribution than the diploids (Zohary and Feldman 1962; Feldman 1963; Kimber and Feldman 1987). Several allotetraploids, Ae. neglecta, Ae. geniculata, Ae. biuncialis, Ae. triuncialis, Ae. cylindrica and Ae. crassa, have a very broad distribution, other allotetraploids, Ae. columnaris, Ae. ventricosa, Ae. peregrina and Ae. kotschyi, have an intermediate distribution, while the hexaploid species, Ae. recta, Ae. vavilovii, Ae. juvenalis, and hexaploid Ae. crassa, have a somewhat more restricted distribution. The polyploids of the U-genome group (section Aegilops) are distributed in the central and western parts of the genus distribution (except for Ae. triuncialis and Ae. columnaris, which extend to the east, and Ae. kotschyi, which is found only in the south-east), those of the D-genome group (section Vertebrata) are in the eastern part (except for Ae. ventricosa, which is found in the western part), and the allotetraploid species of section Cylindropyrum, Ae. cylindrica, is distributed all over the central and northern part (Table 9.1). In most cases, the distribution of the allotetraploid species overlaps, completely or partly, with that of their putative diploid parents. Exception is Ae. ventricosa, which does not overlap with the distribution of either of its parents, the N genome and the D genome donors, namely, Ae. uniaristata and Ae. tauschii, respectively. In some cases, the donor of one of the allotetraploid subgenomes is unknown, e.g., the diploid donor of the X^c subgenome to Ae. crassa, Ae. vavilovii and Ae. juvenalis and the diploid donor of the Xⁿ subgenome to Ae. neglecta, Ae. recta and Ae. columnaris (Dvorak 1998).

The distribution areas of the allotetraploid species of the U-genome group and of Ae. cylindrica from the D-genome group, are larger than those of each of their diploid parents (only that of Ae. columnaris and Ae. peregrina is equal or somewhat smaller than that of Ae. umbellulata). The distribution area of the allotetraploids and allohexaploids of the D-genome group, is smaller than that of Ae. tauschii, the D-genome donor. The distribution of Ae. ventricosa is larger than that of the diploid donor of its second genome, N.

There is a large difference in morphological variation in diploid versus tetraploids. While the diploids have clear-cut boundaries or morphological discontinuities, the tetraploids are characterized by blurred morphological boundaries. This feature of the tetraploid species was already described by Zhukovsky (1928) and Eig (1929a). Eig, in particular, reported on overlapping variation ranges and presence of intermediate linking forms between the various tetraploid species.

There are also striking differences between the patterns of geographical distribution and ecological affinities of the diploids and allotetraploid species (Zohary and Feldman 1962; Feldman 1963). All the diploids are distributed either in or around the center of the genus distribution area (except for Ae. tauschii, which grows in the eastern part). They are relatively restricted in their distribution (Table 9.1) and are much more specialized than the tetraploids in their ecological requirements, usually occupying well-defined habitats with specific edaphic or climatic conditions. Some of the diploids (Ae. tauschii, Ae. umbellulata, Ae. caudata and, to some extent, Ae. speltoides) show wider ecological amplitudes, which correlate with their weedy and segetal tendency.

In contrast, the allotetraploids have a larger distribution area and wider ecological amplitude than the diploids. Ae. triuncialis, Ae. geniculata, Ae. neglecta, Ae. biuncialis and Ae. cylindrica occupy large parts of the distribution area of the genus. The tetraploids do not show the marked ecological specificity of the diploids, as evidenced by their growth in a very wide array of edaphic and climatic conditions. Their weedy nature is reflected in the ability to rapidly and efficiently colonize a variety of newly disturbed and secondary habitats. Undoubtedly, the expansion of agriculture and the opening up of many segetal habitats (in cultivated areas), played a key role in the massive distribution of these tetraploid species throughout the range of the genus.

Many tetraploid species are sympatric and tend to grow in mixed stands, usually with several species in each population (Feldman 1965a). However, in various parts of the genus distribution, one tetraploid is the dominant species. Ae. cylindrica is such a species in the northern part of the genus distribution, Ae. triuncialis in the central and western parts, Ae. peregrina in the southern part, Ae. kotschyi in the southeastern part and Ae. crassa in the eastern part. This interregional kind of vicarism reflects the ability of the various tetraploids to adapt themselves to different climatic conditions.

The diploid species tend to grow in separate habitats, sometimes mixed with tetraploid species. The allotetraploids, in sharp contrast, usually grow intermingled with other tetraploid Aegilops species. In most of the localities studied in Israel, Turkey and Greece, the tetraploid species tend to form mixed populations (Zohary and Feldman 1962; Feldman 1965a). This phenomenon is especially apparent in Turkey, where many tetraploid species occur sympatrically. There, it is possible to find mixed populations which consists of five or even six tetraploid species (Ae. triuncialis, Ae. biuncialis, Ae. neglecta, Ae. columnaris, Ae. geniculate and Ae. cylindrica). In such mixed populations, each species typically exhibits variation in morphological traits and represents by several lines which differ morphologically from one another. The number of distinct morphological lines of each species in a given mixed population is generally related to the number of its individuals, with the most prominent species also tending to be the most variable.

One may assume that the tendency of the allotetraploid species to form polymorphic mixed populations consisting of several species, increases the frequency of genetic contact between them and facilitates interspecific hybridizations and gene flow. Indeed, detailed analyses of several mixed populations demonstrated that the intraspecific variation characterizing each species was partly the result of introgression: some of the lines of each species represented established hybrid derivatives that differed from one another (Zohary and Feldman 1962; Feldman 1965a).

In contrast to the broad distribution of the tetraploid species, the distribution area of the hexaploid species of Aegilops (Ae. vavilovii, Ae. juvenalis, Ae. recta and hexaploid Ae. crassa) is smaller than that of their tetraploid and diploid parents. Only Ae. recta have a larger distribution area than that of its diploid parent, Ae. uniaristata. In addition, the ecological amplitudes of the hexaploids are much more restricted than those of their ancestral tetraploids and even than those of their diploid parents. They grow in a smaller number of habitats and often only sporadically. The morphological variation of the hexaploids is also relatively limited.

According to van Slageren (1994), the genus Aegilops L. is subdivided into the following five sections: Sitopsis (Jaub. & Spach) Zhuk., Vertebrata Zhuk. emend. Kihara, Cylindropyrum (Jaub. & Spach) Zhuk., Comopyrum (Jaub. & Spach) Zhuk., and Aegilops.

9.3 Cytology and Cytogenetics

9.3.1 General Description

The species of Aegilops comprise an allopolyploid series with diploids (2n = 2x = 14), allotetraploids (2n = 4x = 28) and allohexaploids (2n = 6x = 42) (Kihara 1954). Genome size (1C DNA amount) in the diploid species ranges from 4.84 pg in Ae. caudata to 7.52 pg in Ae. sharonensis, in the tetraploid species, it ranges from 9.59 pg in Ae. cylindrica to 12.64 pg in Ae. kotschyi, and in the hexaploid species, from 16.22 pg in Ae. recta to 17.13 pg in Ae. vavilovii (Eilam et al. 2007, 2008; Table 9.3). The karyotype of most species is symmetric, with median or submedian centromeres, with the exception of Ae. caudata, Ae. umbellulata, Ae. comosa, Ae. uniaristata and allopolyploids containing subgenomes that derived from these diploids and have an asymmetric karyotype (Senyaninova-Korchagina 1932; Chennaveeraiah 1960). All species contain two chromosome pairs with a satellite (SAT chromosomes), except for Ae. tauschii and Ae. uniaristata that have only one satellite pair (Chennaveeraiah 1960). Several allopolyploid species contain the sum of the SAT chromosomes of their diploid progenitors, but many more exhibit a smaller number due to amphiplasty, i.e., nucleolar dominance (Chennaveeraiah 1960).

Table 9.3 Nuclear and organellar genome, and genome size of the species of Aegilops

The satellite is a chromosome segment that is separated from the rest of the chromosome by a constriction, called the secondary constriction, whose region is active in nucleolus formation and referred to as nucleolar organizer region (NOR) (McClintock 1934). This NOR contains ribosomal (rDNA) genes that code for the 18S-5.8S-26S (18S-26S). Dubcovsky and Dvorak (1995b) and Badaeva et al. (1996b) presented evidence showing that the major NOR loci are located in homoeologous groups 1, 5, and 6 of the diploid Aegilops species. Using in situ hybridization, additional minor 18S-26S rDNA loci were detected in the genomes of several diploid Aegilops species (Badaeva et al. 1996b).

The 5S rDNA loci have been mapped on chromosomes of the diploid Aegilops species using in situ hybridization (Appels et al. 1980; Castilho and Heslop-Harrison 1995; Friebe et al. 1995c; Badaeva et al. 1996b). Each of the diploid Aegilops species had either one or two 5S rDNA loci on chromosomes of groups 1 and (or) 5 (Badaeva et al. 1996b), either located on the same chromosome arm as the 18S-26S rDNA loci, but unlinked to them, or on different chromosomes (Dvorak et al. 1989; Badaeva et al. 1966b). The variation in chromosomal location and position of major NORs and 5S rDNA loci and the number and distribution of minor NOR loci are characteristic for each diploid species (Badaeva et al. 1996b). The rDNA and the 5S rDNA loci are mobile (Dubcovsky and Dvorak 1995b).

Among the species of section Sitopsis, Aegilops longissima, Ae. sharonensis, Ae. searsii and Ae. bicornis have major NOR loci on chromosomes of groups 5 and 6 and a variable number of minor loci on chromosomes of groups 1, 3, 5, and 6 (Badaeva et al. 1996b). The 5S rDNA loci were observed on chromosomes of groups 1 and 5. On the other hand, Ae. speltoides have a different distribution pattern of NOR and 5S rDNA loci, similar to that of Amblyopyrum muticum (Badaeva et al. 1996b). The major NOR loci are located on chromosomes of groups 1 and 6 and only one 5S rDNA locus was found on the short arm of chromosomes of group 5. Likewise, the distribution of major NOR and 5S rDNA loci in Ae. comosa was similar to that in Ae. speltoides (and in A. muticum), except that minor NOR loci were observed in all seven chromosome pairs. The distribution patterns of NOR and 5S rDNA loci in Ae. umbellulata and Ae. caudata were identical; both loci were located on the short arm of chromosomes of groups 1 and 5 (Gerlach et al. 1980; Miller et al. 1983). However, an additional pair of NOR sites was observed in Ae. umbellulata (Badaeva et al. 1996b).

The distribution of NOR and 5S rDNA loci in Ae. uniaristata and Ae. tauschii is distinct, with only one major NOR locus in the short arm of chromosome 5N and 5D, respectively, and several minor NORs on the short and long arms of chromosomes 1, 6 and 7 in Ae. uniaristata, and on the long arm of chromosome 7 in Ae. tauschii (Badaeva et al. 1996b). Two 5S rDNA loci exist in both species—one in the long arm of chromosomes of group 1 and the other in the short arm of chromosomes of group 5 of Ae. uniaristata. In Ae. tauschii, these two loci are located on the short arm of chromosomes of groups 1 and 5. This mode of distribution of the two 5S rDNA loci is similar in most other diploid Aegilops species (Badaeva et al. 1996b).

In many Aegilops allopolyploid species, the rDNA genes of one parental set are transcribed, while most or all of the rDNA genes inherited from the other parent are silent or absent, a phenomenon known as a nucleolar dominance (Navashin 1928, 1934; Pikaard 1999, 2000). Nucleolar dominance occurs in almost all allopolyploid species of Aegilops (Cermeño et al. 1984b; Feldman et al. 2012). In the allopolyploid species containing the U subgenome of Ae. umbellulata, the U genome completely suppresses the NOR activity of the M, S and D subgenomes of the allopolyploids (Cermeño et al. 1984b).

Since most chromosomes of Aegilops species are morphologically indistinguishable from one another at mitotic metaphase, several methods were developed to identify individual chromosomes, for studying chromosome structure and organization and for genome analysis. Two widely used methods are C-banding and fluorescence in situ hybridization (FISH). The C-banding technique detects heterochromatic regions of the chromosomes, whose distribution can be chromosome- and species-specific. The C-banding patterns of chromosomes at mitotic metaphase were studied in all diploids (Teho and Hutchinson 1983; Teho et al. 1983; Friebe et al. 1992a, 1993, 1995a; Badaeva et al. 1996a, b) and polyploid species (Badaeva et al. 2002, 2004, 2011) of Aegilops. Chromosomes of all species show a distinctive and characteristic C-banding pattern, enabling the identification of their individual chromosomes. The results of the above-mentioned studies indicated that the total amount and the type of distribution of the heterochromatic regions in chromosomes were species-specific. In addition to differences in chromosome morphology (Chennaveeraiah 1960), karyotypes of all species could be distinguished by the distribution of heterochromatic regions in all their chromosomes.

The FISH method exploits repetitive DNA sequences, e.g., probe pSc119 isolated by Bedbrook et al. (1980) from the genome of Secale cereale, or probe pAs1 isolated by Rayburn and Gill (1986) from Aegilops tauschii. This method facilitates the precise location of such sequences on chromosomes may be chromosome-, species-, and probe-specific (Jiang and Gill 2006). A comparative analysis of in situ hybridization with the highly repetitive DNA sequences pSc119 and pAs1 in all the diploid Aegilops species confirmed significant differentiation of their genomes (Badaeva et al. 1996a). In addition to interspecific differences, significant intraspecific polymorphism was also detected in the distribution of repetitive DNA sequences (Badaeva et al. 1996a).

9.3.2 Structure and Distribution of Repetitious DNA

The genomes of diploid and allopolyploid Aegilops species are very large (Eilam et al. 2007, 2008; Table 9.3), and are comprised of about 85–90% repeated nucleotide sequences (Flavell et al. 1979), most of them being transposable elements (TEs), primarily families of retrotransposons (Li et al. 2004; Wicker and Buell 2009; Yaakov et al. 2013; Senerchia et al. 2013; Jia et al. 2013). TEs have the potential to affect genome structure, function and size through transposition (Bennetzen 2005; Slotkin and Martienssen 2007; Fedoroff 2012) and, so, differential proliferation of TEs is considered to be one of the main driving forces of genome size variation in the Triticeae (Charles et al. 2008). It is therefore likely that the large differences in genome size between the various Aegilops species (Eilam et al. 2007, 2008; Table 9.3), derived from differential proliferation of TEs that were active during the speciation processes of these species and that they played an important role in their genomic evolution (Yaakov et al. 2013).

Middleton et al. (2013) found that the abundance of several TE families significantly differs between the Triticeae species, indicating that TE families can thrive extremely successfully in one species while going virtually extinct in another. In this regard, Senerchia et al. (2013) found that ancestral TE families followed independent evolutionary trajectories in several Aegilops species, highlighting the evolution of TE populations as a key factor of genome differentiation. Already in 1979, Flavell et al. (1979) showed that DNA of different Aegilops species hybridized to differing extents with a repetitive probe that derived from Ae. speltoides. These results are consistent with the hypothesis that speciation has been accompanied by quantitative changes in the repeated sequence complements of genomes (Flavell et al. 1979).

Badaeva et al. (1996a), using in situ hybridization with two highly repetitive DNA sequences, pSc119 from Secale cereale (Bedbrook et al. 1980) and pAs1 from Ae. tauschii (Rayburn and Gill 1986), studied genome differentiation in all diploid Aegilops species. While chromosomes of all the diploid species hybridized with the pSc119 probe, the level of hybridization and labeling patterns differed among genomes. Only three species, Ae. tauschii, Ae. comosa, and Ae. uniaristata, showed distinct hybridization with pAs1. The labeling patterns were species-and chromosome-specific, confirming significant differentiation of their genomes.

Similar conclusion was reached by Yaakov et al. (2013), who assessed the relative copy number of 16 TE families in Aegilops species of section Sitopsis and in Ae. tauschii. They reported on a wide variation and genome-specificity of TEs in these species. Likewise, Senerchia et al. (2013) investigated genome restructuring and assessed the evolutionary trajectories of 17 long-terminal repeat (LTR) retrotransposon families after allopolyploidization events. Comparisons between these retrotransposons of the diploid progenitors and the allopolyploids highlighted the proliferation of several TE families and the predominant sequence deletion in others, indicating species-specific and TE-specific evolutionary trajectories following allopolyploidy.

9.3.3 Gametocidal Chromosomes in the Genus Aegilops

9.3.3.1 Opening Remarks

During recurrent backcrossing to produce addition lines of chromosomes from several Aegilops species to common wheat and during recurrent backcrossing to produce alloplasmic lines of wheat containing cytoplasm of Aegilops species substituting the wheat cytoplasm, it was found that certain alien chromosomes from a number of Aegilops species were preferentially transmitted to the offspring (Endo 2007, 2015). When introduced in a single dose to durum or common wheat, as in F₁ hybrids, in backcrossed progeny to wheat, in monosomic addition or monosomic substitution, these chromosomes, ensured their endurance in wheat by inducing severe chromosomal breakage in gametes lacking them, thus, causing their abortion and consequently, leading to their preferential transmission to the offspring of gametes possessing the gametocidal chromosome (Endo 1982, 1985, 1990; Maan 1975; Finch et al. 1984). In consequence, a severe reduction in the fertility of both sexes in wheat plants having a monosomic addition or substitution of one of the gametocidal chromosomes was observed (Endo 1985). Self-pollination of lines carrying such a chromosome yields offspring predominantly bearing a disomic addition or disomic substitution, and restored fertility. These Aegilops chromosomes are termed gametocidal (Gc) chromosomes (Endo 1979, 1982, 2007) or “cuckoo” chromosomes (Miller et al. 1982), and the genes that are responsible for the gametocidal action are called Gc genes (Endo 1982). Using FISH with a probe of a repetitive DNA sequence that marks the Gametocidal (Gc) gene, Friebe et al. (2003) directly demonstrated that chromosome breakage in pollen mitosis occurred only in gametes lacking this gene. The Gc chromosomes derived from different Aegilops genomes (C, S, S^sh, S^l and M^o) and belong to four different homoeologous groups: 2, 3, 4, and 6 (Endo 2007, 2015). Currently known species possessing gametocidal chromosomes are: Ae. sharonensis (genome S^shS^sh) (Maan 1975; Miller et al. 1982; Tsunewaki and Tsujimoto 1983; Tsujimoto and Tsunewaki 1985b; Endo 1990, 2007, 2015), Ae. longissima (genome S^lS^l) (Maan 1975; Endo 1990), Ae. speltoides (genome SS) (Tsujimoto and Tsunewaki 1984, 1988), Ae. caudata (genome CC) (Endo and Katayama 1978; Endo 1985), Ae. triuncialis (genome CCUU) (Endo and Tsunewaki 1975), Ae. cylindrica (genome CCDD) (Endo 1979) and Ae. geniculata (genome UUM^oM^o) (Friebe et al. 1999).

The Ae. sharonensis and Ae. longissima Gc chromosomes were first discovered by Maan (1975). Three such chromosomes were identified in different Ae. sharonensis accessions, (1) by Endo 1982, (2) by Miller et al. 1982, and (3) by Maan 1975, and two such chromosomes in different Ae. longissima accessions, (1) by Maan 1975, and (2) by Panayotov, cited in Endo (1985). The cytological features, homoeology and interrelation of the different Gc chromosomes were studied (Endo 1985). One of the sharonensis Gc chromosomes (no. 1) and one of the longissima Gc chromosomes (no. 2) have the same gametocidal action, and both have an N-banding pattern that resembles that of wheat chromosome 2B, and thus, are homoeologous to wheat group 2, and successfully substitute for any wheat chromosome of wheat homoeologous group 2. The other two sharonensis Gc chromosomes (nos. 2 and 3) and the longissima Gc chromosome (no. 1) are homoeologous to wheat group 4 (Miller et al. 1982), and exhibit relatively similar N-banding patterns. The group 4 chromosomes showing the same gametocidal action, all showed an N-banding pattern rather similar to that of wheat chromosome 4A. Dvorak (1983) presented the similarity between the C-banding pattern of the sharonensis Gc chromosome (no. 2) and wheat chromosome 4A, as evidence supporting his view that chromosome 4A of T. aestivum was contributed by a species of the section Sitopsis and, consequently, belongs to the B genome. The gametocidal chromosomes of group 2 were designated Gc1 and those of group 4 designated Gc2 (Endo 1985, 2007).

Two Gc genes, derived from two different strains, were found in Ae. speltoides (Tsujimoto and Tsunewaki 1984, 1988). These two genes are allelic, located on the Gc chromosome 2S, which is homoeologous to wheat group 2, and consequently, designated Gc1a and Gc1b (Tsujimoto and Tsunewaki 1984, 1988). The two alleles when present in monosomic addition of the Gc chromosome, differ in their ability to induce damage to the offspring of plants lacking the Gc chromosome; Gcla causes endosperm degeneration and chromosome aberrations, whereas Gclb results in abnormal seed lacking the shoot primordium. No correlation between embryo or endosperm degeneration and chromosome breakage was observed (Tsujimoto and Tsunewaki 1984, 1988).

The Gc chromosomes of Ae. caudata and Ae. triuncialis are homoeologous to wheat chromosomes of group 3 (Endo and Tsunewaki 1975). The morphology, pairing homology, selective gametocidality, and effects on plant growth of gametocidal chromosomes of natural and of synthetic Ae. triuncialis are almost the same as those of Ae. caudata (Endo 1979). On the other hand, the Ae. cylindrica Gc chromosome is homoeologous to group 2 (Endo 1979). It differs from the Gc chromosomes of Ae. triuncialis and Ae. caudata in its characteristic appearance and many aborted seeds. In addition, the centromere of the cylindrica Gc chromosome is not so extremely subterminal as that of the caudata and triuncialis Gc chromosome, and the selective gametocidal action of the cylindrica Gc chromosome is not effective in the types of common wheat where the caudata and triuncialis chromosomes exert their gametocidal effect. In respect of the selective gametocidal chromosome, therefore, the C genome of Ae. cylindrica is farther differentiated than that of Ae. caudata and of Ae. triuncialis.

Chromosome 4M^o, which is homoeologous to wheat group 4 chromosomes, is the Gc chromosome of Ae. geniculata (Kynast et al. 2000). When transferred to cv. Chinese Spring of bread wheat as a monosomic addition, it induces chromosome breakage and anaphase bridges at anaphase and telophase of the first and second pollen mitosis. Gc-induced multicentric and ring chromosomes, among other chromosomal aberrations, can be transmitted to the offspring and initiate breakage-fusion-bridge cycles in dividing root tip meristem cells of the derived sporophytes.

9.3.3.2 Interaction Between Gametocidal (Gc) Genes

In double monosomic additions of common wheat with the gametocidal chromosomes of Ae. sharonensis, 2S^sh and 4S^sh, only gametes carrying the alien chromosome 4S^sh were functional. Hence, there are two types of gametocidal chromosomes in Ae. sharonensis, with the 2S^sh chromosome being weaker than the 4S^sh (Endo 1985).

When observing double monosomic addition lines derived from three different Gc chromosomes, Endo (1982) found that the Gc gene of Ae. triuncialis does not interact with the Gc gene of Ae. longissima or Ae. sharonensis. In addition, he found that the activity of the Gc gene of Ae. longissima dominated over that of Ae. sharonensis. Endo (1985) further reported that Gc genes located on chromosome 4S^l of Ae. longissima or 4^sh of Ae. sharonensis are epistatic to those on chromosome 2S^l, irrespective of species. Hence, the Gc chromosomes derived from Ae. triuncialis, Ae. sharonensis and Ae. longissima were found to differ in gametocidal action, as well as in morphology and homoeology (Endo 1982). The mode of action of the Gc genes of Ae. longissima, Ae. sharonensis and Ae. speltoides differs from that of Gc genes in Ae. triuncialis and Ae. cylindrica (Tsujimoto and Tsunewaki 1985b; Tsujimoto and Noda 1989; Endo 1988b).

Using plants with two different Gc genes, Tsujimoto (1995) investigated the functional relationship between six Gc genes and concluded that there are three functional groups. The first group included Gc genes located on the chromosomes of homoeologous group 2; the Gc1 genes of Ae. speltoides showed similar function to those on chromosome 2S^sh of Ae. sharonensis. The second group included the Gc genes on chromosomes 4S^sh of Ae. sharonensis and 4S^l of Ae. longissima, i.e., the Gc2 genes. These genes were epistatic to the Gc1 genes in the first group in terms of gamete abortion and preferential transmission, as indicated by Endo (1985). Although, by themselves, the Gc genes in the first group only cause chromosome breakage at a low frequency (Tsujimoto and Tsunewaki 1985b; Tsujimoto and Noda 1989), they highly enhance breakage by Gc genes of the second group. Conversely, the Gc genes in the second group may enhance breakage induced by those in the first group.

The third group included the Gc gene on chromosome 3C of Ae. triuncialis, proved to have activity independent of that of the Gc genes of the first or second group. The function of the triuncialis Gc gene is suppressed by an inhibitor, Igc1, located on chromosome 3B of some common wheat lines. Based on the interactions between the different Gc genes, Tsujimoto (1995) proposed re-designation of the gene symbols following the rules for gene symbolization in wheat (McIntosh 1988). Tsujimoto (1995) proposed the name Gc1 for the Gc genes in the first group, Gc2 for the Gc genes in the second group, and Gc3 for the Gc genes in the third group, with each designation followed by the name of the genome carrying the gene. The relationship between these Gc genes and those on chromosome 2C of Ae cylindrica, chromosome 4M^o of Ae. geniculata and chromosome 6S of Ae. speltoides has not yet been examined.

9.3.3.3 Mode of Action of Gc Genes

The explanation to the phenomenon of differential transmission of Gc chromosomes in monosomic additions or disomic substitutions of Aegilops Gc chromosomes to common wheat, is that meiospores (microspores and megaspores) with the alien chromosome develop into normal gametophytes, while meiospores lacking the alien chromosome exhibit a wide range of chromosome and chromatid aberrations at first gametophytic mitosis (Endo 2015, and reference therein). This would explain the partial male and female sterility in the monosomic addition or substitution lines. Hence, Gc genes have a dual function, i.e., to induce chromosomal mutations in gametes that lack them and to suppress such mutations in gametes that carry them. Indeed, Endo (1990) and Tsujimoto (2006) hypothesized that two genetic factors are associated with the preferential transmission of the Gc chromosome, the breaker (GcB) that induces chromosome breakage, and the inhibitor that prevents chromosome breakage. Chromosome aberrations do not occur in gametes carrying both elements, as the inhibitor neutralizes the gametocidal action. This hypothesis is supported by the discovery of, Igc1, an inhibitor of the Gc of Ae. cylindrica on 2C (Tsujimoto and Tsunewaki 1985a), and by the isolation of a knockout mutation of the breaker gene of the Gc of Ae. sharonensis on 4S^sh, which renders the breakage function ineffective, while having no influence on the latter inhibition function (Friebe et al. 2003). The Ae. sharonensis 4S^sh breaker element (Gc1B) has been mapped, by C-banding, to the distal end of the long arm of chromosome 4S^sh (Endo 2007). Knight et al. (2015) confirmed this reported location and more specifically defined its location in a region proximal to the sub-telomeric heterochromatin of this chromosome arm. However, the molecular mechanism of the effect of the GcB has not yet been elucidated (Tsujimoto 2006).

It is not known if Gc genes are functional in their species of origin, i.e., in intraspecific hybridization between lines bearing and those lacking Gc chromosomes, nor in interspecific hybrids between various diploid and allopolyploid Aegilops species bearing with those lacking Gc chromosomes. Also, it is not known if some lines of diploid Aegilops and Triticum species possess a gene(s) that suppresses the function of the GcB gene.

9.3.3.4 Modification of Gc Action

The Gc genes action varies, depending on the common wheat cultivar into which a GC chromosome is introduced (Endo 1988b). Chromosome 2C of Ae. cylindrica, for instance, has complete Gc action and is therefore exclusively transmitted to progeny in the common wheat cultivar Jones Fife (JF), whereas its Gc action becomes incomplete in the common wheat cultivar Chinese Spring (CS), where chromosome 2C is lost in part of the progeny (Endo 1988a). Chromosome 3C of Ae. triuncialis has severe Gc action in CS and some other common wheat cultivars, but it displays almost no Gc action in Norin 26 (N26), which possesses the Igc1 Gc-inhibitor gene on chromosome 3B (Tsujimoto and Tsunewaki 1985a). In both cases of incomplete Gc action, semi-lethal chromosomal mutations occur in gametes lacking the Gc chromosome, and structurally rearranged chromosomes are transmitted to the progeny.

Endo (1978) described evidence for the existence of suppressor(s) of Gc genes in certain cultivars of common wheat. He reported that monosomic addition of Gc chromosome 3C of Ae. triuncialis correlated with male and female semi-sterility in the genetic backgrounds of the common wheat cultivars JF and CS, whereas semi-sterility did not appear in the background of cultivar N26. Chromosome 3C was preferentially transmitted to the next generation of both sides in JF but only of the female side in CS. Although Endo (1978) did not mention preferential transmission of Gc chromosome 3C in N26, recovery of fertility in the background of this cultivar indicated that gametes lacking chromosome 3C were also normally transmitted.

Tsujimoto and Tsunewaki (1985a) analyzed the genetic factor in N26 that suppresses the Gc action of Ae. triuncialis chromosome 3C. They crossed the disomic addition line of Chinese Spring carrying chromosome 3C (21ʺw + 1ʺae) with the F₁ progeny of the hybrid Chinese Spring × N26. In the resultant monosomic addition lines, fertile and semi-sterile plants segregated 1:1, indicating that a dominant suppressor gene, termed Igc1, inhibits the action of the Gc gene on chromosome 3C. By monosomic analysis Tsujimoto and Tsunewaki (1985a) localized Igc1 to chromosome 3B of N26. The facts that both the Gc1 gene and its suppressor are located on chromosomes of the same homoeologous group, may indicate relationships between these two genetic factors, i.e., that Igc1 is an antimorph allele of a Gc gene, acting antagonistically to the triuncialis Gc1 gene. Since Igc1 exists only in a number of common wheat cultivars (see below), it is reasonable to assume that it did not derive from the diploid donor of the B genome, but rather, evolved at the polyploid level to counteract the action of Gc genes.

In the JF genetic background, both male and female gametes without chromosome 3C were abortive, whereas in the CS background, pollen without the Gc chromosome functioned and transmitted to the progeny (Tsujimoto and Tsunewaki 1985a). This result suggests the existence of an incomplete suppression in the CS background. In addition, no suppressors for the Ae. longissima, Ae. sharonensis, or Ae. speltoides Gc chromosome actions were discovered among the hundreds of common wheat cultivars tested so far (Tsujimoto, unpublished, cited in Tsujimoto and Tsunewaki 1985a).

Tsujimoto and Tsunewaki (1985a) and Tsujimoto and Tsunewaki (1988) studied the distribution of the inhibitor gene, Igc1, by crossing many cultivars of common wheat with CS following a disomic addition of Ae. triuncialis chromosome 3C, and classifying them into Igc1 carriers and non-carriers, based on the seed fertility of the F₁ progeny. They found that the Igc1 gene exists in Japanese and East and Southwest China cultivars but not in American, African, and Asiatic cultivars. Interestingly, cultivar CS, originally from the Sichuan Basin in China (Yen et al. 1988), is an Igc1 non-carrier in spite of the fact that most Sichuan landraces are carriers. The Japanese cultivar Norin 10, which was used as the source of the semi-dwarf genes Rht1 and Rht2 in modern wheat breeding, carries Igc1, but the suppressor gene did not pass into the high-yielding cultivars containing these dwarfing genes.

Tsujimoto and Tsunewaki (1985b) made note of the fact that the phenomena associated with Gc genes in wheat are similar to those observed in association with hybrid dysgenesis in the fruit fly, Drosophila. These include sterility, lethality, mutation, chromosome breakage, male recombination and segregation distortion, all of which appeared only in the F₁ progeny of a cross between P or I lines of Drosophila males and M or R strain females (Crow 1983; Bregliano and Kidwell 1983). Later, Tsujimoto and Noda (1989) noted the similarity between the nature of Gc genes and the restriction-modification systems found in many bacteria. In bacteria, a restriction endonuclease in the host cuts alien DNA at or around a particular base sequence. The host DNA, by contrast, is protected from digestion by methylation. This restriction-modification system provides a mechanism that could explain chromosome breakage in gametogenesis and in zygotic cells in wheat (Tsujimoto and Tsunewaki 1985b). Tsujimoto and Tsunewaki (1985b) proposed a model for Gc action in which a Gc gene produces both a restriction enzyme (RE) and a modification enzyme (ME), e.g., a methylase. The RE cleaves the specific restriction sites that it recognizes. But, when these sites are protected by DNA methylation, caused by the ME, the RE cannot cleave them. This would be the case in homozygotes for the Gc gene, where no chromosome breakage occurs in any gametes. If ME function is incomplete and cannot protect all of the restriction sites, chromosome breakage may appear at some frequency in all gametes, regardless of presence of the Gc genes.

In plants hemizygotic for a Gc gene, haploid cells without the Gc gene are generated after meiosis. Since DNA replication occurs prior to the first mitotic division in the gametogenesis, cells lacking the Gc gene and therefore, the ME, will not contain modified restriction sites on one of the strands of the replicated DNA. If the RE remains in the cell longer than the ME, or if RE can be supplied by other cells (for example, egg or pollen mother cells), the unmodified restriction sites are broken by the RE. In the following mitosis, unmodified DNA is broken in the same manner. Thus, the gametes without Gc become abortive.

Interestingly, de las Heras et al. (2001) observed that treatment of plants carrying the Ae. sharonensis Gc gene, with the hypomethylation agent 5-azacytidine, induced chromosome breakage in root tip cells. This result supports that the process of chromosome breakage in early seed development was repressed by DNA methylation.

9.3.3.5 Evolutionary Significance of Gc Genes

At the diploid level, Gc genes can restrict intraspecific gene exchange between two different populations that possess non-compensating Gc genes (Endo 2015). If these Gc genes are alleles, located on homologous chromosomes, the intraspecific hybrid between these two populations will be completely sterile, as all gametes will include one of these incompatible Gc alleles. If the two different Gc genes are on non-homologous chromosomes, one-fourth of the gametes produced by the hybrid will be fertile. Thus, sexual isolation will be established within a species between two populations that easily cross-fertilize. As a result, the two populations can develop independent of one another and gradually diverge to two different species. Similarly, two closely related species can be sexually isolated and undergo independent evolutionary development.

An example of sexual isolation within a species is seen in hybrids between two allopatric accessions of Ae. caudata, which have normal meiotic chromosomal pairing, but produce completely sterile pollen (Ohta 1992). This sterility might be explained as the result of the occurrence of two different alleles at the Gc loci on homologous chromosomes of the allopatric accessions.

While all studied accessions of Ae. sharonensis possess gametocidal chromosomes, several lines of Ae. longissima do not and, therefore, addition and substitution lines of chromosomes of these lines were produced in common wheat. Feldman (1975) used Ae. longissima line TL01, from Revivim, central Negev, Israel, as a source for successful production of six different addition lines to common wheat cv. CS. A second complete set of disomic chromosome addition lines was successfully obtained by NA Tuleen (described in Friebe et al. 1993), by crossing Ae. longissima line 4 (an accession collected by G. Hart in Israel, and which is different from TL01), with common wheat cv. CS. Thus, two different lines of Ae. longissima do not possess gametocidal chromosomes. Several lines of Ae. speltoides also lack a gametocidal chromosome (Tsujimoto and Tsunewaki 1984), as also evident by the successful production of a complete series of seven alien addition lines with a low-pairing accession (Friebe et al. 2000).

To date, gametocidal chromosomes have not been found, neither in common wheat nor in the allotetraploids Ae. peregrina and Ae. kotschyi, which contain the S^l subgenome that derived from Ae. longissima. This can be the result of elimination or suppression of the gene(s) causing the gametocidal action during the evolution of the allopolyploid species. Alternatively, these genomes derived from accessions lacking gametocidal chromosomes. The Gc genes of the C and M^o genomes in the allopolyploids Ae. cylindrica, Ae. triuncialis and Ae. geniculata, respectively, induce mild, or semi-lethal, chromosome mutations in alien addition lines of common wheat (Endo 2007). The weak or complete absence of Gc gene activity in allopolyploid species, in contrast to their strong effect in diploid species, suggests that allowance of interspecific hybridization and gene exchange in allopolyploids has a great evolutionary advantage (Zohary and Feldman 1962). This is in contrast to the situation at the diploid level, where restriction of interspecific gene exchange provides an evolutionary advantage.

The presence of incomplete Gc action of some gametocidal genes suggests that the Gc system might also be involved in the evolution of the karyotype in the genus Aegilops (Endo 2015). Incomplete Gc action induces chromosomal rearrangements in hybrids heterozygous for a Gc gene, and gametes with rearranged chromosomes will survive and self-fertilize. The karyotype of the selfed progeny will stabilize when the Gc gene becomes homozygous, and some well-balanced karyotypes might be established in separate populations.

9.3.3.6 Use of Gc Genes in the Production of Deletion and Dissection Lines

The Gc genes of the C and M^o subgenomes in from allotetraploid Aegilops induce only mild, or semi-lethal, chromosome mutations in alien addition lines of common wheat. Consequently, induced chromosomal rearrangements have been identified and established in wheat stocks carrying deletions of wheat and alien chromosomes or wheat-alien translocations. Thus, gametocidal chromosomes may serve as a tool to produce cytogenetic stocks for cytogenetic manipulations (Endo 2007).

Monosomic addition of Ae. cylindrica chromosome 2C showed preferential transmission of 2C in the background of the common wheat cultivar JF but not in the background of CS (Endo 1979, 1988a). The disappearance of Gc action in CS is similar to the case of the Ae. triuncialis chromosome 3C in the background of cultivar N26 (Tsujimoto and Tsunewaki 1985a). Chromosome aberrations caused by chromosome 2C appeared most often in offspring without the alien chromosome. Endo (1988a) suggested that when the gametocidal action is mild, gametophytes without the alien chromosome are fertilized, suffer slight chromosome damage, and develop into plants with chromosome aberrations.

Tsujimoto et al. (1990) observed chromosome fragments, bridges and micronuclei in the first and the second pollen mitoses of monosomic 3C addition to CS and Nasuda et al. (1998) observed similar chromosome aberrations in monosomic 2C addition to CS. The breakpoints of the chromosome aberrations do not appear to be distributed randomly in wheat chromosomes and may be restricted to specific chromosome structures or DNA sequences (Endo and Gill 1996). In their effort to produce a large-scale collection of 1B deletion chromosomes, Tsujimoto et al. (2001) recognized breakage ‘hot spots’. However, the distribution pattern of the breakage hot spots in the studies of Endo and Gill (1996) and Tsujimoto et al. (2001) did not coincide with each other, despite the fact that both studies used the same Gc gene (Tsujimoto et al. 2001; Friebe et al. 2001). The broken end gradually acquired repetitive telomere sequences, indicating that incomplete (or perhaps undetected) telomere sequences were sufficient to heal the broken ends (Tsujimoto 1993). Using the telomere sequence as a primer for PCR, the DNA sequences at the broken ends were amplified and then analyzed. However, no specific sequences were identified (Tsujimoto et al. 1997, 1999).

The abnormal chromosomes induced by the Gc gene of Ae. triuncialis or Ae. cylindrica can be transmitted to the next generation. Because the breakage only occurs in the gametes without the Gc chromosome in monosomic addition lines, the offspring with a chromosome deletion in the next generation were stable and did not induce additional chromosome aberrations. Thus, these deletion lines were useful for mapping and were maintained as the standard for mapping genes to specific chromosome regions in common wheat (Endo and Gill 1996; Tsujimoto et al. 2001).

Using mostly chromosome 2C of Ae. cylindrica, Endo and Gill (1996) produced approximately 350 homozygous deletion lines of CS wheat that contain deletions of various size in specific chromosomes. These lines are useful in cytologic mapping (deletion mapping) of genes and especially of DNA markers to the missing chromosomal regions (Werner et al. 1992; Qi et al. 2004). Most of the CS deletion lines, together with the Gc chromosomes, are available at NBRP-wheat website (http://www.shigen.nig.sc.jp/wheat/komugi/strains/aboutNbrpL.gku.jsp).

In addition, the Gc genes can be usefully applied to induce translocations between alien chromosomes introduced into common wheat and wheat chromosomes. This is of particular relevance in the case of alien chromosomes from species distantly related to wheat, and which show little tendency to undergo homoeologous recombination with wheat chromosomes, even under genetically permissive conditions (Endo 2015). As an example, the Ae. cylindrica Gc chromosome 2C was introduced into CS wheat having all barley chromosomes as disomic addition lines, except for 1H (Shi and Endo 1997). Chromosomal rearrangements were induced by the 2C gametocidal system for each barley chromosome, including 2H (Joshi et al. 2011), 3H (Sakai et al. 2009), 4H (Sakata et al. 2010), 5H (Ashida et al. 2007), 6H (Ishihara et al. 2014) and 7H (Schubert et al. 1998; Serizawa et al. 2001; Masoudi-Nejad et al. 2005; Nasuda et al. 2005). The Gc system was similarly proven to be effective in inducing structural rearrangements in rye chromosome 1R introduced into common wheat (Endo et al. 1994; Masoudi-Nejad et al. 2002; Gyawali et al. 2009, 2010; Li et al. 2013). Since both terminal deletions and wheat-alien translocations enable cytological mapping of alien chromosomes, Endo (2015) has been developing many common wheat lines carrying deletions and translocations of alien chromosomes, collectively named “dissection lines”. Comparative studies of cytological and genetic maps obtained in the above studies revealed that crossing-over is generally more frequent in the distal region than in the proximal region for all the wheat, barley and rye chromosomes that were analyzed.

9.4 Section Sitopsis (Jaub. & Spach) Zhuk.

9.4.1 General Description

Section Sitopsis (Jaub. & Spach) Zhuk. [Syn.: subgen. Sitopsis Jaub. & Spach; sect. Platystchys Eig; Triticum L. sect. Sitopsis (Jaub. & Spach) Chennav.; Sitopsis (Jaub. & Spach) Á. Löve] is characterized by long spikes, of lengths that are at least 20 times their width, with either two-rowed, lemmas of lateral spikelets are awned and disarticulating wedge-type (Ae. speltoides var. ligustica, Ae. bicornis, Ae. sharonensis), or with one-rowed, only lemmas of apical spikelets with awns, disarticulating as one unit, umbrella type (Ae. speltoides var. speltoides and Ae. searsii disarticulate above a basal rudimentary spikelet and Ae. longissima above several fertile spikelets that remain on the culm). Caryopsis is adherent to lemma and palea or free (in Ae. searsii) (Fig. 9.1).

Fig. 9.1

Aegilops species of section Sitopsis; a A spike of Ae. speltoides Tausch var. ligustica (Savign.) Fiori in Fiori & Paoletii; b A spike of Ae. speltoides var. speltoides Tausch; c A spike of Ae. bicornis (Forssk.) Jaub. & Spach; d A spike of Ae. sharonensis Eig; e A plant and a spike of Ae. longissima Schweinf. & Muschl.; f A plant and a spike of Ae. searsii Feldman and Kislev ex Hammer

The Sitopsis species can be distinguished by their morphology, and more easily by their specific habitat, climatic adaptation or area of distribution. Zhukovsky (1928) included four species in section Sitopsis, while Eig (1929a) recognized five, grouping them in two subsections, Truncata and Emarginata. Subsection Truncata, containing one species, Ae. speltoides Taush (Table 9.1), is characterized by many-flowered spikelets and relatively short glumes (about half the length of the florets), terminating in a thick margin, with or without a small tooth on one side. Species of this subsection are found in the central, eastern and northern parts of the Sitopsis distribution area and grow on heavy and moist soils. According to Eig (1929a), subsection Truncata includes two species, namely, Ae. speltoides Tausch and Ae. ligustica (Savigny) Cosson. Yet, Sears (1941b) and Kihara (1954), based on cytogenetic studies, concluded that Ae. ligustica should be included in Ae. speltoides. Zohary and Imber (1963) supported this conclusion by presenting genetic evidence indicating that Ae. speltoides and Ae. ligustica are two genetic forms of the same species. Molecular analysis by Goryunova et al. (2008) also supported this conclusion.

Subsection Emarginata is characterized by few-flowered spikelets and relatively longer glumes than those of subsection Truncata, about 2/3 or more the length of the florets, terminating generally in two teeth (occasionally 0–3), separated by an angle. The species of this subsection are found in the central and southern parts of the distribution area of the section, on light, sandy soils. Subsection Emarginata includes four species: Ae. sharonensis Eig, Ae. longissima Schweinf. et Muschl., Ae. bicornis (forssk.) Jaub. et Sp., and Ae. searsii Feldman and Kislev ex Hammer (Table 9.1). Based on chromosome pairing at meiosis of F₁ hybrids between Ae. sharonensis and Ae. longissima, Kihara (1954) suggested including Ae. sharonensis in Ae. longissima. This suggestion was accepted by several taxonomists and cytogeneticists (e.g., Bowden 1959; Morris and Sears 1967; Mackey 1968). However, Ankori and Zohary (1962) and Waines and Johnson (1972) were inclined to accept Eig’s recognition of Ae. sharonensis as a separate species. Later on, taxonomists (e. g., Hammer 1980; van Slageren 1994) and cytogeneticists (e. g., Teoh and Hutchinson 1983; Yen and Kimber 1990b) presented evidence justifying the specific rank of Ae. sharonensis.

Aegilops searsii was described by Feldman and Kislev (1977) as a taxon with unique habit and habitats, possessing sufficient new characteristics to justify treating it as an independent species. This new species, that was included in subsection Emarginata, differs from the other members of this subsection in morphological, eco-geographical and karyotypic characteristics.

Mendlinger and Zohary (1995) assessed the extent and structure of genetic variation in 21 populations covering the five species of the Sitopsis section, by electrophoretically analyzing water-soluble leaf proteins. All loci were polymorphic across the five species. Over 40% of the alleles were found in all five species and only three rare alleles were species-specific. Genetic diversity was high (D = 0.267), with 51% of the total diversity contributed by within-population diversity, 16% by diversity between populations within a species and 33% by diversity between species. Ae. speltoides was genetically distant from the other four species. Ae. sharonensis was found to be equally close to Ae. longissima and Ae. bicornis, whereas Ae. searsii was equally distant from Ae. longissima, Ae. sharonensis and Ae. bicornis.

Similar results were obtained by Giorgi et al. (2002) who used restriction-fragment-length polymorphism (RFLP) analysis to investigate phylogenetic relationships among the Sitopsis species. A dendrogram derived from a cluster analysis of the complete RFLP dataset showed subdivision of the species into two groups, one comprising the species of the Truncata subsection and the other comprised of the four species of the Emarginata subsection. The findings indicated that Ae. speltoides is the most divergent species within the Sitopsis section, and that Ae. sharonensis and Ae. longissima are closely related species and form a separate subgroup within subsection Emarginata. Ae. bicornis and Ae. searsii also form separate subgroups, where that of Ae. bicornis is closer to the sharonensis-longissima subgroup, and that of Ae. searsii is more distant from the other two subgroups. Similar results were obtained by Goryunova et al. (2008), who used random amplification of polymorphic DNA (RAPD) analysis to study the intraspecific variation and phylogenetic relationships of the Sitopsis species. They found that Ae. speltoides formed the most isolated species in the section, justifying its classification into a separate subsection. In the other subsection, Ae. longissima and Ae. sharonensis were the closest species, Ae. bicornis and Ae. searsii formed separate subgroups, that of Ae. searsii being the most distant. Their findings showed that the extent of intraspecific polymorphism considerably varies among the Sitopsis species: Ae. speltoides is the most polymorphic species of the group, Ae. bicornis and Ae. searsii display the lowest diversity, while Ae. longissima and Ae. sharonensis are intermediate.

That Ae. speltoides differs significantly from the Emarginata species is also evident from studies that used different methodological approaches, e.g., Giemsa C-banding (Teoh and Hutchinson 1983; Friebe and Gill 1996; Friebe et al. 2000), in situ hybridization patterns with DNA probes (e.g., Talbert et al. 1991; (Yamamoto 1992a; Jiang and Gill 1994; Badaeva et al. 1996a, b; Salina et al. 2006; Raskina et al. 2011; Belyayev and Raskina 2013), analysis of the restriction patterns of chloroplast and mitochondrial DNA (Ogihara and Tsunewaki (1988), of biochemical markers (Bahrman et al. 1988), or of repeated sequences (Dvorak and Zhang 1990, 1992a, b; Giorgi 1996) and RFLP analysis (Sasanuma et al. 1996). The separate position of Ae. speltoides within the Sitopsis section and, in contrast, the similarity of the Emarginata species was also shown by Dvorák and Zhang (1992b) who studied variation of repeated nucleotide sequences (RNS). RAPD- and AFLP analyses revealed that Ae. speltoides forms a cluster with polyploid wheats, which is separated from other Sitopsis species (Kilian et al. 2007, 2011; Goryunova et al. 2008). Likewise, study of organellar DNAs by PCR-single-strand conformational polymorphism (PCR-SSCP) revealed high similarity of Ae. bicornis—Ae. sharonensis—Ae. longissima plasmons and their distinctness from plasmon of Ae. speltoides (Wang et al. 1997).

Similar results were obtained by Ruban and Badaeva (2018). These authors studied the relationships between the S-genome species using Giemsa C-banding and fluorescence in situ hybridization (FISH) with several DNA probes. To correlate the C-banding and FISH patterns, they used the microsatellites (CTT)₁₀ and (GTT)₉, which are major components of the C-banding positive heterochromatin in wheat. Their results justify the classification of the Sitopsis species into the two subsections, Truncata and Emarginata, which differ in the C-banding patterns, and distribution of DNA repeats. Evolution of Emarginata species was associated with an increase of C-banding and (CTT)₁₀-positive heterochromatin, as well as amplification of the DNA probe Spelt-52.

In accord with the evidence that Ae. speltoides has less constitutive heterochromatin, as seen from smaller amount of C-banding, than the Emarginata species, the amount of nuclear DNA in subsection Truncata is significantly smaller (5.81 pg 1C DNA) than that in species of subsection Emarginata (6.65 pg 1C DNA in Ae. searsii and 7.52 pg in Ae. sharonensis) (Furuta et al. 1977; Eilam et al. 2007; Table 9.3). No significant intraspecific variation in nuclear DNA size was detected (Furuta et al. 1977; Eilam et al. 2007). Likewise, Li et al. (2022) showed that the five Sitopsis species have variable genome sizes (4.611–6.22 Gb) with high proportions of repetitive sequences (85.99–89.81%); nonetheless, they retain high collinearity with other wheat genomes. Li et al. (2022) concluded that differences in genome size are primarily due to independent post-speciation amplification of transposons rather than to inter-specific genetic introgression.

Hence, although Ae. speltoides has been considered by most taxonomists and cytogeneticists a member of the Sitopsis section, sequencing of chloroplast DNA by Middleton et al. (2014), and Gornicki et al. (2014), and nuclear DNA by Marcussen et al. (2014), showed that Ae. speltoides forms a phylogenetic clade with the B and G subgenomes of allopolyploid wheats and not with the other Sitopsis species. Similar results were presented by Ruban and Badaeva (2018) and by Li et al. (2022). Assembly of chromosome-level genome sequences of all the five Sitopsis species enable Li et al. (2022) to propose that the diploid species and B and G subgenomes of allopolyploid wheats fall into two independent clades, with Ae. speltoides being clustered with the B-subgenome of allopolyploid wheat (B-lineage) while the rest four Sitopsis species being grouped with the D-subgenome (D-lineage) and its diploid donor Ae. tauschii (D-lineage).

Subsection Truncata and subsection Emarginata are genetically isolated from one another, as shown by sterility of hybrids between them. On the other hand, hybrids between Emarginata species are fertile or partially fertile. The F₁ hybrid between Ae. longissima and Ae. sharonensis and the reciprocal hybrid had an 81–85% seed set (Tanaka 1955a; Ankori and Zohary 1962), that between Ae. sharonensis and Ae. bicornis had a 62% seed set (Tanaka 1955a), that between Ae. bicornis and Ae. longissima had a 40% seed set (Waines G, personal communication), and that between Ae. longissima and Ae. searsii had a 35% seed set in one hybrid combination and 7% in another (Feldman et al. 1979).

Ae. speltoides differs from the four species of subsection Emarginata in several additional important features: (i) Ae. speltoides is the only predominantly cross-pollinated species in the section. (ii) Its distribution area is in the central part of the genus area, i.e., in and around the Fertile Crescent, on terra rossa or alluvial soils. In contrast, the species of subsection Emarginata reside in the southern part of the western wing of the Fertile Crescent, with Ae. sharonensis and Ae. bicornis growing on sandy soils, and Ae. longissima on red sandy loam in the coastal plain, on sand derived from Nubian sandstone and on grey calcareous steppe soil or loess in the inland steppe and desert. Ae. searsii grows on terra rossa or basalt soil (Feldman and Kislev 1977; Kimber and Feldman 1987).

The five Sitopsis species differ in their genome size (Eilam et al. 2007), that is primarily due to independent post-speciation amplification of transposons (Yaakov et al. 2013). The sum of repetitive sequences in the different species ranges from 85.99 to 89.81% (Li et al. 2022). Ae. speltoides has the smallest genome, Ae. bicornis and Ae. searsii have a larger genome, and Ae. sharonensis and Ae. longissima have the largest genome.

Ae. speltoides is the only Sitopsis species found in the entire Fertile Crescent and in most parts of Turkey, with both varieties (speltoides and ligustica) often occurring sympatrically and with similar ecological tendencies (Hammer 1980; Kimber and Feldman 1987). Notably, the area of the other four species of the section (Ae. bicornis, sharonensis, longissima, and searsii) occurs within a very limited region in the southeastern corner of the Mediterranean, which marginally overlaps with the southwestern part of the area of Ae. speltoides. It seems as if these species replace Ae. speltoides in the southern climatically and edaphically special environments. Of these, the closely related Ae. sharonensis and Ae. bicornis are an example of bioregional vicariance (Zohary 1962), with the former confined to the coastal plain of the Mediterranean region in Israel and southern Lebanon, and the latter replacing it southwardly up to Lybia and also spreading towards the Saharo-Arabian region in desert sands of the Israeli Negev and Sinai Peninsula. Ae. longissima occurs in two different phytogeographical regions, the Israeli and Egyptian coastal plain (Mediterranean region) and the Israeli Negev, southern Jordan and Sinai Peninsula (Irano-Turanian and Saharo-Arabian regions). Ae. searsii grows in sub-Mediterranean areas in Israel, Jordan, southwestern Syria and southeastern Lebanon and also occurs in the high elevations of the Israeli Negev and southern Jordan. It should also be noted that the distribution of Ae. searsii, similar to Ae. vavilovii, has only recently been defined, and may be larger than hitherto supposed.

9.4.2 Aegilops speltoides Tausch

9.4.2.1 Morphological and Geographical Notes

Ae. speltoides Tausch [Syn.: Triticum speltoides (Tausch) Gren. ex Richter; Sitopsis speltoides (Tausch) Á. Löve] is a predominantly allogamous, annual plant, with 40–70-cm high culms (excluding spikes), and leaves that are usually hairy, and sometimes pendant. Its spike is linear, narrow, tapers to the tip, either two- or one-rowed, and is (6-) 7-11(-15)-cm long (excluding awns). At maturity, its spike disarticulates above the basal rudimentary spikelet, either into single spikelets, with the rachis internode immediately below them (wedge-type dispersal unit), or falling entire as a unit (umbrella-type dispersal unit). Spikelets 7–11, 11–15 mm long, lanceolate or linear, sessile in hollows of rachis, and longer or shorter than the adjacent rachis internode. Each contains 4–8 (usually 4–6) florets, with the upper 1–3 being sterile. There is usually 1 rudimentary spikelet at the base of the spike. Glumes are 5–7 mm long, truncate at the apex, asymmetrical, somewhat keeled, and about 2/3 the length of the lemma. Lemmas are 7–10 mm long and boat-shaped, those of lateral spikelets have either short triangular awns or are not awned at all, while those of terminal spikelets are always have longer awns than those on the lateral spikelets. Anthers are 5–6 mm long. The caryopsis is adherent to lemma and palea (Fig. 9.1a, b).

Ae. speltoides has a median-sized distribution in the central region of the distribution of the genus. It is an eastern Mediterranean element, extending into the steppical (Irano-Turanian) region. It occupies primary and secondary habitats. Ae. speltoides grows in Israel (in the Coastal Plain and Esdraelon Plain on humid, alluvial soil; in Mt. Carmel and western slopes of the Samaria Mts., on terra rossa and Rendzina soils), Jordan, Lebanon, Syria, Turkey, northern Iraq and northwestern Iran. It is a weed in central and western Turkey, Greece, Bulgaria and possibly also Crimea. The center of variation of this species is in north Syria-southeastern Turkey, which is likely the center of its origin.

Ae. speltoides grows sympatrically with Ae. caudata, Ae. umbellulata, Ae. triuncialis, Ae. geniculata, Ae. neglecta, Ae. biuncialis, Ae. peregrina, Ae. cylindrica, wild T. monococcum (ssp. aegilopoides), T. urartu, wild T. timopheevii and wild T. turgidum, and allopatrically with Ae. sharonensis, Ae. longissima and Ae. searsii in its southern distribution area and with Ae. tauschii in its northeastern distribution area.

Ae. speltoides has limited morphological variation, mainly in spike characteristics. It contains two main morphological types that differ markedly in the structure of their fruiting spike and consequently in their mode of seed dispersal. Taxonomists (see van Slageren 1994) regard these two types as varieties, var. speltoides Tausch and var. ligustica (Savign.) Fiori in Fiori & Paoletii.

Var. speltoides [Syn.: Ae. speltoides ssp. speltoides Hammer; Ae. speltoides var. aucheri (boiss.) Fiori in Fiori & Paoletii; Ae. aucheri Boiss.] is characterized by a one-rowed, long and cylindrical spike, with relatively widely interspersed spikelets. Only the apical spikelet is awned and rarely has awns on some lateral spikelets and then shorter and thinner than the awns on the lateral spikelet. The rachis of the spike is tough, except for a brittle node at its base, above a rudimentary spikelet. The spike falls at maturity as a unit (umbrella-type dispersal unit) (Fig. 9.1b).

Var. ligustica [Syn.: Ae. ligustica (Savign.) Coss.; Ae. speltoides ssp ligustica (savign,) Zhuk.; Agropyron ligusticum Savign.] is characterized by a denser, two-rowed spike, in which the lateral spikelets are also awned; awns of terminal spikelet are somewhat sturdier than those of lateral spikelets. The rachis is brittle at every joint, so that the mature spike disarticulates into single spikelets, each with the rachis internode immediately below it (wedge-type dispersal unit). Each individual spikelet thus serves as an independent dissemination unit; it is mechanically adapted to insert itself into the ground and usually buries itself soon after detachment (Fig. 9.1a). The two varieties grow, in most sites, in mixed stands and are cross-fertile; few intermediates are usually found in mixed populations. Presumably, the ligustica form, having the wedge-type dispersal unit typical of many Triticineae species, is the ancestral form. Alternatively, the speltoides form, having one-rowed spike similar to that of several Elymus species, may be the ancestral type.

The morphological differences between var. speltoides and var. ligustica are restricted to the fruiting spike alone. In all other traits, such as vegetative characters and phenological behavior, the types are strikingly similar to each other (Miczynski 1926; Schiemann 1928; Eig 1929a). In addition, F₁ hybrids between the two types are fully fertile and show complete pairing of chromosomes at meiosis (Kihara and Lilienfeld 1932; Sears 1941b; Zohary and Imber 1963). Further indications of the close relationship between speltoides and ligustica were obtained from the usual occurrence of mixed stands of the two types (Eig I929a). There are wide fluctuations in the proportions between ligustica and speltoides in different sites and in different plant formations, but both types were almost universally present in the sites examined. In many stands, ligustica and speltoides plants were, more or less, evenly mixed. Yet, ligustica plants predominated the stands in higher altitudes and speltoides in warmer sites (Zohary and Imber 1963). These observations led to the conclusion that mixed populations are the rule; speltoides and ligustica are apparently spatially inter-connected almost throughout their distribution area (Zohary and Imber 1963).

Rare intermediate types, e.g., plants with a laterally awned but tough rachis (var. polyathera according to Eig 1929a) or also, very occasionally, plants with brittle rachis but laterally awnless spikes, occur in very low frequencies (less than 1%) in mixed speltoides-ligustica populations (Zohary and Imber 1963). Progeny of speltoides x ligustica crosses revealed that the main differences in spike morphology are inherited in an almost monohybrid Mendelian fashion (dominance of ligustica over speltoides in F₁, and 3:1 segregation in F₂) (Miczynsky 1926; Schieman 1928; Kihara and Lilienfeld 1932). These data were first interpreted as indicating the presence of only a single pleiotropic gene, but Sears (1941b), based on occasional occurrence of recombinants or intermediate plants, proposed the operation of a block of closely linked genes. This block should possess at least three genes: one determining the length of rachis internodes (short in ligustica bringing about two-rowed spikes and long in speltoides leading to one-rowed spikes); a second gene that determines awns on the lateral spikelets; and a third gene that determines the type of dispersal unit. Luo et al. (2005) mapped the closely linked genes controlling the ligustica/speltoides spike dimorphism to the centromeric region of chromosome 3S. (the genome symbol of Ae. speltoides is S). The location near the centromere, being a chromosome region with rare recombination events, may explain the rare recombination between these three genes.

Progeny tests performed in plants sampled from natural mixed populations, indicated that speltoides and ligustica plants are genetically interconnected by virtue of their mating system of predominantly cross-pollination. Several ligustica plants, that apparently were F₁, segregated into 3 ligustica: 1 speltoides and several speltoides plants, that were presumably pollinated by ligustica, yielded ligustica progeny (Zohary and Imber 1963). Accordingly, Zohary and Imber (1963) suggested not to regard speltoides and ligustica types as two independent varieties, but as two constituents of dimorphic populations. The mixed stands of the two types are looked upon as a case of genetically determined fruit dimorphism, where two types of seed dispersal apparatus function in the same population in a complementary way (Zohary and Imber 1963). In the speltoides-ligustica pair, the two kinds of dispersal units are borne by different individuals: two types of plants, which differ with regard to their fruit structure, form a common population. This is an instance of genetically determined fruit dimorphism, with a population as its unit of operation.

A model of fruit dimorphism can account for the peculiar mode of inheritance of the many differences between the two types: they are inherited in an almost monohybrid fashion, with only rare cases of recombinants. As already proposed by Sears (1941b), this mode of inheritance suggests the presence of a single block of closely linked genes. A system based on such a single block is essential for the establishment of population dimorphism. The parallel evolvement of fruit dimorphism in two Sitopsis groups, ligustica vs. speltoides on the one hand, and bicornis-sharonensis versus longissima-searsii on the other hand, is an interesting evolutionary phenomenon. The genetic change(s) determining the speltoides and the longissima-spike morphology occurred independently in the two groups, or alternatively, happened once and genes of this system were transferred from one group to the other via interspecific introgressive hybridization. However, it seems there is a difference between the gene system in the two groups; while the genes in ligustica-speltoides are closely linked, those in sharonensis-longissima are not linked, and segregate independently from one another, as is indicated in the scatter diagram of F₂ artificial generation in Ankori and Zohary (1962).

9.4.2.2 Cytology, Cytogenetics, and Evolution

Aegilops speltoides is a diploid species containing seven pairs of homologous chromosomes (2n = 2x = 14), all of which have a median or sub-median centromere and lack distinctive morphological features, except for the SAT chromosomes (Riley et al. 1958; Chennaveeraiah 1960). In contrast, Dong et al. (2017), studying karyotypic polymorphism of Ae. speltoides, found intraspecific variation in the centromere position. In Ae. speltoides var. ligustica, there is one pair with a median centromere, while the rest of the pairs in both taxa have submedian centromeres (Chennaveeraiah 1960). The difference between the karyotypes of the two taxa seems to be very small. However, in var. ligustica, the short arms in the satellite pairs are more or less of the same length. In var. speltoides, there seems to be a difference in the lengths of the short arms of the satellite pairs (Chennaveeraiah 1960). The karyotype of both varieties of Ae. speltoides contains two pairs of SAT chromosomes with fairly large satellites (Pathak 1940; Riley et al. 1958; Chennaveeraiah 1960).

In Ae. speltoides, the genes coding for the 18S and 26S ribosomal RNA (rDNA genes) are located in the NOR regions on the short arms of chromosomes of groups 1 and 6 (Dvorak et al. 1984). There is only one 5S rDNA locus on the short arm of group 5 chromosomes that codes for 5S rRNA (Badaeva et al. 1996b). In contrast, Ae. longissima, Ae. sharonensis, Ae. bicornis and Ae. searsii, exhibit a different distribution pattern of NORs and 5S DNA loci. They contain major NOR loci on the short arms of chromosomes of groups 5 and 6 and two 5S rDNA loci on the short arms of chromosomes of groups 1 and 5 (Badaeva et al. 1996b). Thus, the distribution pattern of the NORs and 5S loci in the five Sitopsis species support the subdivision of this section into the two subsections.

The genome of Ae. speltoides is the smallest among the five Sitopsis species; 1C DNA is 5.81 pg (Furuta et al. 1977; Eilam et al. 2007) and 5.1 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. speltoides genome after genome sequencing and assembly to be somewhat smaller amount, i.e., 4.6 Gb (=4.5 pg), and Avni et al. (2022), also after sequencing and assembly, reported 5.13 Gb (=5.02 pg). No significant difference was noted in the genome size of the two taxa (Furuta et al. 1977; Eilam et al. 2007). Teoh and Hutchinson (1983), used an improved C-banding technique, found that all five Sitopsis species have telomeric, interstitial, and centromeric bands, varying in size and staining intensity. However, Ae. speltoides has exceptionally distinctive and extensive centromeric bands. The extent of polymorphic variation was found to vary between chromosomes of Ae. speltoides and plants were found to be heterozygous for the banding pattern (Teoh and Hutchinson 1983). Yet, the intraspecific variation in C-banding patterns neither differentiated between var. ligustica and var. speltoides, nor corresponded with impaired meiotic chromosomal pairing, since hybrids between the two varieties exhibited complete pairing and full fertility (Teoh and Hutchinson 1983). Badaeva et al. (1996a, b) and Ruban and Badaeva (2018) also found that Ae. speltoides have a different C-banding pattern than those of the other species of section Sitopsis.

Friebe and Gill (1996) established the homoeology between chromosomes of Ae. speltoides and those of T. aestivum, on the basis of similarities in chromosome morphology and C-banding patterns. Maestra and Naranjo (1998) determined the homoeologous chromosome relationship by analyzing homoeologous pairing at meiotic first metaphase of F₁ hybrids between Triticum aestivum and Ae. speltoides carrying the homoeologous–pairing suppressor gene of common wheat, Ph1, or deficiencies ph1b for Ph1, and ph2b for Ph2. The chromosomes and their arms were identified, in both species, by C-banding. Data from relative pairing affinities were used to determine homoeologous relationships between Ae. speltoides chromosomes and bread wheat. All arms of the seven chromosomes of the speltoides genome showed normal homoeologous pairing with the wheat chromosomes, implying that no apparent chromosome rearrangements occurred in the evolution of Ae. speltoides relative to chromosomal structure in wheat. There was agreement between Friebe and Gill (1996) and Maestra and Naranjo (1998) in the assignment of five speltoides chromosomes to homoeologous groups 1, 4, 5, 6 and 7, but discrepancy with regards to chromosomes 2S and 3S.

Using in situ hybridization (ISH), Badaeva et al. (1996a) found that all Sitopsis species were similar to each other and to Amblyopyrum muticum in the distribution of the hybridization sites of two highly repetitive DNA sequences, pSc119 and pAsl. Yet, chromosomes of Ae. speltoides contain rich repetitive DNA sequences highly homologous to the pSc119 probe and, on the other hand, lack such homology to probe pTa53 (Dong et al. 2017). The distribution of pSc119 on chromosomes of Ae. speltoides show differences between accessions, between plants of one accession and even between homologous chromosomes in one plant.

Ae. speltoides comprises many of TEs, particularly long terminal repeat (LTR) retrotransposons, of which Ty1-copia superfamilies (Angela, Barbara, and Wis-A) and Ty3-gypsy superfamilies (Fatima and Erika) make up the main fraction (Middleton et al. 2013; Yaakov et al. 2013). Non-LTR retrotransposons (Ramona and Paula) and DNA transposons (Baldwin, Rong, and Charon) also exist in Ae. speltoides. All these TEs families vary greatly in copy number among different accessions of Ae. speltoides (Yaakov et al. 2013). Likewise, Hosid et al. (2012) analyzed intraspecific variation of four LTR retrotransposons (WIS2, Wilma, Daniela, and Fatima) in 13 different populations of Ae. speltoides from all over the distribution area of this species and found significant diversity in retrotransposon distribution. The various genotypes significantly differ with respect to the patterns of the four explored LTR retrotransposons, indicating a constant ongoing process of LTR retrotransposon fraction restructuring among and within populations of Ae. speltoides. Maximum changes were recorded in genotypes from small, stressed populations. The data of Hosid et al. (2012) revealed dynamic changes in LTR retrotransposon fractions in the Ae. speltoides genome, that are continually reshaping the genome of Ae. speltoides, particularly in stressful environments (Raskina et al. 2004a, b, 2008).

In Ae. speltoides euchromatin, widely interspersed TEs are clustered non-randomly, and may affect chromosomal structure. For instance, Raskina et al. (2004a) reported on the involvement of the transposable element system En/Spm [Enhancer (En) and Suppressor-Mutator (Spm)] in ongoing chromosomal repatterning in a small, isolated, peripheral population of Ae. speltoides. Cytogenetic analysis of the dynamics of En/Spm transposons in meiosis indicated that this transposon is active during male gametogenesis, changing the position of the rDNA sites (Raskina et al. 2004a). Such findings may indicate the importance of TEs in genome architecture (Belyayev et al. 2001, 2005; Altinkut et al. 2006), as well as their most important role in intraspecific divergence (Belyayev et al. 2010). The variability of TE content in Ae. speltoides, as well as in other Aegilops and Triticum species, might has a great impact on the dynamic, ongoing evolution of their genomes (Charles et al. 2008; Yaakov and Kashkush 2012).

Repetitive DNA sequences form chromosome-specific heterochromatin patterns in Ae. speltoides (Salina et al. 2006; Raskina et al. 2008). The species-specific tandem-repeat Spelt1 forms distinct clusters almost exclusively in subtelomeric chromosome positions, while Spelt52 clustering was most prominent at interstitial sites (Salina et al. 2006; Raskina et al. 2011). These two repetitive DNA sequences exhibit high intraspecific variation in both number and size (Salina et al. 2006; Raskina et al. 2011). Using fluorescence in situ hybridization, Salina et al. (2006) observed considerable polymorphisms in the hybridization patterns of Spelt1 between and within the studied lines of Ae. speltoides. There was a distinct ecogeographical gradient in the abundance of Spelt1 and Spelt52 blocks in Ae. speltoides; in marginal populations, the number of Spelt 1 chromosomal blocks was, at times, 12–14 times lower than in the center of the species distribution (Raskina et al. 2011). Likewise, Ae. speltoides had distinct Spelt52 hybridization patterns in the studied lines, but several distal Spelt52 sites, namely, on chromosome arms 3SL, 6SS, and 7SS, were common to all.

Aegilops speltoides contains a genetic mechanism that, in hybrids with allopolyploid wheats, suppresses the activity of the homoeologous-pairing suppressor (Ph1) gene of allopolyploid wheats, Triticum aestivum and T. turgidum. Riley and Law (1965) and Dover and Riley (1977) pointed out that increased homoeologous chromosome pairing at meiosis of common wheat x Ae. speltoides occurs only if Ph1 of common wheat is present and therefore proposed that the Ae. speltoides genome can promote homoeologous chromosome pairing by overpowering the activity of Ph1. Chen and Dvorak (1984) and Dvorak et al. (2006) found that the speltoides genes were ineffective in diploid hybrids lacking a gene like Ph1. Consequently, in accord with the above conclusion of Riley and Law (1965) and Dover and Riley (1977), they concluded that Ae. speltoides genes act on the Ph1 locus by suppressing its expression in common wheat x Ae. speltoides hybrids. Dvorak et al. (2006) mapped these, Ae. speltoides suppressor genes and demonstrated their significant effects as Mendelian loci on the long arms of chromosomes 3S and 7S. The chromosome 3S locus was designated Su1-Ph1 and the chromosome-7S locus was designated Su2-Ph1 (Dvorak et al. 2006). A QTL with a minor effect was mapped on the short arm of chromosome 5S and was designated QPh.ucd-5S (Dvorak et al. 2006). The expression of Su1-Ph1 and Su2-Ph1 increased homoeologous chromosome pairing in common wheat × Ae. speltoides hybrids by 8.4 and 5.8 chiasmata/cell, respectively. Su1-Ph1 was fully epistatic to Su2-Ph1, and, when acting together, the two genes increased homoeologous chromosome pairing in common wheat × Ae. speltoides hybrids to the same level when Su1-Ph1 acting alone. QPh.ucd-5S expression increased homoeologous chromosome pairing by 1.6 chiasmata/cell in common wheat × Ae. speltoides hybrids and was additive to the effect of Su2-Ph1. It is hypothesized that the products of Su1-Ph1 and Su2-Ph1 affect pairing between homoeologous chromosomes by regulating the expression of Ph1, while the product of QPh.ucd-5S primarily regulates recombination between homologous chromosomes.

The Ae. speltoides genes that suppress Ph1 activity were transferred from a high-pairing accession of Ae. speltoides to common wheat (Chen et al. 1994). It appeared that two genes, presumably Su1-Ph1 and Su2-Ph1, were transferred. These suppressor genes were transferred to several lines of common wheat and used to induce homoeologous pairing in hybrids between these lines and wild relatives of wheat and, consequently, enhanced the gene transfer from wild relatives to common wheat (Li et al. 2011).

Yet, it is puzzling to assume that the homoeologous-pairing promoters of Ae. speltoides (and also of Amblyopyrum muticum) evolved at the diploid level as a preadaptation, to repress the expression of the pairing suppressors at the polyploid level. These promoters may also promote homologous and homoeologous pairing per se or counteract the activity of suppressors that exist at the diploid level. Meiotic chromosome pairing data, collected by Sears (1941b), from hybrids of various diploid Triticineae species were interpreted by Waines (1976) to indicate that intraspecific genotypic differences control the amount of homoeologous chromosome pairing in diploid inter-specific hybrids. He suggested that homoeologous-pairing suppressors analogous to the Ph1 allele in bread wheat, are already present in genotypes of diploid species and do not necessarily have to arise by mutation or translocation after allopolyploid formation. He suggested that these homoeologous-pairing suppressors reinforce the isolating mechanism among the diploid species. The existence of Ph1-like genes in diploid species of the wheat group was also suggested by Okamoto and Inomata (1974) and by Maan (1977a). Feldman (1978) presented evidence for the existence of homoeologous-pairing suppressors in a low-pairing line of Ae. longissima.

There is also evidence that the promoters of the diploid species promote homoeologous pairing per se, not necessarily via the repression of Ph1 expression. Feldman and Mello-Sampayo (1967) concluded that, Ae. speltoides carries a promoter(s) that directly promotes homoeologous pairing in bread wheat x Ae. speltoides hybrids lacking the Ph1 gene. Similarly, all the promoters that exist in bread wheat promote pairing rather than suppress the expression of Ph1 (Feldman 1966). Indication that Ae. speltoides (and also A. muticum) contain genes that promote homoeologous pairing in the absence of Ph1 may be deduced from the following pairing data. The F₁ hybrids between Ae. speltoides and T. monococcum and between Ae. speltoides and Ae. comosa analyzed by Sears (1941b), showed a similar amount of chromosome pairing to that observed by Ohta (1990, 1991) in the F₁ hybrids between T. monococcum and A. muticum, between Ae. comosa and A. muticum, and between Ae. speltoides and A. muticum.

Massive chromosome restructuring was observed during the substitution of chromosome 6B of cultivar Chinese Spring of bread wheat by chromosome 6S from Ae. speltoides (Kota and Dvorak 1988). The chromosome rearrangements including deletions, translocations, ring chromosomes, dicentric chromosomes and a paracentric inversion in both euchromatic and heterochromatic regions of both wheat and Ae. speltoides chromosomes. The frequency of chromosome rearrangements was high among the B-subgenome chromosomes, moderate among the A-subgenome chromosomes, and low among the D-subgenome chromosomes (Kota and Dvorak 1988). In the B subgenome, the rearrangements were nonrandom, and occurred most frequently in chromosomes 1B and 5B. These observations indicate that wheat genomes can be subject to uneven rates of structural chromosome differentiation, even when within the same nucleus. Other examples of massive chromosome aberrations in wheat and Aegilops species are rare but were observed in root-tip cell of the Brazilian semi-dwarf cultivar IAS-54 of bread wheat (Dos Santos Guerra et al. 1977), in mitotic and meiotic cells of an individual plant of Elymus farctus (=Agropyron junceum) (Heneen 1963), and in first meiotic metaphase of an Ae. longissima line (TL02) (Feldman and Strauss 1983). Chromosome instability was also associated with gametocidal effects caused by several alien chromosomes of homoeologous group 4 in the monosomic state (Maan 1975; Finch et al. 1984; Tsujimoto and Tsunewaki 1984; Endo 1985).

Aegilops speltoides and Amblyopyrum muticum, the two-primitive species of the wheat group, are predominantly cross-pollinated, and contain genes promoting homoeologous pairing. On the other hand, these two species contain B chromosomes that suppress such pairing (Simchen et al. 1971; Mendelson and Zohary 1972; Zarchi et al. 1974; Mochizuki 1957, 1960, 1964; Ohta 1995a). B chromosomes are only known to occur spontaneously in cross-pollinating taxa (Müntzing et al. 1969) and are fully absent in self-pollinators (Jones et al. 2008). In both species, Ae. speltoides and A. muticum, the B chromosomes are absent in the roots but stably present in the aerial tissue (Mendelson and Zohary 1972; Ohta 1995a); a maximum of eight Bs per cell has been reported in Ae. speltoides (Raskina et al. 2004b). A comparable situation of tissue type-specific B chromosome distribution is also known for Agropyron cristatum (Baenziger 1962). The B chromosomes of Ae. speltoides and A. muticum are not homologous since they do not pair in F₁ hybrids between these two species (Vardi and Dover 1972). The presence of one to three Bs has a positive effect on the plant, whereas a higher number of Bs reduces fertility and vigor (Mendelson and Zohary 1972; Belyayev et al. 2010).

The B chromosomes of Ae. speltoides are also characterized by a few A-chromosome-localized repeats, like Spelt1, pSc119.2 tandem repeats, 5S rDNA and Ty3-gypsy retroelements (Friebe et al. 1995c; Raskina et al. 2011; Hosid et al. 2012; Belyayev and Raskina 2013). It is assumed that these B chromosomes originated from the standard set of A chromosomes as a consequence of interspecific hybridization or, more likely, from trisomic (2n + 1) plants. Several lines of Ae. speltoides form a low frequency of unreduced gametes as a result of meiotic disorders or spontaneous non-disjunction of the entire chromosome complement, leading to the formation of triploid plants upon the fusion of the 2n gamete with 1n gamete (Belyayev and Raskina 2013; Feldman M, unpublished). Self-pollination of such triploids yields aneuploids and trisomic plants. Several trisomic Ae. speltoides plants were occasionally found in plants that originated from different populations (Feldman M, unpublished). Potential donors of the B chromosome of Ae. speltoides are the A chromosomes 1S, 4S and 5S of the Ae. speltoides genome (Friebe et al. 1995c; Belyayev and Raskina 2013). All Bs in Ae. speltoides have a single intercalary Spelt1 tandem repeat cluster and a 5S rDNA cluster in both arms (Raskina et al. 2011). In addition, a large intercalary cluster of Ty3-gypsy elements was found in close proximity to the 5S rDNA and Spelt1 blocks (Belyayev and Raskina 2013). Since chromosome 4S is the only Ae. speltoides chromosome that carries the intercalary Spelt1 cluster, and chromosome 5S, which is an exclusive source of 5S rDNA in the Ae. speltoides genome, it may be involved in the heterologous synapses and recombination resulting in the formation of Bs in this species (Raskina et al. 2011). Support of the idea that B derived from A chromosome(s) also comes from the observation that Bs pair at meiosis, although very rarely, with A chromosome(s) (Belyayev and Raskina 2013). The similarity in the B chromosome structures throughout the species range indicated that they were generated from a similar heterologous recombination of certain A chromosomes.

Ae. speltoides B chromosomes suppress homoeologous meiotic pairing of A chromosomes in intergeneric hybrids with bread wheat lacking the ph1 gene (Vardi and Dover 1972; Dover and Riley 1972). The effect of B chromosomes is like that of wheat Ph1 on homoeologous pairing in wheat hybrids with alien species. If Ae. speltoides B chromosomes derived from one of the A chromosomes, then one of the first essential prerequisites for the establishment of B chromosome as an independent entity should be to prevent pairing and recombination between the newly formed B and the ancestral A chromosome(s). Thus, the development of a genetic system suppressing homoeologous pairing was necessary for the independent existence and evolution of B chromosomes.

B chromosomes suppress homoeologous pairing in interspecific hybrids and thus, counteract the activity of the pairing promoters of Ae. speltoides. In a predominantly cross-pollinated species, the presence of genes that promote pairing is important, either to assure complete pairing in intraspecific hybrids or to increase pairing and recombination in interspecific ones. Yet, the presence of B chromosomes in some individuals of this species helps to conserve the integrity of their genomes.

9.4.2.3 Crosses with Other Species of the Wheat Group

Riley (1966a), Vardi and Dover (1972), and Ohta and Tanaka (1983) determined mean chromosome pairing at first meiotic metaphase in F₁ hybrids of Ae. speltoides x Amblyopyrum muticum with and without B chromosomes (Tables 8.1 and 8.2). The hybrid speltoides x muticum without B chromosomes had mean pairing much higher than hybrid with four B chromosomes. Seed fertility of the F₁ hybrids was very low (about 2.0%). The hybrids with B chromosomes, that suppress homoeologous pairing, indicating that the high pairing in hybrids without B chromosomes results mainly from homoeologous pairing promoted by the homoeologous-pairing promoters that exist in these diploids. Hence, the above pairing data show that the genomes of these two species have substantially diverged from one another.

In general, chromosome pairing at meiotic first metaphase in F₁ hybrids between species of the Sitopsis section has been found to be high (Table 9.4) and consequently, Kihara (1949, 1954) grouped the Sitopsis species all together with the same primary genome symbol, S. The species of subsection Emarginata, namely, Ae. sharonensis, Ae. longissima, Ae. bicornis and Ae. searsii, form fertile F₁ hybrids with one another, whereas F₁ hybrids of any of these species with high- or low-pairing types of Ae. speltoides, the only species of subsection Truncata, show slightly reduced pairing and are sterile (Sears 1941b; Kihara 1949; Tanaka 1955a; Roy 1959; Riley et al. 1961; Kimber 1961; Ankori and Zohary 1962; Feldman et al. 1979). Thus, it is clear that the Ae. sharonensis-longissima-bicornis-searsii are of close affinity, while Ae. speltoides is somewhat more distant.

Table 9.4 Chromosome pairing at first meiotic metaphase of F₁ hybrids between diploid Aegilops species

Hybrids between high-pairing lines of Ae. speltoides and other diploid species of Aegilops, i.e., Ae. caudata, Ae. comosa, Ae. uniaristata, and Ae. umbellulata exhibit about 10.70–11.26 (0.76–0.80%) paired chromosomes, while hybrids between low-pairing type of Ae. speltoides and Ae. caudata, Ae uniaristata, and Ae. umbellulata show only 4.92–7.30 (35–60%) paired chromosomes (Table 9.4). Evidently, the pairing data indicate that Ae. speltoides is more distant to these diploid species than to species of subsection Emarginata.

Chromosome pairing between the allotetraploid species Ae. kotschyi and high-pairing and low-pairing types of Ae. speltoides exhibit somewhat reduced pairing (Table 9.5). The F₁ hybrid between Ae. kotschyi and high pairing line of Ae. speltoides show 14.65 paired chromosomes (69.8%), whereas hybrids between Ae. kotschyi and low-pairing type of Ae. speltoides show only 13.8 (65.7%) paired chromosomes (Rubenstein and Sallee 1973). Hybrid between Ae. peregrina with low pairing type of Ae. speltoides showed 8.02 (38.2%) paired chromosomes (Yu and Jahier 1992). Also, F₁ hybrid between other allotetraploid species of section Aegilops, namely, Ae. biuncialis, Ae. columnaris, Ae. neglecta and Ae. triuncialis, with Ae. speltoides, presumably a high-pairing type, showed similar levels of reduced pairing (Kihara 1949). In all the above diploid and triploid hybrids, the speltoides high-pairing genotypes promoted higher pairing between the homoeologous chromosomes of the studied species than the low-pairing genotypes. Data of chromosome pairing between allohexaploids of Aegilops section Vertebrata, Ae. crassa 6 x and Ae juvenalis, and Ae. speltoides are presented in Table 9.6. These hybrids had higher pairing because of the effect of the homoeologous-pairing promoters in Ae. speltoides.

Table 9.5 Chromosome pairing at first meiotic metaphase of F₁ hybrids between allotetraploids and diploids of the genus Aegilops

Table 9.6 Chromosome pairing at first meiotic metaphase of F₁ hybrids between allohexaploids and diploids of the genus Aegilops

Data of chromosome pairing between allohexaploids of Aegilops section Vertebrata, Ae. crassa 6x, Ae juvenalis, and Ae. vavilovii, and diploid species of Aegilops are presented in Table 9.6. The hybrid Ae. crassa 6x x Ae. tauschii had somewhat higher pairing than most other hybrids indicating high homology between the second D subgenome of the hexaploid and the D genome of the diploid. The remaining hybrid exhibited much less pairing. Chromosome pairing in F₁ hybrids between allopolyploid species of Aegilops indicates that also several chromosomes of the differential subgenomes are involved in pairing in addition to that of he shared subgenome (Table 9.7).

Table 9.7 Chromosome pairing at first meiotic metaphase of F₁ hybrids between allopolyploids of the genus Aegilops

Mean chromosomal pairing at meiosis in F₁ hybrids between high-pairing type of Ae. speltoides and domesticated T. monococcum, ssp. monococcum, was very high, almost complete (Table 9.8). Similar high pairing was observed in the F₁ hybrid between Ae. speltoides (presumably a high-pairing line) and wild T. monococcum, ssp. aegilopoides, was reported by Shang et al. (1989). However, mean chromosomal pairing at meiosis of F₁ hybrids between the low-pairing type of Ae. speltoides and T. monococcum ssp. monococcum was low (Table 9.8) These results indicate that the speltoides homoeologous-pairing promoter genes bring about a high degree of homoeologous pairing in speltoides x T. monococcum hybrids, and, when these genes are absent, as in the low-pairing genotype, the low level of pairing reveals excessive divergence of the genomes of these two species.

Table 9.8 Chromosome pairing at first meiotic metaphase of F₁ hybrids between diploid species of Triticum and Aegilops

McFadden and Sears (1947) reported that mean chromosomal pairing in the triploid hybrid between wild tetraploid wheat T. turgidum ssp. dicoccoides x Ae. speltoides was high, about two-third of the chromosomes paired (Table 9.9). Similar results were obtained by Riley et al. (1958) in hybrids between several subspecies of tetraploid wheat T. turgidum and Ae. speltoides. Yet, F₁ hybrids between T. turgidum ssp. durum x low-pairing type of Ae. speltoides showed much lower pairing, involving very few paired chromosomes (Shands and Kimber 1973).

Table 9.9 Chromosome pairing at first meiotic metaphase of F₁ hybrids between allotetraploid species of Triticum and diploid species of Aegilops

The triploid hybrid between the second tetraploid species of wheat, T. timopheevii and Ae. speltoides showed somewhat higher pairing than that between T. turgidum and Ae. speltoides (Table 9.9). Hybrids between T. timopheevii with the high-pairing Ae. speltoides genotype showed pairing of more than ten chromosomes, while hybrids with the low-pairing Ae. speltoides genotype showed pairing of seven bivalents only (Shands and Kimber 1973). These data indicate greater homology between Ae. speltoides and one of the subgenomes of T. timopheevii than in T. turgidum x Ae. speltoides hybrids.

Mean chromosome pairing at first meiotic metaphase in hybrids between hexaploid wheat, T. aestivum ssp. aestivum (bread wheat) and the high-pairing line of Ae. speltoides is very high (Table 9.10). On the other hand, hybrids with the low-pairing Ae. speltoides genotype, showed much reduced chromosomal pairing (Kimber and Athwal (1972).

Table 9.10 Chromosome pairing at first meiotic metaphase of F₁ hybrids between allohexaploid species of Triticum and diploid species of Aegilops

Maestra and Naranjo (1998) analyzed homoeologous chromosome pairing at first meiotic metaphase in F₁ hybrids derived from crosses of bread wheat, either carrying the homoeologous-pairing suppressor, Ph1, or the deficiency ph1b, lacking this gene, with a high-pairing genotype of Aegilops speltoides. Data from relative pairing affinities were used to predict homoeologous relationships of Ae. speltoides chromosomes to wheat. Chromosomes of both species, and their arms, were identified by C-banding. The Ae. speltoides genotype carried genes that induced a high level of homoeologous pairing in the two types of hybrids analyzed. All arms of the seven chromosomes of the speltoides S genome showed normal homoeologous pairing that implies that no apparent chromosome rearrangements occurred in the evolution of the genome of Ae. speltoides relative to wheat evolution. A pattern of preferential pairing of two types, A-D and B-S, confirmed that the S genome is very closely related to the B subgenome of wheat. Although this pairing pattern was also reported in hybrids of wheat with Ae. longissima and Ae. sharonensis, a different behavior was found in group 5 chromosomes. In the hybrids of bread wheat with Ae. speltoides, chromosome 5B-5S pairing was much more frequent than 5D-5S, while these chromosome associations reached similar frequencies in the hybrids of Ae. longissima and Ae. sharonensis. These results are in agreement with the hypothesis that the B subgenome of wheat is closely related to the B subgenome of Ae. speltoides.

9.4.3 Aegilops bicornis (Forssk.) Jaub. & Spach

9.4.3.1 Morphological and Geographical Notes

Aegilops bicornis (Forssk.) Jaub. & Spach [Syn.: Triticum bicorne Forssk.; Sitopsis bicornis (forssk.) Á. Löve] is predominantly an autogamous annual plant. Culms are slender, erect, 15–45-cm-high. Spikes are 4–5(-8)-cm-long (excluding awns), two-rowed, mostly awned, with 8–15 (-19) spikelets, disarticulating at maturity into spikelets, each spikelet falls with the rachis internode immediately below it (Fig. 2.3; wedge-type dispersal unit). Several lower spikelets often remain attached at the tip of culm. Spikelets are 5.5–8.5-mm-long (excluding awns), mostly 3-flowered (2 lower florets are fertile). The attached rachis internode is half the length of the spikelet. Rudimentary spikelets at the base of the spike are absent. Glumes are 4.5–5.5-mm-long, emarginated or with an angle between teeth, asymmetrical and keeled. Lemmas are 4–6-mm-long, boat-shaped, mostly ending in a slender, 4.5–6.0-cm-long awn (in lower spikelets, awns are short or absent); the awn is usually not flanked by lateral teeth. The caryopsis adheres to lemmas and palea (Fig. 9.1c).

The species includes two varieties: var. bicornis (var. typica Eig) and var. anathera [var. mutica (Aschers.) Eig; Triticum bicorne muticum (Aschers.) Eig]. In var. bicornis, all spikelets are awned, sometimes, the lowest is awnless, whereas, in var. anathera, all spikelets, except for the uppermost 1–3, are awnless. Var. anathera is rare and sporadic, grows together with var. bicornis.

Ae. bicornis exhibits limited morphological variation, involving mainly spike and spikelet size, plant size, and degree of awn development. It is close in spike morphology to Ae. sharonensis, Ae. speltoides var. ligustica, Triticum monococcum ssp. aegilopoides and T. urartu.

Ae. bicornis is a semi-desert element, distributing in southeastern Mediterranean and in Saharo-Arabian regions. It occurs in the coastal regions of Libya (Cyrenaica), Egypt (lower Egypt and Sinai) and southern Israel (western Negev), as well as in inland desert area in Israeli Negev and in Southern Jordan (Edom). Few populations were found in northeastern Cyprus. It grows from sea level to 200 m above sea level (in southern Jordan, it is found at 200–900 m), in areas with an annual rainfall of 75–275 mm (the driest part of the Mediterranean), usually on stable sandy soils, in open dwarf shrubs or herbaceous steppe-like or desert-like formations, xeric coastal and desert plains, in plantations, edges of cultivation and roadsides. It is common in coastal regions, sometimes in dense populations in the coastal plain of southern Israel, Sinai, and lower Egypt, and sporadic in inner sandy deserts. It is a very early maturing type. Ae. bicornis may have originated in the southern part of the Fertile Crescent. Currently, its center of variation is lower Egypt and the Sinai Peninsula. In the Israeli Negev and south Jordan (and possibly also in Egypt), Ae. bicornis grows sympatrically with Ae. longissima, and Ae. kotschyi. In south Israel, it grows allopatrically with Ae. sharonensis and Ae. peregrina.

9.4.3.2 Cytology, Cytogenetics, and Evolution

Ae. bicornis is a diploid (2n = 2x = 14) whose nuclear genome is a modified S genome (designated S^b by Kihara 1954, Kimber and Tsunewaki 1988; Dvorak 1998) and its organellar genome is unique (designated S^b by Ogihara and Tsunewaki 1988). Its nuclear 1C DNA content is relatively high (6.84 ± 0.097 pg; Eilam et al. 2007; 7.1 pg; The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. bicornis genome after genome sequencing and assembly to be 5.73 Gb (=5.6 pg), a value that is smaller than those above that were obtained by flow cytometry measurement.

Ae. bicornis has a symmetric karyotype (one chromosome pair has an almost median centromere and the rest of the pairs have submedian centromeres), with one pair with large and another with small satellites on the short arms (Senyaninova-Korchagina 1932; Riley et al. 1958; Chennaveeraiah 1960). The two SAT-chromosomes belong to homoeologous groups 5 and 6 (Friebe and Gill 1996). According to Riley et al. (1958), the karyotype of Ae. bicornis is similar to that of Ae. sharonensis and Ae. longissima.

Ae. bicornis, like the other species of subsection Emarginata, namely, Ae. longissima, Ae. sharonensis, and Ae. searsii, has major NOR loci on chromosomes of groups 5 and 6 and a variable number of minor loci on chromosomes of homoeologous groups 1, 3, 5 and 6 (Badaeva et al. 1996b). The 5S rDNA loci were observed in these species on chromosomes of groups 1 and 5, distal to minor NOR loci in the short arm of chromosome 1S^b and proximal to a major NOR locus in the short arm of chromosome 5S^b. Ae. bicornis differs from the other species of subsection Emarginats, with the exception of Ae. searsii, by having a lower heterochromatin content. In addition, Ae. bicornis differs from the other three Emarginata species in the size of the two 5S rDNA sites, and by the presence of a minor polymorphic NOR locus in a distal part of the long arm of chromosome 5S^b (Badaeva et al. 1996b).

The karyotype of Ae. bicornis differs from that of Ae. longissima, Ae. sharonensis, and Ae. searsii, in both its C-banding pattern and by the size of C-bands detected in the karyotype, all of which are small (Teho and Hutchinson 1983; Teho et al. 1983a; Badaeva et al. 1996a), and most resembled that of Amblyopyrum muticum (Friebe et al. 1996). Ae. longissima and Ae. sharonensis are highly heterochromatic species; C-bands are present in intercalary, telomeric, and proximal regions of the chromosomes, while Ae. searsii and Ae. bicornis have much less C-heterochromatin and have relatively smaller centromeric bands (Teoh and Hutchinson 1983; Badaeva et al. 1996a).

Strong hybridization with the noncoding, highly repetitive DNA sequence, pSc119, derived from Secale cereale, was observed at the telomeres and at some subtelomeric regions of several chromosomes (Badaeva et al. 1996a). A strong signal with the pScll9 probe was detected at both telomeres of chromosomes 1S^b and one polymorphic site was found in the subterminal region of the long arm of 1S^b. Chromosomes 2S^b, 3S^b, and 6S^b (a SAT chromosome) had similar labeling patterns with pSc119 but differed in chromosome morphology. The hybridization pattern of chromosome 4S^b included one telomeric site in the short arm and a double site in the long arm. An additional site was detected in the subterminal region of the long arm. Chromosome 5S^b, the second SAT chromosome, had one pSc119 site at the telomere of the short arm. Two telomeric pScll9 sites and one subterminal site were detected in the short arm of chromosome 7S^b. The distribution of pScll9 hybridization sites in the Ae. bicornis genome was like that of Ae. longissima, Ae. sharonensis, Ae. searsii and A. muticum.

Using FISH in Ae. bicornis, Salina et al. (2006) did not find any detectable FISH signal with the two tandem repeated sequences Spelt1 and Spelt52 probes. This confirms earlier studies that concluded that Spelt52 is absent in Ae. bicornis (and in Ae. searsii), while it occurs in Ae. sharonensis and Ae. longissima (Anamthawat-Jonsson and Heslop-Harrison 1993; Zhang et al. 2002; Salina et al. 2004a; Belyayev and Raskina 1998; Raskina et al. 2011). Ae. sharonensis and Ae. longissima have similar pSc119 labeling patterns as those seen in Ae. bicornis and Ae. searsii but differ from them in having more heterochromatic chromosomes (Badaeva et al. 1996a; Friebe and Gill 1996).

9.4.3.3 Crosses with Other Species of the Wheat Group

The chromosomes of Ae. bicornis pair relatively well with those of the other Sitopsis species, but only the F₁ hybrid with Ae. sharonensis was fertile (Tanaka 1955a). Tanaka (1955a) reported that in the F₁ hybrid of a biotype of Ae. sharonensis with one of Ae. bicornis, meiotic chromosome pairing was fairly regular, with six to seven bivalents. Pollen and seed fertility in this hybrid was high, even though the plants were dwarf-like and weak. Similarly, high chromosomal pairing was observed in meiosis of a hybrid between a biotype of Ae. longissima and Ae. bicornis (Table 9.4) indicated that these species differed by a reciprocal translocation and the hybrid was more or less self-sterile (Kimber 1961). However, a backcross to either parent set seed when the hybrid was used as the female parent (Waines and Johnson 1972). Kihara (1949) reported 5–7 bivalents (seven bivalents were the mode) in first meiotic metaphase of F₁ hybrids between Ae. speltoides and Ae. bicornis, however, the hybrids were sterile.

Few studies have been performed on crosses with other Aegilops species. Meiocytes of the F₁ hybrid between Ae. bicornis and Ae. tauschii exhibited from zero to six bivalents and a rare one trivalent (Kihara 1949). Those between Ae. bicornis and tetraploid Ae. crassa as well as with Ae. uniaristata, had similar levels of chromosomal pairing, i.e., one to six bivalents and few multivalents (Kihara 1949). The F₁ hybrid between Ae. bicornis and Ae. umbellulata, Ae. biuncialis and Ae. columnaris had an even lower level of pairing (0–5 with 3 bivalents as mode and few trivalents; Kihara 1949), while that with Ae. peregrina had higher pairing (Table 9.5), indicating that the genome of Ae. bicornis is related to one of the subgenomes of Ae. peregrina, presumably subgenome S^v.

Ohta (1990) produced F₁ hybrids between Ae. bicornis and Amblyopyrum muticum with and without B chromosomes (Tables 8.1 and 8.2). In studies of chromosomal pairing at first meiotic metaphase of the hybrids that did not have B chromosomes, he observed very little pairing (0.003 mean arm pairing per cell) whereas in hybrid with B chromosomes the pairing was significantly higher (0.77 mean arm pairing per cell; 82% of the cells had seven bivalents). No cells had multivalents and the frequency of univalents was very low. This high pairing in this hybrid presumably resulted from the activity of the homoeologous pairing promoters that exist in A. muticum rather than from the high degree of homology between the chromosomes of the two species.

Crosses between Ae. bicornis and Triticum monococcum ssp. monococcum was studied by Sears (1941b) (Table 9.8). At meiosis of this F₁ hybrid, 5.38 chromosomes paired on average, most of which in the form of rod bivalent. In other crosses between Ae. bicornis and T. monococcum ssp. aegilopoides, the offspring were inviable or poorly viable, whereas hybrid with another variety of ssp. aegilopoides was viable (Sears 1944a). This lack of viability is apparently mono-factorially determined (Sears 1944a). Poor pairing was observed in the triploid hybrids between wild tetraploid wheat, T. turgidum subsp. dicoccoides x Ae. bicornis, and T. timopheevii ssp. timopheevii and Ae. bicornis (Table 9.9), and between hexaploid wheat, T. aestivum subsp. aestivum and Ae. bicornis (Table 9.10). Evidently, very little homology exists between the chromosomes of diploid wheat and Ae. bicornis; the triploid and tetraploid hybrids had even less pairing, due to the presence of the Ph1 gene of the allopolyploid wheats.

9.4.4 Aegilops sharonensis Eig

9.4.4.1 Morphological and Geographical Notes

Aegilops sharonensis Eig, common name: Sharon goat grass, [Syn.: Aegilops bicornis (Forssk.) Jaub. & Spach var. major Eig; Triticum sharonnse (Eig) Feldman & Sears; T. longissimum (Schweinf. & Muschl.) Bowden ssp. sharonensis (Eig) Chennav.; Aegilops longissima Schweinf. & Muschl. ssp. sharonensis (Eig) Chennav.; Ae. longissima Schweinf. & Muschl. var. major (Eig) Hammer; Sitopsis sharonensis (Eig) Á. Löve] is a predominantly autogamous annual plant. Its culm is (40-) 50–70 (-100)-cm-high (excluding spikes). Spikes are more or less broad, linear, two-rowed, usually awned (except in var. mutica), and 7–13-cm-long (excluding awns). Rachis is zig-zagged, with each segment being bow-shaped, disarticulating to individual spikelets at maturity, each spikelet with the rachis internode immediately below it (wedge-type dispersal unit; Fig. 2.3). The lower-most spikelet or the few lowest spikelets remain attached at the tip of the culm. Spikelets are 8–13-mm-long (excluding awns), linear, elliptical, grow smaller toward the tip, and are more or less flattened. There are 3–5 florets per spikelet, with the upper 1–3 being sterile. The attached rachis internode is shorter than the spikelets and curved. Glumes are 6–7-mm-long, with two small points, one of which is sometimes elongated into a small awn. Lemmas are 8–11-mm-long, with a 40–60-mm-long awn, there are two short broad teeth at the base. Awns increase in length toward the tip of the ear. The 1–3 lowest spikelets are usually almost awnless. Anthers are 5–6-mm-long. The caryopsis adheres to the lemma and palea (Fig. 9.1d).

Since its discovery, the status of Ae. sharonensis as an independent species has been in dispute and has been changed on several occasions. Zhukovsky (1928) considered Ae. sharonensis as a subspecies of Ae. longissima, while Eig (1928a) described it as a variety of Ae. bicornis and named it Ae. bicornis (forssk.) Jaub. & Spach. var. major Eig. Later, Eig (1928b), based on a re-examination of the plant morphology, his advanced knowledge of the various habitats, and the absence of hybrid swarms or intermediate forms in the contact zones of Ae. sharonensis and Ae. bicornis, elevated the former to the species rank and named it Ae. sharonensis Eig. However, its validity as a biological species had since been doubted. Based on complete chromosome pairing of the F₁ hybrids at meiosis and their high fertility, Kihara (1937, 1940a, b, 1949, 1954), and Lilienfeld 1951) treated it as a subspecies or variety of Ae. longissima and assigned the two taxa the same genome formula (S^l). This treatment was supported by studies of the fertility and chromosome pairing in pollen mother cells (PMCs) of hybrids between these two taxa (Tanaka 1955a; Roy 1959; Kimber 1961). Also, Bowden (1959), while integrating the genus Aegilops into Triticum, considered Ae. sharonensis a subspecies of T. longissimum. Chennaveeraiah (1960), based on karyomorphology and plant morphology, also considered Ae. sharonensis to be a subspecies of Ae. longissima. The merging of Aegilops into Triticum was adopted by several cytogeneticists (e.g., Morris and Sears 1967; Mac Key 1968; Kimber and Sears 1983; Kimber and Feldman 1987), but these scientists considered T. sharonense as a separate species, named T. sharonense (Eig) Kimber and Feldman. While reorganizing the tribe Triticeae in genera based on genome homology, Löve (1984), recognized section Sitopsis as a genus including five diploid species bearing the S genome. Under this taxonomic treatment, Aegilops sharonensis was named Sitopsis sharonensis (Eig) A. löve. However, taxonomists (e.g., Baum 1977, 1978a, b; Hammer 1980; Feinbrun-Dothan 1986a; van Slageren 1994) kept Aegilops as a separate genus. Ankory and Zohary (1962) reported that the two species, Ae. sharonensis and Ae. longissima, are well-differentiated in nature and were apparently separated by seasonal and ecological factors for natural hybridization and, therefore, treated them as different species. Using the same arguments, Kimber and Feldman (1987) also treated them as separate species. Using C-banding pattern of mitotic metaphase chromosomes, Teoh and Hutchinson (1983) found that the differences between Ae. sharonensis and Ae. longissima are large enough to justify their treatment as two separate species. Likewise, studies of electrophoretic mobility of isoenzymes (Brody and Mendlinger 1980; Nakai and Tsuji 1984) and analysis of water-soluble leaf proteins (Mendlinger and Zohary 1995) showed Ae. sharonensis to be a valid species. Waines and Johnson (1972), studying the electrophoretic pattern of ethanol-extracted seed proteins, found that Ae. sharonensis was intermediate between Ae. longissima and Ae. bicornis and concluded that, Ae. sharonensis is genetically different from these two species and should be considered a separate species. Finally, Yen and Kimber (1990b) hybridized Ae. sharonensis with autotetraploid lines of Ae. speltoides, Ae. longissima and Ae. bicornis and analyzed chromosomal pairing at meiotic first metaphase of these hybrids. They found that Ae. sharonensis is almost equally related to Ae. speltoides and Ae. longissima, while it is distant from Ae. bicornis, and concluded, therefore, that Ae. sharonensis should not be treated as a subspecies or variety of Ae. longissima.

Ae. sharonensis includes two varieties: var. sharonensis (var. typica Eig), in which lemmas are awned in all spikelets except in the lowest, and var. mutica (Post) Eig, where lemmas are awnless in all spikelets or awned in the upper-most spikelets only. The latter is rare and occurs in mixed populations in Acre Plain and Sharon Plain of Israel.

Morphological variation involves differences in spike color and size, spikelet number and size, degree of expression of two-rowed nature and awn development. Studies on genetic and phenotypic diversity indicated that Ae. sharonensis is a diverse species, in spite of its limited geographic distribution and not highly variable environments (Olivera and Steffenson 2009). Morphologically, Ae. sharonensis is like to Ae. bicornis by having two-rowed spikes, awns in all the spikelets, spike disarticulation into single spikelets, and the shape of the lemmas (Eig 1929a), but have larger plant and spikelet sizes, larger grains and a somewhat laxer rachis. Edaphically, Ae. sharonensis is closer to Ae. bicornis than to Ae. longissima, though sympatric with the latter in several localities and allopatric with the former.

Olivera et al. (2010) used microsatellites to study genetic diversity and population structure of Ae. sharonensis from different sites in Israel and identified the Sharon Plain as the region exhibiting the highest level of allele richness and average gene diversity. Because it is located in the center of the geographic distribution of the species and includes sites with the highest level of diversity, the authors suggested that the Sharon Plain might be the center of origin and center of variation of Ae. sharonensis. There are two possibilities to explain the origin of Ae. sharonensis. First, it derived from Ae. bicornis that penetrated the coastal plain of southern Israel and absorbed genes from Ae. longissima via introgressive hybridization. Indeed, Waines and Johnson (1972), based on studies of seed protein patterns, proposed that, Ae. sharonensis derived from a hybridization between Ae. bicornis and Ae. longissima. Alternatively, Raskina et al. (2004b), based on studies of intrapopulation variability of rDNA in marginal populations of Ae. speltoides and Ae. sharonensis, suggested that Ae. sharonensis derived from Ae. speltoides.

Phenologically, Ae. sharonensis and Ae. bicornis are the earliest heading species of the genus Aegilops. Both species flower from March to May, albeit Ae. sharonensis starts flowering two weeks later than Ae. bicornis (Eig 1929a). Ae. speltoides, Ae. longissima and Ae. searsii head later; these species flower from April to June and in mountainous areas, even up to July (Feinbrun-Dothan 1986a; van Slageren 1994).

Ae. sharonensis is an east Mediterranean element. It has a very limited distribution in the south-central distribution region of the genus. It is endemic to the coastal plain of Israel (the Acre Plain, Sharon Plain, and Philistean Plain) and south Lebanon (Eig 1928b, 1929a, 1936; Millet 2006; Olivera and Steffenson 2009). The name Ae. sharonensis refers to the Sharon Pain in Israel, where this taxon was first described (Eig 1928a). It grows at 0–100 m above sea level, locally common, often in dense stands, on well-drained sandy soils, consolidated sand dunes, and on marine dilluvial rocks (kurkar rocks), in open park-, shrub-, and herbaceous-plant formations, abandoned fields, disturbed habitats and roadsides (Eig 1928b, 1929a; Post 1933; Ankori and Zohary 1962; Witcombe 1983; Kimber and Feldman 1987; van Slageren 1994). It grows sympatrically with Ae. longissima (in few sites in the Israeli Coastal Plain), where intermediate types between these two species in mixed populations, mainly in disturbed habitats in the Sharon Plain, were reported by Eig (1929a) and Ankori and Zohary (1962), and with Ae. peregrina And Ae. geniculata, and allopatrically with Ae. longissima (in most sites in the Sharon and Philistean Plains), Ae. bicornis, and Ae. speltoides. Aegilops sharonensis has a rich source of genes providing resistance to important wheat diseases and abiotic stresses (Olivera and Steffenson 2009 and references therein). Some forms grow in salt-marshes (e.g., Na’aman salt-marsh, north of Haifa) and may contain genes for salt tolerance. With the most limited distribution of any species in the genus Aegilops, Ae. sharonensis is rapidly losing its habitats, owing to the combined effects of modern agricultural intensification and expansion of urban and industrial areas (Millet 2006; Olivera and Steffenson 2009).

9.4.4.2 Cytology, Cytogenetics, and Evolution

Ae. sharonensis is diploid (2n = 2x = 14), having a modified S genome, designated S^l by Kihara (1954) and Dvorak (1998), or S^sh by Theo and Hutchinson (1983) (from here on, the genome symbol of Ae. sharonensis will be S^sh). Its organellar genome is quite similar to that of Ae. longissima, and consequently, is designated S^l (Ogihara and Tsunewaki 1988). According to Eilam et al. (2007), this species has the largest genome in comparison to all other diploid Aegilops species (1C DNA = 7.52 ± 0.100 pg; 7.1 pg; The Angiosperm C-value database at Kew Botanic gardens (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. sharonensis genome after genome sequencing and assembly to be 6.07 Gb (=5.93 pg), and Avni et al. (2022), also after sequencing and assembly, reported 6.71 Gb (=6.56 pg). Furuta et al. (1986) described the genome size of Ae. sharonensis as equal to that of Ae. longissima and larger than those of Ae. searsii and Ae. speltoides. The karyotype of Ae. sharonensis is symmetric; all chromosome pairs have submedian centromere (Chennaveeraiah 1960). Riley et al. (1958) and Chennaveeraiah (1960) found that the karyotype of Ae. sharonensis is similar to that of Ae. longissima and, in both species, there is one pair with large and another pair with distinctly small satellites. The smaller satellites are slightly less than half the size of the larger ones. The karyotypes of Ae. sharonensis and Ae. longissima are quite similar to each other except for minor differences in the arm ratios of the non-satellited pairs (Chennaveeraiah 1960).

Ae. sharonensis, like the other three species of subsection Emarginata, has major NOR loci on chromosomes of groups 5 and 6 and a variable number of minor loci on chromosomes of groups 1, 3, 5, and 6 (Badaeva et al. 1996b). The 5S rDNA loci were observed on chromosomes of groups 1 and 5, distal to minor NOR loci in the short arm of chromosome 1S^sh and proximal to a major NOR locus in the short arm of chromosome 5S^sh (Badaeva et al. 1996b).

Teoh and Hutchinson (1983) and Teoh et al. (1983) studied the C-banding patterns of chromosomes at mitotic metaphase in all 10 diploid Aegilops species and found that all diploid species exhibit characteristically different patterns that enable the chromosomes of any complement to be individually identified. Ae. sharonensis has a unique C-banding pattern with telomeric, interstitial and exceptionally distinctive centromeric bands, varying in size and staining intensity. Hence, from the specific C-banding pattern, Teoh and Hutchinson (1983) concluded that the genome of Ae. sharonensis is different from that of Ae. longissima, and, consequently, gave it the symbol S^sh. The C-banding pattern of Ae. sharonensis indicates that this species is much more closely related to Ae. speltoides and Ae. longissima than to Ae. bicornis and Ae. searsii.

A repetitive DNA sequence thought to be a noncoding, highly repeated, 260-bp DNA fragment derived from the B subgenome of bread wheat (Hutchinson and Lonsdale 1982), was used in FISH experiments on genomes of all the diploid species of Aegilops (Teoh et al. 1983). This repetitive sequence was found in variable amounts in all diploid species but was restricted to specific regions of the chromosomes. Ae. sharonensis, Ae. speltoides and Ae. longissima possess many copies of the sequence and its distribution is correlated with their respective C-banding patterns. The strongest labelling was observed in Ae. longissima and Ae. sharonensis, both showing localization near the centromeres and also interstitially along the chromosomes.

Salina et al. (2006), using two telomere-associated tandem repeat sequences of Ae. speltoides, Spelt1 and Spelt52, found that the in-situ hybridization patterns of these probes in Ae. speltoides, Ae. longissima and Ae. sharonensis are different. The FISH signal with Spelt1 was only observed in Ae. speltoides, while the FISH signal with Spelt52 was found, in addition to Ae. speltoides, in Ae. sharonensis and Ae. longissima as well. Similar results were obtained by Anamthawat-Jonsson and Heslop-Harrison (1993), Zhang et al. (2002), Salina et al. (2004b) and Raskina et al. (2011). Thus, Spelt1 was completely absent from the genomes of the four Emarginata species, Ae. sharonensis, Ae. longissima, Ae. searsii and Ae. bicornis.

Badaeva et al. (1996a), using C-banding and in situ hybridization with two highly repetitive DNA sequences, pSc119, and pAsl, found that Ae. sharonensis was, together with Ae. speltoides and Ae. longissima, the very heterochromatic species of the diploid Aegilops species. While they did not observed hybridization with the pAsl probe on Ae. sharonensis, Ae. speltoides or Ae. longissima chromosomes, Ae. sharonensis, as well as the other Emarginata species (Ae. longissima, Ae. searsii and Ae. bicornis), showed strong labeling with the pSc119 probe in the telomeric chromosomal regions. Yet, the karyotypes of these species exhibited different distributions of C-heterochromatin (Badaeva et al. 1996b). Ae. sharonensis, like Ae. longissima, was highly heterochromatic; C-bands were present in intercalary, telomeric, and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis had much fewer C-bands in these regions.

Yaakov et al. (2013) utilized quantitative real-time PCR to assess the relative copy numbers of 16 TE element families in various Triticum and Aegilops species. They found that the Latidu family of TEs showed specific proliferation in Ae. sharonensis, with more Latidu than in Ae. speltoides and Ae. longissima, and much more than in Ae. searsii. The Rong family was also observed in relatively high quantities in Ae. sharonensis.

9.4.4.3 Crosses with Other Species of the Wheat Group

Crosses between Ae. sharonensis and three other Sitopsis species, i.e., Ae. longissima, Ae. bicornis and Ae. speltoides, produced F₁ hybrids that exhibited complete or almost complete chromosomal pairing at first meiotic metaphase. Thus, F₁ hybrids between Ae. sharonensis and Ae. longissima and their reciprocal hybrid, had 5 bivalents (mostly ring bivalents) and a quadrivalent (or trivalent and univalent), indicating complete homology between the genomes of these two species and the existence of a reciprocal translocation between them (Tanaka 1955a; Kimber 1961; Ankori and Zohary 1962). Only Roy (1959) observed 6.90 bivalent and no multivalents in these hybrids. However, when assessing the photographs presented by Roy, Ankori and Zohary (1962) noticed that the longissima line used was already highly introgressed with Ae. sharonensis, which explains why Roy did not encounter a translocation configuration in his hybrids. Hybrid between Ae. sharonensis and Ae. bicornis exhibited 6–7 bivalents (Tanaka 1955a) and the hybrid between Ae. sharonensis and Ae. speltoides had similar levels of pairing (6.85 bivalents per cell of which 5.35 were ring bivalents; Kihara 1949; Sears 1941b; Kimber 1961), whereas Tanaka (1955a) reported on somewhat less pairing in this hybrid (4–7 bivalents). The F₁ hybrids between Ae. sharonensis and Ae. longissima was almost fully fertile (more than 80% seed set; Tanaka 1955a; Ankori and Zohary 1962) and the hybrid Ae. sharonensis x Ae. bicornis had an approximate 62% seed set (Tanaka 1955a), while the hybrid between Ae. sharonensis and Ae. speltoides was sterile (Sears 1941b).

Data of chromosome pairing in F₁ hybrids between Ae. sharonensis and other diploid Aegilops species are presented in Table 9.4. These hybrids exhibited low pairing, indicating that the genome of Ae. sharonensis is more distant from the genomes of these species than from those of Sitopsis species. The F₁ hybrid between the tetraploid species Ae. kotschyi (genome S^vS^vUU) and Ae. sharonensis exhibited 4.28 bivalents (of which 2 were ring), 1.50 trivalents, 0.46 quadrivalents and 0.09 pentavalents (Rubenstein and Sallee 1973), indicating high homology between the genome of Ae. sharonensis and one of the subgenomes of Ae. kotschyi, most probably genome S^v.

In analyzing F₁ hybrids between Ae. sharonensis and Ambliopyrum muticum without B chromosomes, Ohta (1990) found chromosome pairing at first meiotic metaphase ranging from 6.22 to 6.50 bivalents (of which 3.10 to 4.48 were ring bivalents) (Table 8.1). Several hybrids had few multivalent configurations. These hybrids showed very good pairing, presumably due to the promotion of homoeologous pairing by the genome of A. muticum but were completely sterile (Ohta 1990).

Ae. sharonensis was crossed with diploid, tetraploid, and hexaploid Triticum species (Tables 9.8, 9.9 and 9.10). The F₁ hybrid between Ae. sharonensis and T. monococcum ssp. monococcum had very little pairing and were completely sterile (Kushnir amd Halloran 1981), indicating little homology between the S^sh and A^m genomes. The F₁ hybrid between Ae. sharonensis and tetraploid wild wheat T. turgidum ssp. dicoccoides had 2.23 bivalents (of which 0.03 were ring bivalents) (Kushnir and Halloran 1981), whereas McFadden and Sears (1947) reported 5.18 bivalents and 1.05 trivalents in the reciprocal hybrid. Hybrids between domesticated tetraploid wheat T. turgidum subsp. turgidum and Ae. sharonensis exhibited 2.22 bivalents and 0.02 trivalents (Riley et al. 1958). A similar level of chromosomal pairing in this hybrid combination, i.e., 2.50 bivalents per cell, was reported by Roy (1959). These F₁ triploid hybrids had very little chromosome pairing at meiosis, indicating that, in the presence of the homoeologous-pairing suppressor, Ph1, of allopolyploid wheats, Ae. sharonensis chromosomes rarely pair with the wheat chromosomes and, likewise, the wheat chromosomes rarely pair with each other. Similar low chromosomal pairing exists in F₁ hybrids between hexaploid wheat T. aestivum ssp. aestivum and Ae. sharonensis (Table 9.10). Interestingly, the amount of chromosome pairing in hybrids with hexaploid wheat is lower than that measured in hybrids with tetraploid wheat.

The homoeologous relationship between Ae. sharonensis and chromosomes of T. aestivum was determined by Friebe and Gill (1996), by comparing the C-banding pattern of Ae. sharonensis chromosomes, described by Teoh and Hutchinson (1983), with those of Ae. longissima chromosomes, whose homoeologous relationships to T. aestivum chromosomes were previously established by Friebe et al. (1993). Then, Friebe and Gill (1996) assigned the seven chromosomes of Ae. sharonensis to each homoeologous group of T. aestivum. Later, Maestra and Naranjo (1997) confirmed the homoeologous relationship between Ae. sharonensis and T. aestivum chromosomes, by C-banding analysis of specific pairing at first meiotic metaphase in F₁ hybrids between Ae. sharonensis and T. aestivum ssp. aestivum. They analyzed chromosomal pairing in hybrids between three different genotypes of Triticum aestivum, each carrying (Ph1), or lacking it (ph2b mutant), and Ae. sharonensis, in order to establish the homoeologous relationships of Ae. sharonensis chromosomes to those of hexaploid wheat. Since Ae. sharonensis chromosomes show a distinctive C-banding pattern (Teoh and Hutchinson 1983), and thus, could be distinguished from those of wheat, C-banding was used by Maestra and Naranjo (1997) to identify the chromosomes of both species and their arms. Normal homoeologous relationships for the seven chromosomes of the S^sh genome of Ae sharonensis, and their arms, were revealed in this study. The pattern of pairing between chromosomes of Ae. sharonensis and ssp. aestivum indicated that no apparent chromosome rearrangement occurred during the evolution of the Ae. sharonensis genome relative to the subgenomes of hexaploid wheat. Thus, the chromosome structure of the ancestral genome from which subgenomes A, B, and D of allohexaploid wheat derived, was also preserved in genome S^sh. All three types of hybrids with Ph1, demonstrating low pairing level, however, with ph2b, demonstrating intermediate pairing level was observed, showing preferential pairing between A-D and B-S^sh. A close relationship between the S^sh genome and the B subgenome of bread wheat was confirmed, but the results provided no evidence that the B subgenome was derived from Ae. sharonensis. Similar results were obtained by Fernández-Calvín and Orellana (1993).

On the basis of the results of homoeologous pairing between chromosomes of Ae. sharonensis and T. aestivum, Maestra and Naranjo (1997) were able to identify the homoeologous relationships between all of the chromosomes of the S^sh genome and bread wheat. Their homoeologous pairing findings confirmed the homoeology of chromosome 4S^sh to wheat chromosomes of group 4, reported by Miller et al. (1982), as well as the assignment of the S^sh genome chromosomes to the seven homoeologous groups of wheat, and the arm designation, suggested by Friebe and Gill (1996), based on chromosomal morphology and C-banding.

9.4.5 Aegilops longissima Schweinf. & Muschl.

9.4.5.1 Morphological and Geographical Notes

Aegilops longissima Schweinf. & Muschl. [Syn.: Aegilops longissima (Schweinf. & Muschl.) ssp. longissima Hammer; Triticum longissimum (Schweinf. & Muschl.) Bowden; Sitopsis longissima (Schweinf. & Muschl.) Á. Löve] is a predominantly autogamous annual plant. Its culm is 40–110-cm-high (excluding spikes). Spike is narrow, linear, one-rowed, tapering slightly to the tip, and 10–20-cm-long (excluding awns). At maturity, they are fragile near, but usually not at, the base, with the greater part of the spike falling entire, while lower spikelets remain on the culm or fall later singly or in 2–3 pieces. Spikelets 8–15 are 12–14-mm-long (excluding awns), become thinner and shorter toward the tip of the spike, and are appressed to the rachis segment. There are 3–5 florets in each spikelet, the upper 1–3 being sterile. The rachis internodes are nearly as long as spikelets in the middle of the spike. Glumes are 6–8-mm-long, tough, and usually with two teeth separated by a membranous edge. Glumes of the terminal spikelet sometimes have three teeth, with the center one sometimes elongated into a very short awn. Lemmas of the lateral spikelets are canoe-shaped and awnless. Lemmas of the terminal spikelet have a 7–12-cm-long, broad, convex awn, often with a small unequal tooth on each side. Anthers are 5–6-mm-long. The caryopsis adheres to both lemma and palea (Fig. 9.1e).

Limited morphological variation is mainly seen in spike length, spikelet number, rachis form (zig-zagged or straight) and awn length. Like Ae. speltoides and Ae. searsii, Ae. longissima ripens later than Ae. sharonensis and Ae. bicornis. The center of variation is in steppic regions of Israel and Jordan.

Morphologically, Ae. longissima is taller than Ae. sharonensis and differs from it by its long, narrow, linear, and one-rowed spike, by the absence of awns on the lemmas of the lateral spikelets, and by the presence of long and broad awns on the terminal spikelet. At maturity, the spike breaks near the base and falls almost entire and, at the end of the summer, the spike may disarticulate into single spikelets with the rachis internode beside them (barrel-type disarticulation). Ankori and Zohary (1962) reported that the two species are well-differentiated in nature and apparently separated by seasonal and ecological factors for natural hybridization. Cytologically, the two taxa have minor differences in their karyotype, and even though the hybrid between them exhibits complete chromosomal pairing at meiosis, the two taxa differ by a reciprocal translocation. The occurrence of a reciprocal translocation between the distal regions of chromosome arms 4S^lL and 7S^lL in Ae. longissima was later confirmed by Friebe et al. (1993). This translocation is not present in other Sitopsis species (Kihara 1949; Tanaka 1955a; Riley et al. 1961; Kimber 1961; Feldman et al. 1979).

Ae. longissima has a relatively limited distribution in the south-central region of the distribution area of the genus. A steppical (Irano-Turanian) element extends into sub-Mediterranean and desert (Saharo-Arabian) regions. It grows in Egypt (lower Egypt and Sinai), Israel (coastal plain, northern, and central Negev and eastern Judea and Samarian Mountains up to the east to Ein Gev, the eastern shore of the Lake of Galilee), Jordan, Lebanon (southern coastal plain), and Syria. Alt: 0–900 m above sea level (Table 9.1). It grows on sandy loams derived from the sandstone of the coastal plain, rarely on somewhat heavier soil, in the coastal plain of Egypt, Israel and Lebanon, and on sand derived from Nubian sandstone, in grey calcareous steppe soil or loess in the inland steppe or desert regions, in open dwarf shrub or herbaceous steppe-like or desert-like formations, plains, abandoned fields, edges of cultivation and roadsides. It is common, and often abundant in the coastal plain and in several steppic habitats bordering on the Mediterranean region in Israel, Jordan, and Syria.

Like Ae. bicornis, it grows in the hot and dry parts of the south-east Mediterranean, but its distribution area is closer to the center of the distribution area of the genus. The center of variation is in the steppic regions of Israel and Jordan, where it presumably originated and from which it invaded the coastal plain. It is sympatric with Ae. sharonensis [hybrid swarms with Ae. sharonensis can be found in the coastal plain (Ankori and Zohary 1962)] and Ae. peregrina in the coastal plain and with Ae., bicornis, Ae. kotschyi and Ae. vavilovii in the steppical region. It is allopatric with Ae. speltoides in the coastal plain, with Ae. searsii in southern Judea Mts., with Ae. umbellulata, Ae. geniculata and Ae. biuncialis in the sub-Mediterranean area in Lebanon and Syria and with Ae. crassa in the steppical regions of Syria.

9.4.5.2 Cytology, Cytogenetics, and Evolution

Ae. longissima is diploid (2n = 2x = 14) with a modified S genome, designated S^l by Kihara (1949, 1954, 1963, 1970). Its organellar genome was designated S^l2, as a subtype of the S^l plasmon (Wang et al. 1997; Table 9.3. This species has the second largest genome among diploid Aegilops species (1C DNA = 7.48 ± 0.082 pg) (Eilam et al. 2007), 6.0 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. longissima genome after genome sequencing and assembly to be somewhat smaller amount, i.e., 6.22 Gb (=6.07 pg), and Avni et al. (2022), also after sequencing and assembly, reported 6.70 Gb (=6.55 pg). Furuta et al. (1986) reported that the genome size of Ae. longissima is equal to that of Ae. sharonensis but larger than those of Ae. searsii and Ae. speltoides.

The karyotype of Ae. longissima is symmetric; the satellite chromosomes have median centromeres, while all the non-satellited chromosomes have sub-median centromeres (Senyaninova-Korchagina 1932; Chennaveeraiah 1960). Riley et al. (1958) reported that the karyotypes in Ae. longissima and Ae. sharonensis are similar, and that in both species, there is one pair with large and another with distinctly small satellites. These findings were confirmed by Chennaveeraiah (1960), who stated that the smaller satellites are slightly less than half the size of the larger ones, and that there are minor differences in in the karyotypes of Ae. longissima and Ae. sharonensis in the arm ratios of the non-satellited pairs.

Based on the presence of secondary constrictions (Chennaveeraiah 1960; Chen and Gill 1983; Teoh and Hutchinson 1983), in situ hybridization analysis using a radioactive rDNA probe (Miller et al. 1983; Teoh et al. 1983), and analysis of nucleolar activity by Ag-NOR banding (Cermeño et al. 1984a, b), two pairs of NORs were identified in the secondary constrictions of Ae. longissima. Yet, Friebe et al. (1993) and Badaeva et al. (1996b), using in situ hybridization with a 18S–26S rDNA probe, which enabled high sensitivity, detected, in addition to the two major-active NORs on the short arms of chromosomes 5S¹ and 6S¹, an additional minor NOR on chromosomes 1S^l, and a polymorphic minor NOR on chromosome 3S^l. The minor NORs usually do not form secondary constriction, indicating that these NORs are not transcribed. The 5S rDNA loci were detected in Ae. longissima on chromosomes 1S^l and 5S^l, distal to minor NOR loci in the short arm of chromosome 1S^l and proximal to a major NOR locus in the short arm of chromosomes 5S^l (Badaeva et al. 1996b).

In situ hybridization (ISH) patterns with both pTa7l (rDNA) and pTa794 (5S rDNA) probes revealed similar distribution of the two probes in Ae. longissima and Ae. sharonensis (Badaeva et al. 1996a). However, while ISH patterns were not polymorphic in Ae. sharonensis, intraspecific polymorphism was found in Ae. longissima. On the other hand, Ae. longissima chromosomes were similar to those of Ae. searsii in hybridization patterns with both probes (Badaeva et al. 1996a). This confirms the high degree of similarity of their genomes, as first observed by the distribution of highly repetitive DNA sequences (Teoh et al. 1983; Badaeva et al. 1996b). Aegilops bicornis differed from Ae. longissima by its lower heterochromatin content, differences in size between the two 5S rDNA sites, and the presence of a minor polymorphic NOR locus in a distal part of the long arm of chromosome 5S^b (Badaeva et al. 1996b).

The C-banding patterns of Ae. longissima chromosomes were determined at mitotic metaphase by Teoh and Hutchinson (1983), who showed that all seven pairs of Ae. longissima chromosomes can be identified by their characteristic C-banding patterns. Like all other Sitopsis species, Ae. longissima also had telomeric, interstitial and centromeric bands, that varied in size and staining intensity. The proximal and telomeric bands were smaller than the centromeric ones (Teoh and Hutchinson 1983).

The karyotypes of Ae. longissima, Ae. sharonensis, Ae. searsii and Ae. bicornis have different C-banding patterns (Badaeva et al. 1996a). Ae. 1ongissima and Ae. sharonensis are highly heterochromatic species; C-bands are present in intercalary, telomeric, and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis have much less C-heterochromatin compared with the two other Emarginata species.

Teoh et al. (1983) used a noncoding repetitive sequence, derived from T. aestivum, as a probe in in situ hybridization experiments on the genome of diploid Aegilops species. Ae. longissima, like Ae. speltoides and Ae. sharonensis, possesses many more copies of the noncoding sequence than Ae. bicornis and Ae. searsii. The distribution of the copies in each species correlated with their respective C-banding patterns.

Friebe et al. (1993) analyzed C-banding polymorphism in 17 accessions of Ae. longissima from Israel and Jordan and established a generalized idiogram of this species. Polymorphism for C-band size and C-band location was observed between different accessions but did not prevent chromosome identification. The C-banding patterns of Ae. longissima chromosomes they reported were similar to the N- and C-banding patterns reported earlier for this species (Chen and Gill 1983; Jewell and Driscoll 1983; Teoh and Hutchinson 1983; Kota and Dvorak 1985; Hueros et al. 1991). However, in most earlier reports, only one accession was analyzed and, therefore, no data were available on C-band polymorphisms in this species. Moreover, not all Ae. longissima chromosomes assigned correctly according to their homoeologous groups (Friebe et al. 1993).

A complete series of Ae. longissima addition and substitution lines were produced in the T. aestivum background (Hart and Tullen 1983). Sporophytic and gametophytic compensation tests were used to determine the homoeologous relationships of Ae. longissima chromosomes (Friebe et al. 1993). All Ae. longissima chromosomes compensated rather well and fertility was restored even in substitution lines involving wheat chromosomes 2A, 4B and 6B that contain major fertility genes.

The homoeologous relationships between Ae. longissima and wheat were determined for several chromosomes by their ability to compensate for the absence of wheat chromosomes in substitution lines (Jewell and Driscoll 1983; Kota and Dvorak 1985) and by all the chromosomes by C-banding analysis of Ae. longissima chromosomes in disomic and ditelosomic addition lines of Ae. longissima to T. aestivum and in substitution lines of Ae. longissima chromosomes for their T. aestivum homoeologues (Friebe et al. 1993).

The homoeologous relationships between Ae. longissima and wheat chromosomes have been established earlier by isozyme, storage protein, and morphological markers, as well as by analyzing their compensating ability in substitution lines (Hart and Tuleen 1983; Jewell and Driscoll 1983; Netzle and Zeller 1984; Kota and Dvorak 1985; Levy et al 1985; Millet et al. 1988; Hueros et al. 1991). The results obtained by Friebe et al. (1993) largely agreed with those reported previously. However, there were some discrepancies caused by chromosome misidentification in earlier studies.

C-banding patterns and morphology of Ae. longissima chromosomes 1S^l, 3S^l and 5S^l are very similar to those of chromosomes 1B, 3B and 5B of T. aestivum (Gill et al. 1991a). The C-banding pattern of the short arm of 4S^l is almost identical to that of the short arm of wheat chromosome 4B. The remaining Ae. longissima chromosomes showed differences in C-banding patterns and arm ratio with their wheat homoeologues.

Salina et al. (2006) and Raskina et al. (2001), using fluorescence in situ hybridization (FISH), observed considerable polymorphisms in the hybridization patterns of two tandemly repeated sequences, Spelt1 and Spelt52, among Aegilops species. While there was no detectable Spelt1 FISH signal in any species of section Sitopsis, except for Ae. speltoides, hybridization patterns of Spelt52 were species-specific in Ae. speltoides, Ae. longissima and Ae. sharonensis (Salina et al. 2006). Two very small and dim Spelt52 blocks were detected in Ae. searsii, whereas Ae. bicornis did not contain any Spelt52 repeats in its genome (Raskina et al. 2011). On the other hand, Ae. longissima and Ae. sharonensis showed that probe pSc119ƒffƒ (from Secale cereale) labeling patterns similar to Ae. bicornis and Ae. searsii but differed from them by having more heterochromatic chromosomes (Badaeva et al. 1996a; Friebe and Gill 1996).

Badaeva et al. (1996a), using both in situ hybridization (ISH) with the highly repetitive DNA sequences pSc119, from Secale cereale, and pAsl, from Ae. tauschii, as probes, and C-banding, analyzed genome differentiation in all diploid Aegilops species. The level of hybridization and labeling patterns differed among genomes. All five Sitopsis species had different C-banding pattern but they were similar to each other and to Amblyopyrum muticum in the distribution of pSc119 hybridization sites. On the other hand, no hybridization was observed with the pAsl probe on Ae. speltoides, Ae. sharonensis and Ae. longissima chromosomes, whereas a few minor pAsl sites were observed in Ae. searsii and Amblyopyrum muticum (Badaeva et al. 1996a). The ISH Patterns showed only minor intraspecific variations. Ae. bicornis and A. muticum had a low amount of C-heterochromatin, whereas Ae. longissima and Ae. sharonensis were the most heterochromatic species, and Ae. searsii possessed intermediate amounts of C-heterochromatin (Badaeva et al. 1996a). Ae. longissima, like the other three species of subsection Emarginata, Ae. sharonensis, Ae. searsii, and Ae. bicornis, had strong labeling in the telomeric chromosomal regions with the pSc119 probe (Badaeva et al. 1996a).

The C-banding pattern of the A. muticum genome chromosomes was different from that of the chromosomes of Ae. speltoides, and most resembled those of Ae. bicornis (Friebe et al. 1996). Distribution of pScll9 hybridization sites in the A. muticum genome also differed from that of Ae. speltoides but was similar to those of Ae. longissima, Ae. sharonensis, Ae. searsii and Ae. bicornis. Yet, the A. muticum genome was easily morphologically differentiated from the genome of the Sitopsis species.

During its evolution, Ae. longissima suffered a translocation involving the long arm of chromosome 4S^l and the long arm of chromosome 7S^l (Hart and Tuleen 1983; Friebe et al. 1993; Naranjo 1995). Analysis of chromosomal pairing at first meiotic metaphase showed a quadrivalent or a trivalent plus univalent in F₁ hybrids between Ae. longissima and all other members of Sitopsis, indicating that Ae. longissima differs from the other four species by a reciprocal translocation (Feldman et al. 1979 and references therein). Since F₁ hybrids between these four species form only bivalents at meiosis, it is reasonable to conclude that all four Sitopsis species, except Ae. longissima, have the ancestral chromosome structure.

Ae. longissima is probably involved in the parentage of the allotetraploid species Ae. peregrina (genome S^lS^lUU) and Ae. kotschyi (genome S^lS^lUU) (Kihara 1954; Feldman 1963; Kimber and Feldman 1987; Kimber and Sears 1987; Yen and Kimber 1989; Zhang et al. 1992; Dvorak 1998). Ae. longissima is a good source for genes conferring resistance to mildew, leaf and stem rust, heat and drought tolerance, and determination of high grain protein.

Some forms of Ae. longissima have genetic mechanisms capable of partially suppressing the Ph1 gene of polyploid wheat in hybrids with T. aestivum and T. turgidum. Mello-Sampayo (1971b) reported that a line of Ae. longissima promoted homoeologous pairing in meiosis of the F₁ hybrid with T. aestivum, thereby, partially counteracting the suppressive effect of wheat-Ph1. This line was designated an intermediate-pairing (IP) line to distinguish it from the more common low-pairing (LP) lines. Upadhya and Swaminathan (1967) reported on another IP line of Ae. longissima that promoted pairing in hybrids with common wheat. In studies of chromosome pairing in F₁ hybrids between ditelosomic lines of T. aestivum cv. Chinese Spring and the IP line of Ae. longissima, discovered by Mello-Sampayo (1971b), an intermediate amount of pairing was observed (5.49 bivalents, 0.39 trivalents and 0.06 quadrivalents per pollen mother cells (PMCs), most of which was between chromosomes of the wheat B subgenome and those of Ae. longissima (Feldman 1978).

To assess the effect of LP and IP genotypes on homologous pairing, Avivi (1976) produced autotetraploids from both lines of Ae. longissima and followed chromosomal pairing at first metaphase of meiosis. While the two induced autotetraploids did not differ in chiasma frequency or in the number of paired chromosomal arms, they differed significantly in multivalent frequency; the IP autotetraploid exhibited the same multivalent frequency as that expected on the basis of random pairing between the four homologues, namely, 4.7 multivalents per/cell. In contrast, the LP autotetraploid exhibited a significantly lower than expected frequency of multivalents. Avivi (1976) assumed that the LP genotype in the autotetraploid does not affect meiotic pairing per se but modifies the pattern of homologous association by separating the four sets of homologous chromosomes in somatic and premeiotic cells into two groups of two. In contrast, the IP genotype does not affect the spatial arrangement of the chromosomes. Accordingly, the gene Ph1 of common wheat suppresses homoeologous pairing in the aestivum x longissima hybrid by separating the wheat genomes from that of the longissima genome, thereby preventing pairing of distantly located chromosomes, the IP genotype does not do it and, consequently, enables some pairing between common wheat and longissima chromosomes (Avivi 1976).

9.4.5.3 Crosses with Other Species of the Wheat Group

Several intraspecific crosses were performed between lines of Ae. longissima from different habitats and geographical regions (Feldman et al. 1979 and Feldman M, unpublished). All the hybrids showed seven bivalents, most of which were ring bivalents. Seed set was normal except in one combination between a line from the Israeli Negev (steppe area) and a line from the Israeli coastal plain (Mediterranean region), in which seed fertility was low (53%) (Feldman et al. 1979).

Crosses between Ae. longissima and all of the other four Sitopsis species, i.e., Ae. sharonensis, Ae. bicornis, Ae. searsii and Ae. speltoides, produced F₁ hybrids that exhibited complete or almost complete chromosomal pairing at first meiotic metaphase (Table 9.4). Thus, F₁ hybrids between Ae. longissima and Ae. sharonensis and the reciprocal hybrids exhibited almost complete chromosome pairing at first meiotic metaphase, with 5 bivalents, one quadrivalent or trivalent and univalent, indicating the existence of high homology and the presence of a reciprocal translocation between these two species (Tanaka 1955a; Kimber 1961; Ankori and Zohary 1962). The F₁ hybrids are almost fully fertile, with an 82–86% seed set (Tanaka 1955a). Similarly, high pairing indicating great chromosomal homology and the presence of a reciprocal translocation was observed in all the hybrids of Ae. longissima and the other four Sitopsis species (Table 9.4). Nevertheless, the F₁ hybrids involving Ae. longissima and the Emarginata species, namely, Ae. sharonensis, Ae. bicornis and Ae. searsii, were highly or only partially fertile. In contrast, the F₁ hybrid Ae. longissima x Ae. speltoides was completely sterile in contrast to the high pairing (Riley et al. (1961), implying that the chromosomes of the two species differ by cryptic structural hybridity or by genetic barriers. Thus, Ae. longissima diverges from the other Sitopsis species by a translocation and is totally isolated from Ae. speltoides by partial internal chromosomal or genetic barriers which are superimposed upon predominant self-fertilization and ecological specialization of these two species. Since all intraspecific hybrids of Ae. longissima studied had seven bivalents, i.e., they were homozygous for the translocation, it is reasonable to assume that the translocation originated during the formation of this species.

Kihara (1949) crossed Ae. longissima with Ae. caudata, Ae. comosa, Ae. uniaristata and Ae. umbellulata analyzed chromosome pairing in the F₁ hybrids and in the reciprocal combinations. In longissima x caudata Kihara observed 3–6 bivalents and 0–2 trivalents, in longissima x comosa up to 6 bivalents and 0–1 trivalents, in longissima x uniaristata 2–5 bivalents and in Ae. longissima x Ae. umbellulata 0–4 bivalents (with mode of 2) and 0–1 trivalent These hybrids were completely sterile. These findings indicate that these four-specie are not close to Ae. longissima and have diverged quite considerably from it.

Chromosome pairing in F₁ hybrids between Ae. longissima and Amblyopyrum muticum (without B chromosomes) was relatively high (Tables 8.1 and 8.2). These hybrids showed very good pairing, presumably due to the promotion of homoeologous pairing by the genome of A. muticum, but were completely sterile (Ohta 1990). The presence of multivalents indicated that the two species differ by a reciprocal translocation. There was a slight difference in the amount of pairing between these two hybrids. Since the same accession of A. muticum was used in the two hybrids, Ohta (1990) suggested that the frequency and configuration of chromosome pairing observed in these two hybrid plants might be caused by small differences in the chromosomal structure or in the genotypes affecting homoeologous chromosome pairing between the two accessions of Ae. longissima. In hybrids Ae. longissima x A. muticum with B chromosome pairing was considerably reduced (Table 8.2).

Data from analyses of chromosomal pairing at first meiotic metaphase in F₁ triploid hybrids between Ae. longissima and a number of allotetraploid species of section Aegilops are presented below. Kihara (1949) reported the presence of 6–8 bivalents (with mode of 7) and 0–1 trivalents in the F₁ Ae. peregrina x Ae. longissima. Similar data were also reported by other researchers (Table 9.5) indicating that one of the two subgenomes of Ae. peregrina is homologous to the genome of Ae. longissima. Similar results were observed in the F₁ triploid Ae. kotschyi x Ae. longissima (Table 9.5). Chromosomal pairing in the F₁ Ae. biuncialis x Ae. longissima exhibited 0–5 bivalents and 0–2 trivalents, and the F₁ Ae. geniculata x Ae. longissima hybrid had 2–6 bivalents (with mode of 4), and 0–1 trivalents (Kihara 1949). Evidently, there is reduced homology between the genomes of these species.

Several crosses were performed between Ae. longissima and diploid, tetraploid and hexaploid Triticum species (9.8, 9.9 and 9.10). Kihara (1949) studied chromosome pairing in the F₁ Ae. longissima x T. monococcum ssp. monococcum and found 0–3 bivalents at meiosis. Feldman (1978) reported 1.49 bivalents, of which 0.02 were ring bivalents, and 0.07 trivalents in F₁ longissima x ssp. monococcum hybrid. These data indicate that the two species have diverged quite considerably from one another.

Chromosome pairing was analyzed in several Ae. longissima x T. turgidum hybrids (Table 9.9). In all these hybrids pairing was very low, indicating that very little homology exists between the chromosomes of Ae. longissima and those of T. turgidum. Kihara (1949) also crossed Ae. longissima with the second tetraploid species of wheat, T. timopheevii, and also found very little pairing in this hybrid (0–7 bivalents and 0.01 trivalents), indicating that the chromosomes of this tetraploid species are not homologous to those of Ae. longissima.

Table 9.1 in McFadden and Sears (1947) includes data on chromosomal pairing in the F₁ hybrid between Triticum turgidum subsp. dicoccoides and Aegilops sharonensis, which showed the formation of 5.18 bivalents 0–3 multivalents. But, on page 1016 of their article Fig. 9.2i is showing Ae. longissima and not Ae. sharonensis. Therefore, the pairing data in Table 9.1 is between subsp. dicoccoides and Ae. longissima, and, judging from the level of pairing in this hybrid, as well as in hybrids between T. aestivum and intermediate-pairing (IP) type Ae. longissima (Mello-Sampayo 1971b; Feldman 1978), it is most likely that the longissima parent in the McFadden and Sears (1947) paper is an IP type of Ae. longissima, and not of Ae. sharonensis.

Fig. 9.2

Aegilops species carrying the D genome; a A. spike of Ae. tauschii Coss. ssp. tauschii (formerly ssp. eusquarrosa Eig); b A spike of Ae. tauschii Coss. ssp. strangulata (Eig) Tzvelev; c A spike of Ae. ventricosa Tausch; d A plant and a spike of 4x Ae. crassa Boiss.; e A spike of Ae. vavilovii (Zhuk.) Chennav.; f A spike of Ae. juvenalis (Thell.) Eig; g A spike of Ae. cylindrica Host

Several crosses were performed between low-pairing (LP) type and IP type Ae. longissima and hexaploid wheat, T. aestivum. Hybrids involving LP types of Ae. longissima yielded very low pairing, with 1.45–1.96 bivalents and 0.01–0.08 trivalents (Riley et al. 1961; Riley and Chapman 1963; Riley 1966a; Ceoloni et al. 1986; Yu and Jahier 1992; Naranjo and Maestra 1995). Chromosomal pairing in these hybrids also showed little homology between the genomes of Ae. longissima and those of T. aestivum. On the other hand, hybrids involving IP types of Ae. longissima, that contains homoeologous pairing promoter(s), exhibited higher chromosomal pairing, namely, 4.79–5.49, 0.19–0.39 trivalents and 0.04–0.07 quadrivalents (Mello-Sampayo 1971; Feldman 1978; Ceoloni et al. 1986).

9.4.6 Aegilops searsii Feldman & Kislev Ex Hammer

9.4.6.1 Morphological and Geographical Notes

Aegilops searsii Feldman & Kislev ex Hammer [Syn.: Triticum searsii Feldman & Kislev; Sitopsis searsii (Feldman & Kislev ex Hammer) Á. Löve] is a predominantly autogamous annual plant. Its culm is 20–40 (-50)-cm-high (excluding spikes). The spike is narrow, linear, 5–11-cm-long (excluding awns), one-rowed, and tapers slightly towards the tip. At maturity, it disarticulates above the lowest spikelet and falls as a unit. There are 8–12 spikelets, which are linear, and generally adorned with 3 florets per spikelet in middle of the spike, of which 2 are fertile, 1 in the lower and one in the upper part of spike. The uppermost floret in the middle of spike is sterile. The rachis internode is more or less as long as the spikelets. Glumes feature two teeth, separated by a membranous edge. Glumes are ¾ the length of the spikelet. Lemmas of the terminal spikelet are awned. One floret has a short awn, triangular in cross section, while the other floret has a very long (equal to or longer than the spike), flat awn. Awns are flanked at the base by 1–2 unequal, short–articulate teeth; the middle tooth at glumes in terminal spikelet sometimes lengthened into an up to 1-cm-long awn. Anthers are short (2–3 mm). The caryopsis is more or less free at maturity (Fig. 9.1f).

A unique character of Ae. searsii is the unequal (one very long and the second short or sometimes absent) awns in the terminal spikelet, which undoubtedly results from the presence of only one fertile floret in the terminal spikelet. Generally, in the related species of the section, two developed awns form at the top of the ear, due to the occurrence of two fertile florets.

It is assumed (Feldman and Kislev 1977) that Ae. searsii is a young species which returned from the relatively dry habitats of the other Emarginata species to more mesophilic habitats. The very low stature of the plant, as well as its short ear, the almost equal length between the glumes and the florets, as well as the free caryopsis, are considered characteristics of advanced members of the genus (Eig 1929a).

Ae. searsii has limited morphological variation, involving mainly spike size, spikelet number and awn length. Israel and Jordan are the center of variation, where it probably originated. Morphologically, Ae. searsii is close to Ae. longissima, but differs from it in habitat and in the following characteristics: shorter plant, shorter spike, fewer spikelets, fewer fertile florets, larger glumes to spikelet ratio, awns of lemma in the terminal spikelet are very unequal, shorter anthers and free caryopsis.

Analysis of electrophoretically discernible water-soluble leaf proteins (Mendlinger and Zohary 1995) and random amplified polymorphic DNA (RAPD) (Goryunova et al. 2008), showed that Ae. searsii (and Ae. bicornis) displays the lowest diversity among the Sitopsis species. This lies in agreement with earlier studies (Dvorak and Zhang 1992b; Sasanuma et al. 1996; Giorgi et al. 2002; Goryunova et al. 2008), which have shown that Ae. searsii appears to be remote from the other three Emarginata species, equally distant from Ae. longissima, Ae. bicornis and Ae. sharonensis (Mendlinger and Zohary 1995). Likewise, meiotic pairing analyses in triploid hybrids of Ae. searsii with autotetraploid Ae. longissima, autotetraploid Ae. speltoides and autotetraploid Ae. bicornis, indicated the S^s genome of Ae. searsii is equally distant to the S genome of Ae. speltoides, the S^l genome of Ae. longissima and the S^b genome of Ae. bicornis (Yen and Kimber 1990a).

Ae. searsii has a relatively limited distribution region in the south-central area of the distribution of the genus. It occupies east Mediterranean primary and secondary habitats. It is a sub-Mediterranean element, extending into the Irano-Turanian region. Alt: 200–1000 m above sea level. Ae. searsii is limited to Israel (Judea, Samaria, the Golan Heights, and the higher Negev), Jordan (Gilead, Ammon, Moav, and Edom), southeast Lebanon and southwest Syria. It grows on terra rossa, sometimes mixed with loess, or on basalt soil in the destroyed sub-Mediterranean habitat of Sarcopoterium spinosum, in open-park herbaceous formations, in the degraded deciduous steppe-maquis, in small shrub (Batha) formations, abandoned fields and edges of cultivation. Locally common, sometimes dense stands in Judea.

Ae. searsii grows sympatrically with Ae. peregrina, Ae. geniculata, Ae. biuncialis and T. turgidum subsp. dicoccoides (in Judea and Samaria), with Ae. triuncialis, T. monococcum subsp. aegilopoides and T. urartu (in Lebanon and Syria), and with Ae. kotschyi and Ae. vavilovii (in southern Jordan and Israeli Negev). It grows allopatrically with Ae. speltoides, Ae. longissima, Ae. caudata, Ae. umbellulata and Ae. neglecta. Ae. searsii is involved in the parentage of Ae. vavilovii (mixed populations of these two species occur in the higher central Negev in Israel and in southern Jordan), and it presumably introgressed with wild tetraploid wheat, T. turgidum subsp. dicoccoides, Ae. peregrina and Ae. kotschyi. This is the only diploid Aegilops species that grows in mixed populations with wild T. turgidum in the southern Fertile Crescent. It also has massive contact with T. urartu in southeastern Lebanon and southwestern Syria. Variation in the ability to suppress the Ph1 gene of polyploid wheat has not yet been demonstrated in this species.

9.4.6.2 Cytology, Cytogenetics, and Evolution

Ae. searsii is a diploid (2n = 2x = 14), with a modified S genome, designated S^s by Feldman et al. (1979). Its organellar genome was designated S^v (Ogihara and Tsunewaki 1988; Wang et al. 1997) since it differs from those of the other Sitopsis species (Wang et al. 1997). The genome of Ae. searsii is the smallest genome in the Emarginata subsection (1C DNA = 6.65 ± 0.091 pg; Eilam et al. 2007), 5.9 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. searsii genome after genome sequencing and assembly to be 5.55 Gb (=5.42 pg). This genome size is somewhat smaller than that of Ae. bicornis and considerately smaller than those of Ae. sharonensis and Ae. longissima, but larger than that of Ae. speltoides. The karyotype of Ae. searsii is symmetric, with two metacentric chromosome pairs and five sub-metacentric pairs (Feldman and Kislev 1977; Feldman et al. 1979). One of the metacentric pairs and one of the submedian pairs are satellited chromosomes; the metacentric pair has large satellites and the submetacentric one has a pair of medium-size satellites. Ae. searsii differs from the other three Emarginata species in that it has a pair of the medium-size satellites, whereas the other species contain a pair of smaller satellites.

Karyotype analysis showed the presence of two secondary constrictions in Ae. searsii (Feldman and Kislev 1977; Feldman et al. 1979; Teoh and Hutchinson 1983). In situ hybridization analysis, using a radioactive rDNA probe (Teoh et al. 1983), revealed that two pairs of NORs are located in the secondary constriction of Ae. searsii. Yet, hybridization with a 18S–26S rDNA probe, which enabled high sensitivity, revealed that the two-major active NORs are located on the short arms of chromosomes 5S^s and 6S^s (Badaeva et al. 1996b). The numbers and chromosomal locations of the NORs corresponded to the number of satellited (SAT) chromosomes (Feldman and Kislev 1977; Feldman et al. 1979; Teoh and Hutchinson 1983), indicating that these regions are actively transcribed. An additional pair of minor NORs was detected on the short arm of chromosome 1S^s (Badaeva et al. 1996b). The minor NORs do not usually form secondary constriction, indicating that these NORs are not transcribed. In situ hybridization with the pTa794 (5S rDNA) probe showed that the 5S rDNA loci are located in Ae. searsii on the short arms of chromosomes 1S^s and 5S^s (Badaeva et al. 1996b).

Ae. longissima chromosomes were similar to Ae. searsii chromosomes in hybridization patterns with both the pTa7l and pTa794 DNA probes, confirming the high degree of similarity between their genomes (Teoh et al. 1983; Badaeva et al. 1996b).

Teoh and Hutchinson (1983), using an improved C-banding technique, found that the C-banding patterns of mitotic metaphase chromosomes of Ae. searsii exhibited a characteristically different pattern from those of the Sitopsis species. Moreover, the C-banding pattern enables the chromosomes of this species to be identified individually. Ae. searsii, like other Sitopsis species, has telomeric, interstitial, and centromeric bands. However, Ae. searsii and Ae. bicornis have relatively smaller centromeric bands (Teoh and Hutchinson 1983).

Teoh and Hutchinson (1983) analyzed only one accession per species. C-banding polymorphism in a larger number of accessions of the diploid Aegilops species was studied by Friebe et al. (1992b, 1993, 1995b) and Friebe and Gill (1996). C-banding polymorphism was analyzed in 14 accessions of Ae. searsii from Israel, enabling establishment of a generalized idiogram of the species (Friebe et al. 1995b). All seven Ae. searsii chromosome pairs were individually identified on the basis of their C-banding patterns. No variation was observed within the accessions, but C-band polymorphism was detected between the different accessions. Ae. searsii was easily distinguished from Ae. longissima by their distinct C-banding patterns; Ae. searsii had fewer and smaller C-bands, and, in this respect, was similar to Ae. bicornis (Friebe et al. 1995b). Isozyme studies (Pietro et al. 1988) showed that there is no translocation 4/7 in Ae. searsii, also differing in this respect, from Ae. longissima.

C-banding analysis was also used to identify the seven disomic addition lines of Ae. searsii chromosomes to T. aestivum cv. Chinese Spring, 14 ditelosomic chromosome addition lines, 21 disomic substitution of whole chromosome and 31 ditelosomic chromosome substitution lines, produced by NA Tuleen. The identity of these lines was further confirmed by meiotic pairing analysis. Sporophytic and gametophytic compensation tests were used to determine the homoeologous relationships of the Ae. searsii chromosomes. The results show that the Ae. searsii chromosomes do not compensate well for their wheat homoeologues. The short arm of Ae. searsii, 3S^sS, causes spike fragility; the spike breaks between the first and the sixth spikelets. Chromosome arm 2S^sS causes tenacious glumes.

In situ hybridization with the highly repetitive DNA sequences pSc119 and pAsl, and C-banding analysis, enabled the genome differentiation in all the diploid species of Aegilops. The ISH patterns of chromosomes with pSc119 showed only minor intraspecific variations in the subsection Emarginata species. Chromosomes of all these diploid species hybridized with the pSc119 probe; however, the level of hybridization and labeling patterns differed among genomes (Badaeva et al. 1996a). All these species had strong pSc119 hybridization sites located mainly in the telomeres. Ae. longissima and Ae. sharonensis are highly heterochromatic species; C-bands were present in intercalary, telomeric and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis have much less C-heterochromatin compared with the other two Emarginata species and hence, these species form a third subgroup (Badaeva et al 1996a).

On the other hand, the Sitopsis species differ in their ability to hybridize with the pAs1 probe (Badaeva et al. 1996a). No hybridization was observed with the pAs1 probe on Ae. speltoides, Ae. sharonensis and Ae. longissima chromosomes. A few minor pAs1 sites were observed in Ae. searsii and Ae. bicornis.

The evolution of two tandemly repeated sequences, Spelt1 and Spelt52, was studied in species of Aegilops sect. Sitopsis (Salina et al. 2006). Fluorescence in situ hybridization showed considerable polymorphisms in the hybridization patterns of Spelt1 and Spelt52 repeats between and within Aegilops species. Hybridization patterns of Spelt52 in Ae. speltoides, Ae. longissima and Ae. sharonensis were species-specific. There was no detectable Spelt1-associated FISH signal in Section Sitopsis, with the exception of Ae. speltoides. Spelt52 and its analogues pGc1R-1 and pAesKB52, were found in Ae. speltoides, Ae. longissima and Ae. sharonensis, but not in Ae. bicornis or Ae. searsii (Anamthawat-Jonsson and Heslop-Harrison 1993; Zhang et al. 2002; Salina et al. 2004a).

Yaakov et al. (2013) assessed the relative copy numbers of a number of TE families in five accessions of Ae. searsii and in several other Sitopsis species. The analysis of six Gypsy families revealed that Fatima is very abundant in Ae. searsii and in Ae. speltoides. On the other hand, several elements, i.e., Latidu, Sabrina, BAGY2 (all Gypsy retrotransposon) and Charon (Mutator DNA transposon), had specific proliferation in the Ae. searsii genome, in comparison to the other Sitopsis species.

9.4.6.3 Crosses with Other Species of the Wheat Group

Very few hybrids between Ae. searsii and other Aegilops and Triticum species were produced and analyzed. Feldman et al. (1979) noticed that the crossability between Ae. longissima with Ae. searsii is apparently very low. Over 380 pollinated florets (about 15 spikes) in both directions yielded about 10 seeds, and then, only when Ae. longissima was the female parent. Most of the seeds were shriveled and their embryos poorly developed; they either did not germinate, even after placing the embryo on culture medium, or the seedlings died very young. Only three seeds, representing two hybrid combinations, germinated and survived the seedling stage. When studying chromosome pairing at first meiotic metaphase of these F₁ hybrids, Feldman et al. (1979) found that meiosis was irregular and chromosomal pairing was somewhat reduced (Table 9.4). One of the rod bivalents was heteromorphic, indicating that the total length and the arm ratio of the two pairing partners were not the same. Moreover, most meiocytes exhibited one asymmetrical multivalent, indicating that the two species differ in one asymmetric reciprocal translocation. The asymmetrical configuration implies that the translocated segments were unequal. Further indication of chromosomal differentiation between Ae. searsii and Ae. longissima was the occurrence of a chromatid bridge and an acentric fragment at first anaphase of the two hybrids, indicating heterozygosity for a paracentric inversion. A comparison of the karyotypes of the two species confirms the existence of several structural differences between the chromosomes of the two species (Feldman et al. 1979). Pollen and seed fertility of the F₁ hybrids was low; pollen fertility was 30–37% and seed fertility 6.7–35%. The significant reduction in meiotic pairing in these hybrids, as manifested by reduced numbers of chiasmata, shows the incomplete homology between the Ae. searsii and Ae. longissima genomes. Clearly, in one or both species, several chromosomes have undergone independent re-patterning, as indicated by the asymmetrical translocation, heteromorphic bivalent and the paracentric inversion.

Ae. searsii and Ae. longissima are presumably isolated via several different mechanisms that include low crossability, partial hybrid sterility and differences in ecological requirements and geographical distribution. In addition to these, hybridization between the two species in nature is also greatly reduced because of the predominance of self-pollination. The multiplicity of isolating mechanisms is probably responsible for the rarity of hybrids between these species in the contact zone between them.

Yu and Jahier (1992) produced F₁ hybrid between the allotetraploid Ae. peregrina and Ae. searsii. Analysis of chromosome pairing at first meiotic metaphase (Table 9.5) revealed that the S^v subgenome of Ae. peregrina is not completely homologous to that of Ae. searsii. From the level of pairing in the triploid hybrid, Yu and Jahier (1992) concluded that subgenome S of Ae. peregrina is very close to the genome of Ae. longissima, and relatively more distant from that of Ae. searsii.

Hybrids between hexaploid wheat, Triticum aestivum ssp. aestivum and Ae. searsii were produced and analyzed by Feldman (1978) and Yu and Jahier (1992) (Table 9.10). The low pairing observed in these hybrids indicated that very little homology exists between the S^s genome of Ae. searsii and all of the three subgenomes of T. aestivum. Hence, Ae. searsii is not the source of the B subgenome of allopolyploid wheat.

9.5 Section Vertebrata Zhuk. Emend. Kihara

9.5.1 General Description

Section Vertebrata Zhuk. emened Kihara [Syn.: Pachystachys Eig, Gastropyrum (Jaub. & Spach) Zhuk.; Polyploids Zhuk.; Aegilonearum Á. Löve] consists of annual, predominantly autogamous species. The plants are robust, with a more or less long spike, (3-) 6–10 (-15)-cm-long, mostly thick, 3–7 mm wide, cylindrical or moniliform, mostly awned and disarticulating at maturity into spikelets, each with the laterally adjacent rachis-internode (barrel-type dispersal unit). Spikelets are more or less ventricose or linear and glumes are mostly awnless. Lemmas of apical spikelets, sometimes of several upper spikelets, with one well-developed broad and mostly flat awn. The caryopsis adheres to lemmas and palea (Fig. 9.2).

Section Vertebrata contains five species, one diploid (Ae. tauschii Coss., genome DD) and two tetraploids [Ae. ventricosa Tausch (genome DDNN) and Ae. crassa Boiss. (genome D^cD^cX^cX^c). The latter also has a hexaploid cytotype (genome D^cD^cX^cX^cDD)], and two hexaploids (Ae. juvenalis (Thell.) Eig (genome D^cD^cX^cX^cUU) and Ae. vavilovii (Zhuk.) Chennav. (Genome D^cD^cX^cX^cS^sS^s). Kimber and Zhao (1983) and Zhao and Kimber (1984), based on analysis of meiotic chromosome pairing, decided that the allopolyploid species containing the D subgenome can be divided into three clusters: (1) T. aestivum, Ae. cylindrica and Ae. ventricosa, in which the D subgenome has undergone little modification from the genome of the diploid progenitor, Ae. tauschii; (2) tetraploid and hexaploid Ae. crassa, in which the D subgenome is somewhat modified; (3) Ae. juvenalis and Ae. vavilovii, in which the D subgenome is substantially modified. Dubcovsky and Dvorak (1995a), Dvorak (1998) and Dvorak et al. (2012) arrived at a similar conclusion but pointed out that the D subgenome of Ae. crassa differentiated considerably from that of the diploid parent and consequently, it was designated D^c (Dvorak 1998). Analyses of the plasmon of these species clearly showed that Aegilops tauschii is the maternal parent of the two allotetraploid species, Ae. crassa and Ae. ventricosa, and that the tetraploid cytotype of Ae. crassa is the maternal parent of three hexaploids, Ae. juvenalis, Ae. vavilovii and 6x Ae. crassa (Tsunewaki 1993, 2009; Ogihara and Tsunewaki 1988; Wang et al. 1997). These studies suggested that the D-genome containing allotetraploids originated at three different times from Ae. tauschii in the following order: Ae. crassa, Ae. cylindrica, and Ae. ventricosa. Ae. crassa is the oldest allotetraploid containing the D genome, while Ae. cylindrica and Ae. ventricosa originated later. (Tsunewaki 1993, 2009; Ogihara and Tsunewaki 1988; Wang et al. 1997). Thus, 4x Ae. crassa appears to be an ancient allotetraploid that originated from hybridization of primitive Ae. tauschii with an ancient species in the evolutionary lineage leading to the section Sitopsis (Dubcovsky and Dvorak 1995a; Badaeva et al. 2002).

The Vertebrata species are characterized by, more or less, high morphological resemblance. All the allopolyploid species contain the D subgenome or a modified D subgenome that derived from the genome of the diploid species, and the impact of this genome is clearly apparent in the morphology and dispersal unit type of all the allopolyploid species. In addition, the organellar genome of all the allopolyploid species derived from that of the diploid species.

The central and northeastern part of the Fertile Crescent (i.e., eastern Turkey, Syria, Iraq and Iran) is the region with the greatest diversity for the Vertebrata species (Table 9.1). Only one species, Ae. ventricosa, occurs exclusively west of the Fertile Crescent, while other three, Ae. tauschii, Ae. crassa and Ae. juvenalis, have spread eastward, and Ae. vavilovii grows in the southeastern part of the genus distribution area. With the exception of Ae. juvenalis and, to a lesser extent, Ae. vavilovii (both hexaploids), all Vertebrata species are widespread.

The distribution of Ae. ventricosa is the most difficult to explain in an evolutionary sense, as it distinctly does not overlap with the areas of any of its putative parents. Ae. tauschii is the donor of one subgenome to Ae. ventricosa (Kihara 1949) and the second genome is derived from Ae. uniaristata (Yen and Kimber 1992b). There seems only a theoretical overlap on the Istrian peninsula of Croatia of uniaristata and ventricosa, but Ae. tauschii is still far away.

Some Vertebrata species are also found at higher altitudes and may show better adaptation to cold than most species of Aegilops. Several species grow in areas with relatively little annual rainfall and may be drought-tolerant. On the other hand, the D-genome of these species may be responsible for the overall poor performance of species with this genome in resisting rust infection (Hammer 1987).

9.5.2 Aegilops tauschii Coss.

9.5.2.1 Morphological and Geographical Notes

Ae. tauschii Coss. [Syn: Ae. squarrosa L.; Triticum squarrosum (L.) Rasp.; Triticum aegilops P. Beauv. ex Roem. & Schult.; Triticum tauschii (Coss.) Schmahlh.; (Coss.) Á. Löve], also known as Tausch’s goat grass and rough-spike hard grass, is a predominantly autogamous, annual plant. It is tuft, few- to many-tillered plant, its culms are 20–45-cm-high (excluding spikes) and it defoliates in lower parts at maturity. The spike is cylindrical, thick or thin, one-rowed, tapers slightly to the tip, and is relatively long (4–10-cm-long, excluding awns). At maturity, the spike disarticulates into individual spikelets, each with its adjacent rachis segment (barrel-type dispersal unit). The number of spikelets per spike is 5–13, cylindrical, barrel-shaped, usually equal in length to the adjacent rachis segment. Basal rudimentary spikelets are absent, or rarely, there are 1 or 2. There are 3–5 florets, usually 4, with the upper 1–3 being sterile. Glumes are almost rectangular, narrow, equally spaced and with small nerves. The tip of the glume is truncated, with a clearly thickened edge, with one or no teeth. Lemmas are membranous, with a keel that terminates with a small tooth or awn, up to 4-cm long, and sometimes accompanied by 1–2 short, wide lateral teeth. The lemmas of the apical spikelet have longer awns, that up to 5.5-cm-long. The awns are triangular in cross section and are shorter on the lower spikelets. The caryopsis adheres to lemmas and palea (Fig. 2.1).

Ae. tauschii possesses a very wide morphological variation, mainly in spike shape, length and width, awned to awnless, straight or winding rachis, cylindrical to square spikelets, number of spikelets per spike and length of awns. Several accessions have a tough rachis (Waines et al. 1982; Knaggs et al. 2000). The morphological variation and ecological amplitude of Ae. tauschii exceed those of any other diploids of the Triticum-Aegilops group (Zohary et al. 1969). This variation led Eig (1929a) to classify the various morphological forms in two subspecies, ssp. eusquarrosa Eig (now ssp. tauschii), which has elongated, cylindrical spikelets (Fig. 9.2a), and ssp. strangulata (Eig) Tzvelev, which have a curved rachis segment, noticeably longer and narrower than the adjacent spikelets, giving the spike a markedly moniliform appearance, as well as square spikelets, equally long as wide (Fig. 9.2b). The latter subspecies is morphologically distinct, being taller with greater seed weight and rounded seed shape than in ssp. tauschii. Despite its variability, ssp. strangulata has been considered a discrete taxon (Jaaska 1981). Ssp. tauschii has been found throughout the geographic range of the species while ssp. strangulata mainly occurs in narrow belts along the southeastern Caspian Sea in Iran (Zohary et al. 1969; Tanaka 1983; van Slageren 1994).

Ssp. tauschii was further divided into three varieties: typica Eig (now var. tauschii), anathera (Eig) Hammer (awnless), and meyeri (Griesb.) Tzvelev (has a slender spike). var. anathera is rather easily distinguishable by awnless type and is shorter than the other varieties, whereas var. tauschii and var. meyeri proved difficult to identify and were not easily distinguished due to many intermediates between them (Knaggs et al. 2000). In spite of this difficulty, subdivision of Ae. tauschii on the basis of morphology appears to be reasonably valid (Knaggs et al. 2000).

Eig’s classification was based primarily on variation in spike morphology. Hammer (1980) accepted Eig’s intraspecific classification of Ae. tauschii and added var. paleidenticulata (Gandiljan) Hammer to ssp. tauschii. Hammer (1980) recommended using Ae. tauschii as the correct name for this species, renamed ssp. eusquarrosa as ssp. tauschii and var. typica as var. tauschii. Knaggs et al. (2000), based on morphological study of a large number of Ae. tauschii accessions, also identified the two subspecies, tauschii and strangulata, and the three varieties of ssp. tauschii. Kihara and Tanaka (1958) adopted Eig’s (1929a) intraspecific classification of this species and presented a detailed account of the morphological and genetic variation of Ae. tauschii. Yet, Kihara and Tanaka (1958) and Kihara et al. (1965) showed all three varieties of ssp. tauschii are interfertile and described many intermediates between them. Only ssp. strangulata is more distinct, and its occurrence is limited to a narrow belt on the southern shores of the Caspian Sea (Kihara and Tanaka 1958; Zohary et al. 1969) and in Armenia, Azerbaijan, and Turkmenistan (Kim et al. 1992). In the southern shores of the Caspian Sea, in northern Iran, ssp. strangulata grows next to many robust types of Ae. tauschii with large, thick spikes that presumably have segetal growth (Zohary et al.1969). Kim et al. (1992) pointed out that intermediate forms exist among all morphologically distinct forms of Ae. tauschii and that gene flow takes place quite frequently between the two subspecies. Moreover, they found that a ribosomal DNA genotype of an accession of ssp. strangulata from Armenia was different from other strangulata genotypes and similar to genotypes of the tauschii subspecies. Consequently, Kim et al. (1992) considered intraspecific classification of Ae. tauschii on morphological grounds inadequate. Similarly, van Slageren (1994) thought it justified not to classify the observed intraspecific variation of Ae. tauschii. To this day, the status of subspecies and varieties is still under discussion (Hammer 1980; van Slageren 1994; Dudnikov 2000; Knaggs et al. 2000). Nevertheless, the morphologically-based intraspecific classification of Ae. tauschii has been used by many cytogeneticists.

Since the discovery that Ae. tauschii donated its genome (genome D) to hexaploid wheat (McFadden and Sears 1944, 1946; Kihara 1944), Ae. tauschii has been a subject of intensive genetic, cytogenetic, and molecular studies which seek answers to many questions, including the center of genetic diversity of Ae. tauschii and the place of origin of hexaploid wheat (Gill 2013). A large portion of these studies have concentrated on assessment of the intraspecific genetic diversity of this species in order to understand the factors leading to the evolutionary success of Ae. tauschii and to exploit its rich gene pool for wheat improvement.

Very wide intraspecific variation in Ae. tauschii has been demonstrated through morphological (Kihara and Tanaka 1958; Hammer 1980; Knaggs et al. 2000), isozyme (Nishikawa et al. 1980; Jaaska 1981; Dudnikov 2014 and reference therein), restriction fragment length polymorphism (RFLPs) (Lubbers et al. 1991; Dvorak et al. 1998c), amplified fragment length polymorphism (AFLPs) (Saeidi et al. 2008) and microsatellite (Pestsova et al. 2000; Saeidi et al. 2006) analyses. Diversity was studied in accessions of Ae. tauschii that were collected from the western (Syria, Turkey, and Georgia) to the eastern (Central Asia and western China) range of its distribution. The above studies showed that accessions from the region along the Southern Caspian Sea exhibited the highest genetic variation, suggesting that this region is the center of variation of Ae. tauschii. Genetic variability within the D subgenome of wheat is much lower than it is within Ae. tauschii (Appels and Lagudah 1990; Lagudah et al. 1991a; Lubbers et al. 1991), so the wild progenitor offers great potential for wheat improvement. Utilization of Ae. tauschii for wheat improvement is further aided by the ability of the chromosomes of Ae. tauschii and the D subgenome chromosomes of bread wheat to naturally recombine.

Isozyme analysis conducted by Jaaska (1981) revealed intraspecific differentiation of Aegilops tauschii into two groups of biotypes, which essentially correspond to its two-morphological subspecies, tauschii and strangulata. Dudnikov (2014) pointed out that the isozymic variation in Ae. tauschii reflects adaptive intraspecies divergence: ssp. strangulata favors the habitats of the Caspian seaside climate, with warm and moist winters, while ssp. tauschii mostly occupies the habitats, which have a rather continental climate, with relatively cold and dry winters.

RFLP marker analysis of genetic diversity, performed by Tsunewaki et al. (1991) and Lubbers et al. (1991), found close similarities between ssp. tauschii var. meyeri and ssp. strangulata. A molecular study by Dvorak et al. (1998c) found evidence of gene migration between the different divisions in accessions from the southwest Caspian area of Iran. The greatest amount of variation was found in Ae. tauschii accessions collected in Iran and western Transcaucasia (Lubbers et al. 1991; Tsunewaki et al. 1991). Var. meyeri, although formally placed in ssp. tauschii, was found to be genetically closer to ssp. strangulata than to ssp. tauschii, and consequently, Lubbers et al. (1991) concluded that var. meyeri actually belongs to ssp. strangulata.

Actually, Lubbers et al. (1991) identified two genetically diverse groups, one consisting of ssp. tauschii vars. tauschii and anathera, and the other of ssp. strangulata and ssp. tauschii var. meyeri. Their analysis strongly supported the suggestion the that Caspian Sea region is the center of genetic diversity and origin of Ae. tauschii. Likewise, Dvorak et al. (1998c), in a large study of RFLP markers, confirmed the existence of two genetically diverse groups in Ae. tauschii, which crosscut taxonomic groupings, but, in contrast to Lubbers et al. (1991), proposed Armenia as the center of genetic diversity and origin of hexaploid wheat. However, Wang et al. (2013b), using 7815 single nucleotide polymorphisms (SNPs) previously mapped by Luo et al. (2009), that provide complete coverage of the genome, to analyze 402 accessions of Ae. tauschii, 75 hexaploid wheats, and seven tetraploid wheats, concluded that southwestern Caspian Iran is the center of genetic diversity of Ae. tauschii and the center of origin of hexaploid wheat.

As seen from the above, the intraspecific botanical classification of Aegilops tauschii agrees poorly with the genetic relationships (Dvorak et al. 1998c). The most apparent contradiction is encountered with var. meyeri, which is assigned to ssp. tauschii on the basis of morphology but is genetically closely related to ssp. strangulata (Lubbers et al. 1991; Dvorak et al. 1998c). In genetic studies, therefore, the use of categories based on genetic subdivision of A. tauschii is preferable to those based on formal taxonomy (Wang et al. 2013b). In fact, recent genetic and molecular studies have shown that two major lineages exist in Ae. tauschii (Takumi et al. 2008; Mizonu et al. 2010; Sohail et al. 2012; Wang et al. 2013b). These two lineages were named by Takumi et al. (2008) and Mizuno et al. (2010) lineage 1 (L1) and lineage 2 (L2). L1 consisted of accessions of ssp. tauschii from the eastern habitats, mainly Afghanistan and Pakistan, whereas L2 included ssp. tauschii accessions of the western habitats and all accessions of ssp. strangulata. Accordingly, both lineages included accessions of ssp. tauschii, whereas ssp. strangulata belonged only to lineage 2. The two varieties of ssp. tauschii, meyeri and anathera were classified into L2 and L1, respectively.

Nucleotide sequence variations of 10 nuclear genes were used to construct a phylogenetic tree for each gene, which were compared to the SSR phylogenetic tree (Takumi et al. 2008). Although some discrepancies were found between the trees, the results supported the subdivision of Ae. tauschii into two major lineages and suggested that ssp. strangulata derived from only one of the two lineages, i.e., lineage 2. This phylogenetic tree was consistent with a previous report of a similar SSR analysis (Pestsova et al. 2000).

Mizuno et al. (2010) conducted an AFLP analysis to study population structure of 122 accessions of Ae. tauschii. Their phylogenetic and principal component analyses revealed, similar to the finding of Takumi et al. (2008), two major lineages in Ae. tauschii, i.e., lineages one (L1) and two (L2). The results of the study of Wang et al. (2013b) also supported the subdivision of Ae. tauschii into two lineages (L1 and L2) that have been reproductively isolated in nature. Particularly informative was a comparison of the SNPs tree, obtained by Wang et al. (2013b), with the AFLP tree reported by Mizuno et al. (2010). In both trees, branches in L2 were longer than those in L1, indicating that L2 was more diverse than L1. Wang et al. (2013b) found that each lineage consists of two closely related sub-lineages that appear to be geographically isolated. Within L1, sub-lineage 1W is located in Turkey, Transcaucasia, and western Iran, whereas sub-lineage 1E is located from central Iran to China. Sub-lineages 1W and 1E are predominantly found at high elevations (400–3000 m above sea level). Within L2, sub-lineage 2W occupies elevations between 400 and 1500 m in Transcaucasia (Armenia and Azerbaijan), whereas sub-lineage 2E occupies elevations ≤25 m and is distributed across Azerbaijan and Caspian Iran. The 2E sub-lineage is morphologically heterogeneous, including both the typical moniliform Ae. tauschii ssp. strangulata in southern Caspian Iran and morphologically intermediate forms classified as ssp. tauschii vars. meyeri and tauschii in southern and southwestern Caspian Sea.

The poor agreement between morphological and genetic relationships among A. tauschii accessions (Lubbers et al. 1991; Dvorak et al. 1998c; Mizuno et al. 2010; Sohail et al. 2012; Wang et al. 2013b) was reconciled by two mutually exclusive hypotheses (Wang et al. 2013b): (1) the morphological traits of Ae. tauschii and the subsequent classification are trustworthy, but the taxa are genetically heterogeneous as a result of gene flow between them; or (2) Ae. tauschii is genetically clearly subdivided but the subdivision is not faithfully reflected by morphology and taxonomic classification. Clear genetic separation of lineages 1 and 2 and the paucity of intermediate genotypes, led Wang et al. (2013b) to favor the second alternative.

Ae. tauschii is the donor of the D genome to hexaploid wheat, T. aestivum (McFadden and Sears 1944, 1946; Kihara 1944), to which it confers many important traits, including bread making quality (Kerber and Tipples 1969; Orth and Bushuk 1973), cold hardiness (Limin and Fowler 1981; Le et al. 1986) and salt tolerance (Schachtman et al. 1992). In addition, many accessions of Ae. tauschii, particularly those of subsp. strangulata, showed resistance to various diseases (Yildirim et al. 1995; Cox et al. 1995; Appels and Lagudah 1990; Knaggs et al. 2000). Limin and Fowler (1981) rated cold hardiness in a large number of accessions from species which share a common genome with hexaploid wheat and found that more than half of the Ae. tauschii accessions survived the coldest temperatures of the Canadian winter and had a hardiness level which approached that of the hardiest winter cultivars of hexaploid wheat. Le et al. (1986) obtained similar results. These results support Tsunewaki’s (1968) suggestion that the addition of the D genome to tetraploid wheat made spread of the cultivation of the resulting hexaploid to colder northern countries possible. Indeed, study of intervarietal substitution lines, in which a chromosome of a cold hardiness cultivar of hexaploid wheat substituted its homologous chromosome in a spring cultivar, showed that chromosomes 4D and 5D accounted for much of the difference in cold hardiness between these two cultivars (Law and Jenkins 1970; Cahalan and Law 1979).

The differentiation of A. tauschii into two lineages brings forth several questions relevant to the origin of wheat and shaping of its diversity (Wang et al. 2013b). Recurrent hybridization and introgression between wheat and Ae. tauschii were known to have played a role in the origin of wheat D-subgenome diversity, although the magnitude is unknown (Dvorak et al. 1998a, b, c; Talbert et al. 1998; Caldwell et al. 2004; Akhunov et al. 2010). Did only lineage 2 contribute germplasm to the wheat D subgenome? If so, why lineage 2 and not lineage 1? Diversity is uneven among and along the wheat D-subgenome chromosomes (Akhunov et al. 2010). So, does the distribution of diversity along wheat chromosomes have anything to do with its distribution along the A. tauschii chromosomes, and what is the cause of this pattern?

The wheat D subgenome, unlike the other two subgenomes of hexaploid wheat, shows great fluctuation in diversity among chromosomes (Akhunov et al. 2010). Similar uneven distribution of diversity among all D-genome chromosomes was noted by Wang et al. (2013b) in Ae. tauschii. They also showed that diversity correlated with recombination rates along the chromosomes. Similar correlations were also observed along each chromosome in both lineages of Ae. tauschii. Likewise, RFLP was shown to correlate with recombination rates in Ae. tauschii (Dvorak et al. 1998a).

Previous genetic studies placed the origin of wheat in Transcaucasia and southwestern Caspian Iran (Tsunewaki 1966; Nakai 1979; Jaaska 1980; Dvorak et al. 1998c) or southeastern Caspian Iran (Nishikawa et al. 1980). The consensus has been that ssp. strangulata was the wheat progenitor (Nishikawa 1973; Nakai 1979; Jaaska 1980; Hammer 1980; Nishikawa et al. 1980; Lagudah et al. 1991a; Lubbers et al. 1991; Dvorak et al. 1998c, 2012).

Jaaska (1981), by way of isozyme analysis, identified ssp. tauschii as the contributor of the D genome to the allotetraploid Ae. cylindrica (genome DDCC) and the third subgenome to allohexaploid Ae. crassa Boiss. (genome D^cD^cX^cX^cDD), and subsp. strangulata as the contributor of the D genome to the allohexaploid Triticum aestivum, to the allotetraploids Ae. crassa (genome D^cD^cX^cX^c) to Ae. ventricosa (genome DDNN), and to the allohexaploid Ae. juvenalis (genome D^cD^cX^cX^cUU).

Wang et al. (2013b) identified 12 Ae. tauschii accessions that are closely related to the D-subgenome of wheat. All 12 accessions belonged to sub-lineage 2E and were members of populations located in southwestern and southern Caspian Iran. In a surprising departure from the belief that ssp. strangulata was the source of the wheat D subgenome, only one of these 12 accessions had been classified as ssp. strangulata on the basis of its morphology. Eleven of the 12 accessions were classified as ssp. tauschii var. tauschii or var. meyeri. However, if it is accepted that morphology does not reflect genetic relationships, as in the second hypothesis of Wang et al. (2013b), this conflict with the previous conclusions regarding the progenitor of the wheat D subgenome becomes irrelevant, as, genetically, these accessions are members of the 2E sub-lineage. Of the 7185 SNP sites in the wheat D subgenome studied by Wang et al. (2013b), 0.8% appeared to originate by introgression from the L1 lineage, while 99% of the D subgenome was contributed by A. tauschii lineage 2. A population within lineage 2E in the southwestern and southern Caspian appears to be the main source of the wheat D subgenome (Wang et al. 2013b).

Given the extensive opportunity for natural hybridization between wheat and Ae. tauschii (Kihara et al. 1965), why does hexaploid wheat appear monophyletic and why the preference for sub-lineage 2E? An answer to this question may reside in the geography of cultivation of tetraploid wheat by early farmers. Aegilops tauschii readily hybridizes with tetraploid wheat, and triploid hybrids often produce so many unreduced gametes, that they are fertile (Zhang et al. 2010). Spontaneous chromosome doubling via union of unreduced (2n) gametes has been thought to be the way that hexaploid wheat originated from the hybridization of T. turgidum with Ae. tauschii. Previous works have observed unreduced gametes in F₁ hybrids of Ae. tauschii with six of the eight T. turgidum subspecies tested (Zhang et al. 2008, 2010). By contrast, hybridization of Ae. tauschii with hexaploid wheat is arduous, and hybrids can only be obtained with the aid of embryo rescue. Introgression from A. tauschii into hexaploid wheat should therefore only be expected in the areas where tetraploid wheat was farmed in mixed populations with hexaploid wheat (Dvorak et al. 1998c).

Today, farming of tetraploid durum wheat is limited to a few mountainous regions in northern Iran (Matsuoka et al. 2008), but the situation could have been different in the past. If wheat farming was predominantly in low elevations in Caspian Iran, and if the distribution of Ae. tauschii was similar to its present-day distribution, the only possible source of the D genome was sub-lineage 2E, as only sub-lineage 2E is found at low elevations.

Among the diploid Aegilops species, Ae. tauschii has the widest geographic distribution. It grows in a very large area in the eastern part of the distribution of the genus. This species is the only diploid that spread eastward from the center of origin of the genus. It grows in the following countries: Crimea (possibly), Ciscaucasia (northern Caucasia) (possibly), Transcaucasia (Georgia, Armenia, and Azerbaijan), Uzbekistan, Turkmenistan, Kazakhstan, Tagikistan, Kyrgyzstan, Afghanistan, China (western slopes of Himalaya), India (Kashmir), Pakistan, Iran, Iraq, Turkey, and northeastern Syria (Table 9.2). Its distribution is limited in the east by the Himalaya mountains, in the south by the deserts of southern Pakistan and southern Iran, in the north by the cold steppes of Central Asia, and in the west by the Mediterranean climate. Zohary et al. (1969) assumed that the southern shores of the Caspian Sea, Turkmenistan, and northern Afghanistan, include the ‘primary habitat’ of the species, while the peripheral localities in e.g., Central Asia, Pakistan, Armenia, Iraq, and Syria, are ‘secondary’ in being always associated with weedy growth, often in (irrigated) wheat fields. They pointed out that the center of diversity and abundance comprises a belt around the southern shores of the Caspian Sea, across northern Iran, Turkmenistan, and northern Afghanistan. In this area, Ae. tauschii attains its widest ecological amplitude and morphological diversity. From this center, it spread westward to the Turkish border and the Syrian steppes, and eastwards to Pakistan, approaching the Chinese border. It is mentioned as a rare plant in the Caucasus and the Crimea, but it is doubtful that it is a native plant there (Zohary et al. 1969). In its broadly scattered distribution area, Ae. tauschii is also a weed and often a weed in cereal fields.

Ssp. strangulata is distributed from Transcaucasia to eastern Caspian Iran (Kihara and Tanaka 1958; Kihara et al. 1965; Zohary et al. 1969; Jaaska 1980). Ssp. tauschii var. meyeri grows in the southwestern and southern Caspian Iran, where there is some overlap with ssp. strangulata. Ssp. tauschii var tauschii, var. anathera, and their intermediate types, have a broad distribution and were found in almost all parts of the distribution of the species (Kihara and Tanaka 1958). Ae. tauschii is also found in several Chinese locations near the Yellow River in Shaanxi and Henan provinces (Yen et al. 1983). These authors assume that Ae. tauschii was introduced to China with emmer wheat (Yen et al. 1983). Adventive presence of Ae. tauschii is reported from USA (New York area) and various countries in south, west and central Europe (van Slageren 1994).

Ae. tauschii occupies a variety of different habitats, from steppes and margins of deserts, to the very wet and more temperate forests of the southern Caspian seashores in Iran and to the cool and dry central Asian steppes. This diploid occurs over a strikingly wide range of climatic conditions, from the dry Artemisia steppes and margins of deserts to the rain-soaked temperate hyrcanic forest belt of the southern coast of the Caspian Sea, and from the hot plains of southern Iran to the extreme continental climate of the Central Asiatic steppes (Zohary et al. 1969). It grows on a variety of soils, i.e., grey-calcareous steppe, marl, alluvial and sandy soils and at altitudes of 200–1800 m above sea level (Hodgkin et al. 1992), infrequently at higher elevation (up to 3000 m above sea level; Wang et al. 2013b). It grows in primary habitats, being a common component of several types of plant formations, such as open areas of deciduous steppe maquis, dwarf-shrub steppe-like formations and steppical plains, as well as in secondary disturbed habitats such as abandoned fields, edges of cultivation, and roadsides, from which it has developed as a successful weed of cultivated cereals (Zohary et al. 1969).

Ae. tauschii grows sympatrically with Ae. umbellulata, Ae. columnaris, Ae. triuncialis, Ae. cylindrica, Ae. crassa and Ae. juvenalis, and allopatrically with Amblyopyrum muticum, Ae. speltoides, Ae. caudata, T. monococcum ssp. aegilopoides, T. urartu, T. timopheevii ssp. armeniacum, T. turgidum ssp. dicoccoides, Ae. geniculata, Ae. neglecta, Ae. biuncialis and Ae. kotschyi.

9.5.2.2 Cytology, Cytogenetics, and Evolution

Ae. tauschii is a diploid species (2n = 2x = 14) bearing the D genome (Kihara 1949, 1954). Its organellar genome was designated D by Tsunewaki (1993, 2009) and Wang et al. (1997). It has a small genome, the second smallest in the genus (1C DNA = 5.17 ± 0.087 pg) (Eilam et al. 2007; Table 9.2). Similar results on genome size were previously obtained by Furuta et al. (1986) and Rees and Walters (1965). Smaller amount of 1C DNA (4.17 pg) was reported by Arumuganathan and Earle (1991). The DNA amount reported by Eilam et al. (2007) is equivalent to 5.056 Mbp, since 978 Mbp exist in 1 pg DNA (Doležel et al. 2003). The main part (about 80% or more) was estimated to be repetitive DNA (You et al. 2011).

The karyotype of Ae. tauschii is symmetric, consisting of four types of chromosomes (Senyaninova-Korchagina 1932). There is only one pair with satellites on the short arm, and the centromeres in the rest of the pairs are submedian. Later studies by Riley et al. (1958) and Chennaveeraiah (1960), confirmed the findings of Senyaninova-Korchagina (1932), but Chennaveeraiah (1960) recognized five types of chromosomes in the Ae. tauschii karyotype. In all analyzed accessions of Ae. tauschii, the size of the satellites was very much the same, varying from 0.7 to 0.8 μ (Chennaveeraiah 1960). The short arm that bears the satellite being about 3 times larger than the satellite (Chennaveeraiah (1960). There is, however, a negligible difference in the lengths of the long arms of this satellite pair. This long arm is roughly twice the size of the short arm that bears the satellite. The karyotypic differences between the varieties seem to be very small; there is more uniformity than there are differences.

Teoh et al. (1983) used a DNA sequence that codes for ribosomal RNA in in situ hybridization experiments on the genome of Ae. tauschii. This sequence consists of 18S and 25S rRNA genes, with associated spacer DNA (Gerlach and Bedbrook 1979). They found a complete fit between the number of satellited chromosomes and the number of rRNA sites; one pair of chromosomes exhibited one such site. Likewise, Badaeva et al. (1996b) studied the distribution of the 5S and 18S-26S ribosomal RNA gene families on Ae. tauschii chromosomes by in situ hybridization with pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) DNA clones. The distribution of hybridization sites with pTa71 in Ae. tauschii was unique. Only one major 18S-26S rDNA locus, that was found in the NOR of Ae. tauschii, was located on the short arm of chromosome 5D. Similar results were obtained by Appels et al. (1980), Lawrence and Appels (1986) and Lassner et al. (1987). In addition, a minor NOR locus was located on the short arm of chromosome 7D (Badaeva et al. 1996b). Hybridization with pTa794 showed the presence of two 5S rDNA loci in the short arms of chromosomes of group 1 and 5 (Badaeva et al. 1996b). This is similar to the location of these loci in most other diploid Aegilops species. The 5S rDNA loci were not associated with NORs.

The chromosomal locations of major and minor NORs and 5S rDNA loci in the D genome of Ae. tauschii were identical to those of the D subgenome of hexaploid wheat (Friebe et al. 1992a; Yamamoto 1992b), with the minor NOR on chromosome 7D being polymorphic among different lines.

The first complete investigation of C-banded chromosomes of Ae. tauschii was carried out by Teoh and Hutchinson (1983), using an improved C-banding technique. They found that Ae. tauschii exhibited a unique pattern, different from that of other Aegilops species. In this pattern, each of the seven chromosomes comprising the haploid set of Ae. tauschii displayed its own characteristic banding pattern and could be identified individually and clearly (Iordansky et al. 1978; Teoh and Hutchinson 1983). The banding pattern described by Teoh and Hutchinson (1983) is not comparable to that reported by Gill and Kimber (1974) but bears some resemblance to that of Iordansky et al. (1978).

Teoh and Hutchinson (1983) studied C-banding in only one accession of Ae. tauschii and therefore, Friebe et al. (1992a) and Badaeva et al. (1996a) extended the study to include more accessions. A generalized C-banded karyotype of Ae. tauschii was established based on chromosome analysis of 15 accessions of Ae. tauschii of diverse origins, including the two subspecies, tauschii (with the varieties tauschii, anathera and meyeri) and strangulata (Friebe et al. 1992a). Whereas only minor variation in C-banding patterns was observed within accessions, a larger amount of polymorphic variation was found between accessions. Yet, this polymorphic variation did not prevent chromosome identification in these accessions. All chromosomes of Ae. tauschii contain centromeric bands of almost equal staining intensity and size, while they differ in the amount and distribution of interstitial, subtelomeric and telomeric bands (Teoh and Hutchinson 1983; Friebe et al. 1992a). One accession (TA 2462) was found to be homozygous for a reciprocal translocation involving the complete arms of chromosomes 1D and 7D (Friebe et al. 1992a). In situ hybridization using the D-genome-specific probe, pAs1, confirmed the presence of this translocation in the accession TA 2462. The C-banding pattern of Ae. tauschii chromosomes was similar to that of the D-subgenome chromosomes of hexaploid wheat, thus permitting their unequivocal identification and homoeologous groups designations (Friebe et al. 1992a).

Badaeva et al. (1996a) studied genome structure through in situ hybridization with Ae. tauschii-derived pAsl probe. As expected, Ae. tauschii showed heavy labeling with this probe, as was found previously (Rayburn and Gill 1986; Cabrera et al. 1995). The labeling pattern in this species was chromosome-specific. Yet, intraspecific variation in the distribution of the pAs1 probe was observed (Badaeva et al. 1996a). The labeling pattern in one accession was similar to that of the D subgenome of hexaploid wheat (Rayburn and Gill 1986, 1987; Mukai et al. 1991), whereas two accessions were similar to the second unmodified D subgenome of hexaploid Ae. crassa. Comparison of the labeling pattern of pAs1 in Ae. tauschii with the distribution of heterochromatin detected by C-banding in this species, showed that Ae. tauschii possessed an intermediate amount of C-heterochromatin. On the other hand, using a highly repetitive, 260-bp, non-coding DNA sequence derived from the B subgenome of hexaploid wheat (Hutchinson and Lonsdale 1982), Ae. tauschii was shown to have very little labeling with this probe, indicating a small amount of heterochromatin (Teoh et al. 1983).

Genetic diversity of Ae. tauschii was further assessed using fluorescence in situ hybridization (FISH) with eleven DNA probes representing satellite and microsatellite DNA sequences as well as the 45S and 5S rRNA gene families and by electrophoretic (EF) analysis of seed storage proteins (gliadins) (Badaeva et al. 2019). A clear genetic differentiation of accessions into groups strangulata and tauschii was observed. These two groups differ in the presence of microsatellite repeats GAAn and ACTn and in the distribution of satellite DNA families, especially pAs1. Based on similarities of labeling patterns of DNA probes used in the study, they concluded that the strangulata group was phylogenetically closest to the D subgenome of common wheat. A comparison of spectra of gliadins revealed the highest similarity of Armenian and Azerbaijani accessions of Ae. tauschii to common wheat, which may indicate a contribution of Transcaucasian members of the strangulata group to the genetic pool of common wheat.

Since the D subgenome of hexaploid wheat and the genome of Ae. tauschii are still homologous, the genome of Ae. tauschii serves as an invaluable reference for wheat genetics and genomics as well as an important resource for wheat improvement. Undoubtedly, the usefulness of Ae. tauschii as a reference for the structure of wheat D subgenome and for the ability to identify and use important Ae. tauschii genes would be further enhanced by a high-quality sequencing of its entire D genome. Still, development of high-quality physical and genetic maps was a necessary step in progressing towards complete sequencing of the Ae. tauschii genome. These maps of Ae. tauschii are continuously updated, with more sequences and particularly, with detailed information about functional elements.

Genetic maps are based on meiotic recombination frequency between different pairs of genetic or molecular markers, with distances between markers expressed in crossover units. Physical maps use molecular biology techniques to align DNA sequences to construct maps showing the positions of sequences, including genes, relative to one another along the DNA helix axis, with actual physical distance between markers expressed in base pairs (bp). Physical distances between markers have been determined by techniques such as radiation hybrid mapping, fluorescence in situ hybridization (FISH) or, ideally, by automated DNA sequencing. Genome assembly involves a multi-step procedure, in which DNA fragments were cloned, sequenced and, on the basis of the markers they were found to contain, ordered relative to each other and to the genetic map. Obtaining sufficient coverage of the genome involved generating much physical and genetic data so that the two maps could be reconciled. Following assembly, the physical and genetic maps were continuously updated with detailed information about functional elements. For the physical sequence map, the primary annotation task was identification of genes for the genetic map. High‐resolution genetic and physical maps serve as the framework for genome sequence assembly. Accession AL8/78 of Ae. tauschii subsp. strangulata was chosen as the standard accession for the construction of physical and genetic maps.

DNA markers and coding sequences such as cDNAs and ESTs (expressed sequence tag) have been used for construction of genetic maps of Ae. tauschii. Kam-Morgan et al. (1989) used restriction fragment length polymorphisms (RFLPs) as genetic markers and determined linkage relationships between RFLP loci. In addition, they demonstrated the use of segregating populations of Ae. tauschii for linkage measurements and the use of wheat aneuploid lines to allocate markers to chromosome arms in the D subgenome of T. aestivum. Gill et al. (1991b), Lagudah et al. (1991a) and Boyko et al. (1999, 2002) used this strategy to enrich the genetic map of Ae. tauschii and that of the D subgenome of T. aestivum by a large number of loci. Boyko et al. (2002) carried on building a high-density map of the Ae. tauschii by inserting additional loci, including retrotransposon loci and microsatellite and ISSR loci. Comparison of the genetic maps of Ae. tauschii with those of the D subgenome of hexaploid wheat confirmed the conserved collinearity between the two D genomes. Yet, accessions of Ae. tauschii revealed greater polymorphism than that observed in the D subgenome of common wheat.

Physical map construction necessitates the production of a large number of bacterial artificial chromosome (BAC) clones, sequencing with a next-generation DNA sequencing platform, assembly into long contiguous sequences and anchoring the contigs on a genetic map (Luo et al. 2013). Genes and transposable elements in the assembled sequences are annotated. However, due to the lack of recombination in certain chromosomal regions, genetic mapping alone is not sufficient to develop high-quality marker scaffolds for a sequence ready physical map. Radiation hybrid mapping has proven to be a successful approach for developing marker scaffolds for sequence assembly in the genome of Aegilops tauschii. This method offers much higher and more uniform marker resolution across the length of the chromosome, compared to genetic mapping, and does not require marker polymorphism per se, as it is based on a presence vs. absence marker assay (Kumar et al. 2012, 2015). Kumar et al. (2012) reported the development of high-resolution radiation hybrid maps for the genome of Ae. tauschii accession AL8/78, the standard accession for the construction of physical and genetic maps, which were then used for anchoring unassigned sequence scaffolds. Their study demonstrated how radiation hybrid mapping, which offers high and uniform resolution across the length of the chromosome, can facilitate complete sequence assembly of large and complex plant genomes.

Several physical maps of Ae. tauschii were constructed in the last decades (e.g., Fleury et al. 2010; Massa et al. 2011; Kumar et al. 2012, 2015; Luo et al. 2013; Hastie et al. 2013; Zhu et al. 2016). The assembled scaffolds of high-quality sequences in these maps represent more than 80% of the D genome, of which about 66% of the sequences are comprised of repetitive elements. More than 43,000 protein-coding genes were identified, 71.1% of which were uniquely anchored to chromosomes. Genes, pseudogenes and transposable elements were annotated.

The Department of Plant Sciences, University of California, Davis and Genomics and Gene Discovery Unit, USDA/ARS Western Regional Research Center, Albany, California, runs a database which accumulates and releases up-to-date information on genome mapping and sequencing, genetic and physical maps, genetic markers and genomic sequences of Ae. tauschii. Gaining sufficient coverage of the Ae. tauschii genome may pave the way to the assembly of a high-quality sequence of the entire genome.

Sequencing of more than 90% of the Ae. tauschii genome was recently achieved by Jia et al. (2013). They identified 43,150 protein-coding genes and found that more than 66% of the Ae. tauschii genome was composed of 410 different transposable element families, of which the 20 most abundant comprised more than 50% of the genome.

However, even though sequencing whole plant genomes has advanced rapidly during the last decades, with the development of next-generation sequencing (NGS) technologies and bioinformatics (Bierman and Botha 2017), complete sequence assembly of the Ae. tauschii genome was problematic due to its large size and about 80% repetitive DNA (Jia et al. 2013). Nevertheless, using an array of advanced technologies, Luo et al. (2017) succeeded to obtain a reference-quality genome sequence for Ae. tauschii ssp. strangulata, accession AL8/78 which is closely related to the wheat D subgenome (see Chap. 3, Sect. 3.7). They show that the Ae. tauschii genome contains unprecedented amounts of very similar repeated sequences, a greater number of dispersed duplicated genes than other sequenced genomes, and its chromosomes have been structurally evolving an order of magnitude faster than those of other grass genomes. The decay of colinearity with other grass genomes correlates with recombination rates along chromosomes. These authors propose that the vast amounts of very similar repeated sequences cause frequent errors in recombination and lead to gene duplications and structural chromosome changes that drive fast genome evolution.

Zhao et al. (2017) generated a chromosome-scale, high-quality reference genome of Ae. tauschii, in which 92.5% sequences have been anchored to chromosomes. Using this assembly, they accurately characterized genic loci, gene expression, pseudogenes, methylation, recombination ratios, microRNAs and especially TEs on chromosomes. In addition to the discovery of a wave of very recent gene duplications, the authors revealed that TEs occurred in about half of the genes, and found that such genes are expressed at lower levels than those without TEs, presumably because of their elevated methylation levels. All wheat molecular markers and mapped allowing the authors to construct a high-resolution integrated genetic map corresponding to genome sequences, thereby placing previously detected agronomically important genes/ quantitative trait loci (QTLs) on the Ae. tauschii genome for the first time.

9.5.2.3 Crosses with Other Species of the Wheat Group

Intraspecific F₁ hybrids between accessions of Ae. tauschii that were collected from different regions, had regular chromosome pairing at meiosis (seven bivalents) and high fertility in most cases, but some hybrids were partially sterile (Kihara et al. 1965). Yet, F₁ hybrids involving one accession from Iran, had a ring of four chromosomes and five bivalents, indicating the presence of a reciprocal translocation in this accession (Kihara et al. 1965). Lagudah et al. (1991b) and Hohman and Lagudah (1993) found that this reciprocal translocation involves chromosomes 1DS-7DL and 7DS-1DL.

Chromosomal pairing at first meiotic metaphase of F₁ hybrids between Ae. tauschii and other species of the wheat group are characterized by relatively low pairing. The F₁ hybrid Ae. tauschii x Ae. bicornis had a low to intermediate level of chromosomal pairing at meiosis (0–6 bivalents and 0–1 trivalents; Kihara 1949). Evidently the D genome of Ae. tauschii has diverged quite considerably from the S^b genome of Ae. bicornis. Likewise, chromosomal pairing at first meiotic metaphase of F₁ hybrids between Ae. tauschii and Ae. caudata had 3–5 bivalents (Kihara 1949) or 3.60 bivalents, of which 0.60 were ring, 1.06 trivalents and 0.04 quadrivalents (Sears 1941b), indicating homoeology rather than homology of the genomes of these two diploid species. Divergence of the genome of Ae. tauschii from that of Ae. uniaristata (genome NN) is obvious from the low pairing in the F₁ hybrids between these two species. Kihara (1949) observed 1–5 bivalents and 0–1 trivalents and Sears (1941b) reported 3.82 bivalents, of which 0.20 were ring bivalents, and 0.14 trivalents. The F₁ hybrid between Ae. umbellulata (Genome UU) and Ae. tauschii had 3–5 bivalents with a mode of 3 bivalents and 1–3 trivalents (Kihara 1949), indicating low homology between the genomes of these two species. Pairing in the F₁ hybrid between Ae. geniculata (genome M^oM^oUU) and Ae. tauschii was also low, with 5–8 bivalents and 0–1 trivalents, (Kihara 1949), indicating that the two subgenomes of Ae. geniculata are not homologous to that of Ae. tauschii.

Chromosome pairing in Ae. tauschii x A. muticum hybrids was almost regular and exceeded the pairing in hybrids between Ae. mutica and Sitopsis species and between A. muticum and diploid Triticum species (Jones and Majisu 1968). Similarly, Ohta (1990) produced F₁ hybrids between Ae. tauschii and A. muticum (presumably without B chromosomes) and observed a relatively high amount of chromosomal pairing at first meiotic metaphase (Table 8.1). This high pairing presumably resulted from the homoeologous-pairing promoters of A. muticum. And indicates that the chromosomes of A. muticum appear to have considerable homoeology with the D genome of Ae. tauschii.

Data of chromosomal pairing in F₁ hybrids between Ae. tauschii and the allotetraploid species of section Vertebrata, namely, Ae. crassa 4x and Ae. ventricosa, are presented in Table 9.5. These pairing data and that of Kihara (1949) clearly show that the two allotetraploids contain one subgenome that is very closely related to the genome of Ae. tauschii. Moreover, the similarity of the spike morphology of these allotetraploids to that of Ae. tauschii further supports the conclusion that Ae. tauschii is the donor of one of the two subgenomes of these allotetraploids. Chromosomal pairing in the F₁ hybrids between Ae. tauschii and the allohexaploid species of section Vertebrata, namely, Ae. crassa 6x, Ae. juvenalis, and Ae. vavilovii, indicates that hexaploid Ae. crassa (genome D^cD^cX^cX^cDD) contains one unaltered and one modified subgenomes of Ae. tauschii, as suggested by Kihara (1949), whereas the other two allohexaploids, Ae. vavilovii (genome D^cD^cX^cX^cS^sS^s) and Ae. juvenalis (genome D^cD^cX^cX^cUU), contain a modified subgenome of Ae. tauschii (Kimber and Zhao. 1983) (Table 9.6). On the other hand, chromosomal pairing in the F₁ hybrid of Ae. cylindrica (genome DDCC) x Ae. tauschii showed 8–9 bivalents and up to 5 trivalents (Kihara 1949), indicating that one of the two genomes of Ae. cylindrica was donated by Ae. tauschii.

Chromosomal pairing in F₁ hybrids with the two subspecies of diploid wheat, T. monococcum (genome A^mA^m) domesticated ssp. monococcum and wild ssp. aegilopoides, showed that the genome of Ae. tauschii diverged from those of diploid wheat although somewhat less than its divergence from the genomes of other diploid Aegilops species (Table 9.8). Chromosomal pairing in F₁ hybrids between Ae. tauschii and the two allotetraploid species of Triticum, namely, the wild subspecies of T. turgidum, ssp. dicoccoides (genome BBAA), and the domesticated subspecies of T. timopheevii ssp. timopheevii (genome GGAA) showed that the subgenomes of the Triticum allotetraploids are not homologous to that of Ae. tauschii, although the pairing in the timopheevii x tauschii hybrid was somewhat higher than in the dicoccoides x tauschii hybrid (Table 9.9). On the other hand, chromosomal pairing in the F₁ hybrid with allohexaploid wheat, T. aestivum ssp. aestivum, showed that the allohexaploid contains one subgenome homologous to that of Ae. tauschii (Table 9.10).

9.5.3 Aegilops ventricosa Tausch

9.5.3.1 Morphological and Geographical Notes

Ae. ventricosa Tausch [Syn: Triticum ventricosum (Tausch) Ces., Pass. & Gibelli; Gastropyrum ventricosum (Tausch) Á. Löve] is an annual plant with few to many rather thick culms, that are 20–40-cm-high (excluding spikes). Its entire length is foliated with broad linear glabrous, seldom hairy, leaves. Spikes are long, moniliform, 4–6 (rarely up to 12)-cm-long, excluding awns, with 5–10 spikelets, more or less rough, and usually awned. Rudimentary spikelets are absent, rarely 1–2 are found. The spike disarticulates into individual spikelets, each falling with its adjacent rachis segment (barrel-type dispersal unit). Sometimes, the entire spike disarticulates at the base. Spikelets are oval, equal in length to the rachis segment, and suddenly become inflated in the lower parts (urn–shaped). There are 4–5 florets; the upper 1–3 are sterile. The glumes are strongly overlapping, with curved nerves, tips somewhat thickened with two teeth separated with a broad sinus; one tooth may be lengthened into an awn (up to 3-cm-long), that is usually shorter than the lemma awn (up to 4-cm-long). Tips of the glumes of the terminal spikelet are 3-toothed, the center one usually elongated into an awn. The lemmas are membranous, thickened in the upper parts, with weak keel at the tip, which is elongated into an awn, at the base of which are 1–2 small teeth. The awns are triangular, with lemma awns stronger than glume awns. All awns become longer toward the tip of the spike. The caryopsis adheres to the lemma and palea (Fig. 9.2c).

Ae. ventricosa exhibits limited morphological variation, mainly in narrowness of the upper part of the spikelets, spike length and width, spikelet number and awning. Some of the spike and spikelet characters are similar to those of Ae. uniaristata, one of the two diploid progenitors of this allotetraploid, but basic characters of the spikes and spikelets indicate that Ae. ventricosa morphology derived from Ae. tauschii, the second diploid parent of this allotetraploid species.

Ae. ventricosa is a Mediterranean element. It grows in the western and northern parts of the Mediterranean Sea, namely, Portugal, Spain, South France, Corsica, and Sardinia, rarely Italy and Sicily, Egypt (near Alexandria, rare), Libya, Tunisia, Algeria, and Morocco (Table 9.2). Ae. ventricosa grows on terra rossa, rendzina and light, sandy soils in the edges and openings of deciduous and sclerophyllous Mediterranean forests and maquis, in degraded dwarf-shrub and semi-steppical formations, pastures, abandoned fields, edges of cultivation, disturbed habitats and roadsides. It is common in a wide array of habitats and invades wheat fields, vineyards, and olive groves as weed. Ae. ventricosa develops well in areas with 200–350 mm annual rainfall but was also found in areas with less than 100 mm and up to 600 mm annual rainfall. It is found in a range of altitudes, from sea level to 1850 m above sea level.

Ae. ventricosa has a medium-sized distribution in the western part of the distribution of the genus. It is relatively isolated from the rest of the group. Interestingly, its distribution does not overlap with that of either of its diploid progenitors (Ae. tauschii and Ae. uniaristata), and it is larger than that of Ae. uniaristata, but smaller than that of Ae. tauschii. Its distribution area is closer to the distribution area of Ae. uniaristata than to that of any other diploid species (excluding Ae. bicornis, with which it may have contact in Libya). Ae. ventricosa distributes sympatrically with other Aegilops tetraploid species, i.e., Ae. geniculata, Ae. neglecta, Ae. biuncialis, Ae. triuncialis, Ae/ perigrina and Ae. cylindrica, and allopatrically with Ae. bicornis, Ae. recta and Ae. kotschyi (in Libya and Tunisia). Natural hybridizations are known with many species, including domesticated tetraploid and hexaploid wheat. Several economically important traits, such as resistance to eyespot, stripe-, leaf- and stem-rust, and tolerance to aluminum, were discovered in Ae. ventricosa (e.g., Maia 1967; Dosba et al. 1980), some of which were already transferred to common wheat (e.g., Jahier et al 1978, 1996; Bariana and Mcintosh 1993; Tanguy et al. 2005).

9.5.3.2 Cytology, Cytogenetics, and Evolution

Ae. ventricosa is an allotetraploid species (2n = 4x = 28; genome DDNN) that evolved as a result of hybridization between two different diploid Aegilops species, followed by chromosome doubling. The existence of a D subgenome in Ae. ventricosa was first directly proved by the observation by Kihara, in 1938, of seven ring bivalents in the Ae. tauschii x Ae. ventricosa F₁ hybrid (see Kihara 1949). The second subgenome of this allotetraploid was first believed to be a modified M genome, designated M^v, derived from Ae. comosa, which underwent changes at the polyploid level (Kihara 1949, 1954, 1963; Kihara and Lilienfeld 1932). However, Chennaveeraiah (1960), Kimber et al. (1983), Kimber and Zho (1983), Zhao and Kimber (1984) and Yen and Kimber (1992b) demonstrated that the second subgenome in Ae. ventricosa is actually a N genome from Ae. uniaristata. Thus, it is currently accepted that the genomic constitution of Ae. ventricosa is DDNN (Kimber and Tsunewaki 1988; Dvorak 1998). Its organellar genome is similar to that of Ae. tauschii, which was the female parent in the formation of Ae. ventricosa, and consequently, was designated D by Ogihara and Tsunewaki (1988) and Wang et al. (1997).

Matsumoto and Kondo (1942) synthesized an amphiploid of Ae. tauschii – Ae. uniaristata, and Matsumoto et al. (1957) synthesized the amphiploid of the reciprocal combination. Morphologically, both synthetic amphiploids were very similar to Ae. ventricosa. The synthetic amphiploids exhibited almost regular pairing at meiosis and had about 60% seed fertility. Hybrids between the synthetic and natural Ae. ventricosa exhibited chromosomal pairing at meiosis that was quite good (average of 12 bivalents and 0–3 trivalents or quadrivalents per cell). In spite of the high pairing, seed fertility was very low (3–5%) (Matsumoto et al. 1957).

Ae. ventricosa has 10.64 pg 1C DNA (Eilam et al. 2008; Table 9.3). This allotetraploid species has 3.18% less DNA than expected from the DNA sum of its two diploids parents, i.e., 10.99 pg (Ae. tauschii contains 5.17 pg and Ae. uniaristata contains 5.82 pg; Eilam et al. 2007). The loss of DNA in the allotetraploid was confirmed by Badaeva et al. (2012) who, based on differential C-banding and in situ hybridization, found that Ae. ventricosa exhibits substantial structural chromosome rearrangements, including deletion of chromosomal segments and reduction of heterochromatin content.

Earlier karyomorphological studies (Emme 1924; Sorokina 1928; Senyaninova-Korchagina 1932) did not find satellites in Ae. ventricosa. Since there is no species of Aegilops that is without at least one pair of satellites, it is quite unlikely that this species is without any satellites. Indeed, the karyotype of this allotetraploid species contains one chromosome pair with a satellite on the short arm (Chennaveeraiah 1960). In agreement with this finding, Orellana et al. (1984), using a silver-staining procedure, analyzed the activity of the nucleolar organizer of Ae. ventricosa and detected only one pair of Ag-NORs, indicating that natural amphiplasty occurs in this allotetraploid species.

The karyotype of Ae. ventricosa is asymmetric, consisting of five pairs with submedian-subterminal centromeres and the rest with submedian centromeres (Chennaveeraiah 1960). One half of the set consist of seven pairs with submedian centromeres, resembling the chromosome set of Ae. tauschii, whereas the other seven pairs closely resemble the set in Ae. uniaristata (Chennaveeraiah 1960). Hence, the karyotype of the D subgenome chromosomes of Ae. ventricosa is symmetric, consisting of metacentric and submetacentrics chromosomes, whereas the karyotype of the N subgenome is asymmetric; five chromosomes are subtelocentrics (Chennaveeraiah 1960). Though this chromosome set agrees mostly with the chromosome set of Ae. uniaristata, it differs from it in the absence of a satellite pair (Chennaveeraiah 1960; Badaeva et al. 2012). Therefore, the N subgenome of Ae. ventricosa is not completely homologous with the genome of its diploid progenitor Ae. uniaristata, but it is slightly modified.

Meiotic pairing analysis revealed only minor modification of the D and N subgenomes in Ae. ventricosa (Kimber et al. 1983; Kimber and Zhao 1983; Zhao and Kimber 1984; Yen and Kimber 1992b). Yet, Badaeva et al. (2002, 2011, 2012), using the C-banding technique and fluorescence in situ hybridization with clones pTa71 (18S-5.8S-26S rDNA), pTa794 (5S rDNA), both clones from T. aestivum, and pAs1 (non-coding repetitive DNA sequence from Ae. tauschii) as probes, found that the N subgenome of Ae. ventricosa substantially differed from that of Ae. uniaristata. The D subgenome also differed, albeit to a lesser extent, from the genome of Ae. tauschii. They observed polymorphism for the presence and size of C-bands and for chromosome aberrations in these subgenomes, including minor changes in the C-banding and pAs1-FISH patterns, complete deletion of the NOR on chromosome 5D and the loss of several minor 18S-5.8S-26S rDNA loci on N subgenome chromosomes. In addition, several chromosomal translocations involving different chromosomes and a pericentric inversion of chromosome 5N, were identified in several accessions of Ae. ventricosa (Bardsley et al. 1999; Badaeva et al. 2012).

Cuñado et al. (1986) pointed out that two chromosome groups in Ae. ventricosa have a distinct C-banding pattern, i.e., 16 chromosomes with prominent C-heterochromatin located in centromeric and pericentromeric regions and 12 almost entirely euchromatic chromosomes. These differences may distinguish between the N and D subgenomes, respectively, although at least one of the heterochromatic chromosomes should belong to the D genome. These different C-banding patterns allowed for estimation of the frequencies of homologous and homoeologous pairing in the hybrids. The N subgenome chromosomes of Ae. ventricosa are also characterized, in addition to their distinct asymmetry, by large pericentric heterochromatin complexes, like in Ae. uniaristata (Badaeva et al. 2012). In contrast, the D subgenome chromosomes of Ae. ventricosa have only a few small, predominantly intercalary C bands, like the chromosomes of Ae. tauschii. Yet distinct reproducible changes associated with chromosome modification that accompanied the formation of the allopolyploid species, were observed in the two Ae. ventricosa subgenomes. As apparent from the appearance or disappearance of C-bands and from changes in band size, the heterochromatin content decreased in some N subgenome chromosomes (1N, 3N, and 7N) and increased in others (2N, 4N, 5N, and 6N). The greatest changes in C banding pattern were observed for chromosome 3N, which lost the large pericentric heterochromatin block in the short arm and the distal C-band marker in the long arm, while several new bands appeared instead (Badaeva et al. 2012). In situ hybridization with the pAs1 clone revealed changes in the N subgenome of Ae. ventricosa, manifested by fewer hybridization sites or less intense signals (Badaeva et al. 2012). The greatest changes were observed for chromosomes 1N, 4N, 6N, and 7N. In addition, the minor NORs of chromosomes 1N and 7N were partly lost, but new, earlier unidentified minor 45S rDNA sites, appeared in the subterminal regions of the short and long arms of Ae. ventricosa chromosome 3D (Badaeva et al. 2011).

Bardsley et al. (1999) analyzed lines of Ae. ventricosa using fluorescent in situ hybridization with probes including rDNA, repeated sequences from wheat and rye, simple-sequence repeats (SSRs) and total genomic DNA. They found that the banding patterns could be used to distinguish most chromosome arms of this allotetraploid. All lines had a single major 18S-25S rDNA site, the nucleolar organizing region (NOR) in chromosome 5N and several minor sites of 18S-25S rDNA and 5S rDNA. A 1NL.3DL-1NS.3DS translocation was identified, and other minor differences were found between the lines.

9.5.3.3 Crosses with Other Species of the Wheat Group

Crosses between Ae. ventricosa with its parental diploids, namely, Ae. ventricosa x Ae tauschii and Ae. ventricosa x Ae uniaristata, showed that the allotetraploid contains one subgenome similar to that of Ae. tauschii and another alike the genome of Ae. uniaristata (Table 9.5). The F₁ hybrid Ae. ventricosa x Ae. tauschii had 6 bivalents and 1 trivalent at meiosis (Kihara 1949; Kimber and Zhao 1983) and the F₁ hybrid between Ae. ventricosa and Ae. uniaristata also had a similar level of pairing, i.e., 6 bivalents and a trivalent (Kimber et al. 1983). Since the F₁ hybrid between the two diploid progenitors of Ae. ventricosa, Ae. tauschii and Ae. uniaristata, exhibited reduced pairing, i.e., 3.82 bivalents (Sears 1941b), indicating homoeologous relationships between the D and the N genomes, the pairing in the triploid hybrids between Ae. ventricosa and its parental diploids was rather homologous. The chromosome pairing in the tetraploid hybrid between Ae. ventricosa and Ae. crassa (4x) (genome D^cD^cX^cX^c) (about 5 bivalents and up to 2 trivalents; Kimber and Zhao 1983) also indicates that the two allotetraploid species share one subgenome, most probably D (Table 9.7).

Chromosome pairing in the F₁ hybrid between tetraploid wheat (T. turgidum subsp. dicoccoides) (genome BBAA) and Ae. ventricosa was very low [0–2 (mode of 0) bivalents per cell] (Kihara 1949). On the other hand, the F₁ hybrid between hexaploid wheat, T. aestivum ssp. aestivum (genome BBAADD), and Ae. ventricosa showed 5 bivalents and up to 2 trivalents (Kimber and Zhao 1983), indicating that allohexaploid wheat has one subgenome homologous to one of the subgenomes of Ae. ventricosa. Since the hybrid with tetraploid wheat had very little pairing, the good pairing in the hybrid with hexaploid wheat is between the D subgenomes of ssp. aestivum and that of Ae. ventricosa.

9.5.4 Aegilops crassa Boiss.

9.5.4.1 Morphological and Geographical Notes

Ae. crassa Boiss. (Persian goat grass) [Syn: Triticum crassum (Boiss.) Aitch. & Hemsl.; Triticum syriacum Bowden; Ae. platyathera Jaub. & Spach; Castropyrum crassum (Boiss.) Á. Löve] is an annual robust plant with many-jointed culms, sometimes thick culms, that are 20–40-cm high (excluding spikes) and often foliated along the entire length. The spikes are usually long (4–10-cm, excluding awns), with 6–10 spikelets, thick, cylindrical, somewhat like a string of beads (moniliform), tapering toward the tip, usually hairy, with barrel-shaped disarticulation, and each spikelet falling with its adjacent rachis segment. Rudimentary spikelets are absent, or rarely, there are 1–2. There are 3–5 florets, with the upper 1–3 being sterile. Glumes are somewhat or not fully overlapped, covered with fine silvery hairs, with a tip blunt, thickened, with 1–4, usually 2, teeth, that are separated by a broad and shallow sinus, and usually not awned or seldom with a short awn. Glume awns are slender, triangular and weaker than the lemma awns of the same spikelet. Lemmas are membranous or cartilaginous, somewhat keeled, usually tipped with a tooth or an awn, flanked by 1–2 lateral teeth. The lemma awns are mostly broad and more strongly developed in the upper spikelets. The caryopsis adheres to lemmas and palea (Fig. 9.2d).

Ae. crassa is morphologically closer to Ae. tauschii than Ae. ventricosa. It is a most variable species, varying in all the diagnostic elements, such as spike and spikelet size, form, structure, color, glume and lemma awn development, form, and place of attachment. The wide morphological variation of Ae. crassa led Eig (1929a) to recognize two varieties within this species: var. typica and var. palaestina (= ssp. vavilovii Zhuk.). [The later was elevated to the specific rank by Chennaveeraiah (1960), who designated it Ae. vavilovii (zhuk.) Chennav.]. Hammer (1980) split Ae. crassa into two subspecies: ssp. crassa, with three varieties (var. crassa (=var. typica), var. glumiaristata Eig. and var. macrathera Boiss.) and subsp. vavilovii Zhuk. Var. typica was subdivided by Hammer (1980) into f. crassa, f. rubiginosa (Popova) Hammer and f. fuliginosa (Popova) Hammer. Indeed, a very wide variation in Ae. crassa from Iran was found in morphological elements (Ranjhar et al. 2007; Bordbar and Rahiminejad 2010), in simple sequence repeat (SSR) DNA markers (Naghavi et al. 2009a, b), in SSR and ISSR markers (Moradkhani et al. 2015), in nuclear microsatellite loci, nuclear rDNA ITS, and in chloroplast trnL-F sequences (Bordhar et al. 2011). The wide variation of Ae. crassa may has accumulated throughout the lifetime of this species, which is considered very old (Tsunewaki 1993, 2009).

Ae. crassa is a steppical (Irano-Turanian) element, penetrating into semi-desert regions. It has a relatively large distribution area in the eastern part of the distribution of the genus, i.e., in western Asia. It grows in Transcaucasia (Georgia, Armenia, and Azerbaijan), southern Turkmenistan, southern Uzbekistan, southernmost part of Kazakhstan, northern Tajikistan, western Kyrgyzstan, Afghanistan, Iran, central and northern Iraq, northern and northeastern Syria and southeastern Turkey. It is rarely present in Jordan and Lebanon. Ae. crassa inhabits a wide range of primary and secondary habitats. It grows on grey-calcareous, loess, and alluvial soils and on stony slopes and gravel, in degraded deciduous steppe maquis, Juniperous forests, dwarf-shrub steppe-like formations, steppical plains, wadis, edges of cultivation, disturbed habitats and roadsides. It is a common weed of cultivation.

Ae. crassa is considered a drought-tolerant species because it grows in areas with 150–350 mm rainfall (Kilian et al. 2011). In this regard, Harb and Lahham (2013) reported that several accessions of Ae. crassa from semiarid and arid areas in Jordan, are drought-tolerant and can be used to improve this trait in domesticated wheat. (A likely possibility is that these authors refer to Ae. vavilovii).

The distribution area of Ae. crassa is somewhat smaller than that of Ae. tauschii, but much larger than that of the M-genome species (Kihara 1963). It grows sympatrically with the following species: Ae. columnaris, Ae. triuncialis, Ae. tauschii, Ae. cylindrica and Ae. juvenalis, and allopatrically with Ae. speltoides, Ae. caudata, Ae. umbellulata, Ae. geniculata, Ae. biuncialis, Ae. neglecta, Ae. vavilovii, wild T. monococcum (subsp. aegilopoides), and wild T. timopheevii (subsp. armeniacum).

Ae. crassa contains two cytotypes, an allotetraploid (2n = 4x = 28; genome D^cD^cX^cX^c) and an auto-allohexaploid (2n = 6x = 42; genome D^cD^cX^cX^cDD) (Table 9.3), that are very similar morphologically. Yet, there is a morphological difference between the two cytotypes; the allotetraploid has more robust spikes and has a greater tendency to display moniliform spikes, while the auto-allohexaploid has more or less cylindrical spikes (Kihara 1963).

The allotetraploid cytotype was found throughout the species distribution area, whereas the distribution of the auto-allohexaploid cytotype is restricted to northern Afghanistan and northeastern Iran (Kihara 1963). This difference in the size of the distribution areas presumably results from the age of the two cytotypes; the tetraploid is an old polyploid, while the hexaploid is much younger (Tsunewaki 1993, 2009). In northern Afghanistan and northeastern Iran, there are mixed populations of the hexaploid and the tetraploid cytotypes. It is assumed therefore, that the hexaploid cytotype was formed there. Variation analysis of the restriction profiles of nuclear repeated nucleotide sequences support this assumption (Dubcovsky and Dvorak 1995a). The allotetraploid Ae. crassa appears to be an ancient allotetraploid (Tsunewaki 1993, 2009) that originated from hybridization of primitive Ae. tauschii with an ancient species in the evolutionary lineage, leading to the section Sitopsis (Dubcovsky and Dvorak 1995a). The auto-allohexaploid cytotype is a young taxon. According to Kihara (1954), the tetraploid originated in Asia Minor and the hexaploid in northern Afghanistan.

9.5.4.2 Cytology, Cytogenetics, and Evolution

The allotetraploid cytotype of Ae. crassa (2n = 4x = 28; genome D^cD^cX^cX^c) evolved as a result of hybridization between two different diploid Aegilops species, and the auto-allohexaploid cytotype (2n = 6x = 42; genome D^cD^cX^cX^cDD) was formed as a result of hybridization between the allotetraploid cytotype and Ae. tauschii. The F₁ hybrids of both hybridizations, i.e., between the two diploid parents of the allotetraploid and between the allotetraploid and Ae. tauschii, are sterile, but chromosome doubling led to the production of fertile polyploids. The existence of a D subgenome in the allotetraploid Ae. crassa was demonstrated by the observation of seven ring bivalents in the F₁ hybrid between Ae. crassa 4x and Ae. tauschii (Kihara 1949, 1957). The C-banding and ISH results reported by Badaeva et al. (1998) support the genome analysis data of Kihara (1949), demonstrating that one of the two subgenomes of 4x Ae crassa was derived from the D genome of Ae. tauschii. Yet, the D subgenome of the allotetraploid differentiated from that of the diploid progenitor (Dubcovsky and Dvorak 1995a) and consequently, was designated D^c by Dvorak (1998). The second subgenome of this allotetraploid was first believed to be a modified M genome of Ae. comosa, designated M^cr (Kihara 1940a, 1957, 1963; Lilienfeld 1951; Kihara et al. 1959; Kihara and Tanaka 1970; Kimber and Feldman 1987), but analysis of variation in nuclear repeated nucleotide sequences (Dvorak 1998 and reference therein) showed this subgenome to differ from the M genome and instead, to be related to genome S of the Sitopsis species. It is assumed therefore, that the X^c genome derived from an extinct ancestor of section Sitopsis and was tentatively designated X^c until its equivalence to a Sitopsis genome will be verified by further studies (Dubcovsky and Dvorak 1995a; Dvorak 1998). Yet, the X^c subgenome exhibits a large number of intra-subgenomic rearrangements, making it difficult to identify the origin of this subgenome (Badaeva et al. 1998). Hexaploid Ae. crassa derived from hybridization between the tetraploid cytotype of Ae. crassa and Ae. tauschii. The speciation of 6x Ae. crassa was accompanied by a reciprocal translocation which is specific for the 6x cytotype. The distinct genetic differences between the tetraploid and hexaploid cytotypes of Ae. crassa would justify classifying them as different subspecies (Badaeva et al. 1998) or even different species.

The D^c subgenomes of 4x and 6x Ae. crassa are highly modified compared with the D genome of the progenitor species Ae. tauschii (Kihara 1940a, 1957; Lilienfeld 1951; Siddique and Jones 1967; Kihara and Tanaka 1970; Chapman and Miller 1978; Nakai 1982; Kimber and Zhao 1983; Zhao and Kimber 1984; Kimber and Feldman 1987; Zhang and Dvorak 1992; Tsunewaki 1993). In situ hybridization with the D-genome-specific DNA clone pAs1 confirmed that the D^c subgenome of tetraploid Ae. crassa is significantly different from the ancestral D genome of Ae. tauschii (Badaeva et al. 1998). The D subgenome of 6x Ae. crassa is different from the D^c subgenome and is similar to the D-genome of Ae. tauschii.

The organellar genome of the allotetraploid cytotypes of Ae. crassa is a subtype of the organellar genome D of Ae. tauschii, designated D² (Ogihara and Tsunewaki 1988; Wang et al. 1997). It is similar to that of Ae. tauschii, which was the female parent in the formation of 4x Ae. crassa, but diverged from it somewhat. The organellar genome of the hexaploid cytotype is similar to that of the allotetraploid and, consequently, designated D² (Ogihara and Tsunewaki 1988; Wang et al. 1997). This indicates that the allotetraploid was the female parent in the hybridization that led to the formation of the auto-allo-hexaploid.

Analysis of meiotic chromosome pairing in F₁ hybrids between 4x and 6x Ae. crassa confirmed that the hexaploid forms were derived from a hybridization between 4x Ae. crassa and Ae. tauschii (Kihara et al. 1959; Kihara 1963; Kimber and Zhao 1983; Zhao and Kimber 1984). The F₁ hybrid between the two cytotypes showed up to 14 bivalents and several multivalents (Kihara 1963), indicating that genome D^cD^cX^cX^c of the tetraploid is homologous to the D^cD^cX^cX^c subgenomes of the hexaploid cytotype, and that the D^c subgenome of the hexaploid is related to the D^c subgenome of the tetraploid.

A synthetic auto-allohexaploid was produced by crossing the allotetraploid cytotype of Ae. crassa with Ae. tauschii (Shigenobu and Sakamoto 1977). This synthetic hexaploid (genome D^cD^cX^cX^cDD) closely resembles the natural auto-allohexaploid cytotype of Ae. crassa (Kihara 1963). The F₁ hybrid between the synthetic and the natural hexaploid had 19 bivalents at meiosis and had high fertility (89%) (Kihara 1963). Since there are mixed populations of tetraploid Ae. crassa and Ae tauschii in northern Afghanistan and northeastern Iran, they had many opportunities to hybridize and to form the hexaploid cytotype (Kihara (1963).

The allotetraploid cytotype of Ae. crassa has 10.86 pg 1C DNA (Eilam et al. 2008; Naghavi et al. 2013) and the auto-allohexaploid has 15.90 pg 1C DNA (Naghavi et al. 2013) (Table 9.3). The expected amount of 1C DNA in the tetraploid cannot be calculated since the donor of the X^c subgenome has yet to be identified. The expected amount of 1C DNA in the hexaploid cytotype is 16.03 pg (10.86 pg of 4x + 5.17 pg of Ae. tauschii; Eilam et al. 2007), 0.82% higher than the content in the natural hexaploid Ae. crassa. The loss of DNA in the two cytotypes was confirmed by Badaeva et al. (1998) who, based on differential C-banding and in situ hybridization, found that Ae. crassa exhibits substantial structural chromosome rearrangements, including deletion of chromosomal segments and reduction of heterochromatin content.

The correct number of chromosomes in the allotetraploid Ae. crassa was first reported by Emme (1924), but this report did not mention the presence of satellites. Chennaveeraiah (1960) described the correct karyotype, which is symmetric, consisting of two pairs with satellites of different sizes on short arms, one pair with a median centromere, and one pair with an almost median centromere. Three chromosome pairs with satellites exist in the hexaploid Ae. crassa. The remaining chromosomes in both cytotypes have submedian centromeres. There are no subterminal centromeres in the set.

With the aim of analyzing the activity of the nucleolar organizer regions (NORs), Cermeño et al. (1984b performed a comparative analysis of somatic metaphase chromosomes in 4x and 6x Ae. crassa by phase contrast, C-banding and Ag-staining. They found that the nucleolar activity of chromosome “C” of the X^c subgenome was much higher than that of chromosome “B” of the D^c subgenome, indicating the incidence of partial amphiplasty. The NORs activity of the D subgenome chromosomes are also inhibited in the 6x Ae. crassa (Cermeño et al. 1984b). ISH analysis using the pTa71 clone, containing 18S, 5.8S, and 26S rDNA loci, detected a third NOR locus in 4x Ae. crassa, but this locus is not actively transcribed (Yamamoto and Mukai 1995).

Using C-banding and in situ hybridization (ISH) analyses, Badaeva et al. (1998) studied the distribution of highly repetitive DNA sequences on chromosomes of the tetraploid and the hexaploid cytotypes of Ae. crassa. The ISH studies were carried out with the pSc119 [120-bp sequence from Secale cereale (Bedbrook et al. 1980)], pAs1 [1 kb from Ae. tauschii (Rayburn and Gill 1986)] and pTa794 [410-bp from T. aestivum containing the 5S rRNA gene unit (120-bp) separated by a 290-bp spacer (Gerlach and Dyer 1980)] DNA clones. All chromosomes were identified by their C-banding and ISH pattern with the pAs1 clone, and the position of C-bands generally coincided with the location of the pAs1 sequence (Badaeva et al. 1998). Only a few pSc119 hybridization sites were observed in the telomeric regions of several chromosomes. Since pSc119 hybridizes with all S-genome chromosomes of Sitopsis species (Badaeva et al. 1996a), the few minor pSc119 ISH sites that were detected in 4x Ae. crassa, weaken the assumption that X^c subgenome originates from a Sitopsis species. Three pTa794 ISH sites (5S rDNA) were detected in tetraploid Ae. crassa and five such sites were identified in the hexaploid cytotype, three having derived from 4x Ae. crassa, and two contributed by Ae. tauschii (Badaeva et al. 1998). All the hexaploid accessions differed from the tetraploids by a reciprocal non-centromeric translocation. Several additional intraspecific translocations were detected between 4 and 6X accessions (Badaeva et al. 1998).

Using C-banding and fluorescence in situ hybridization (FISH) with ten DNA probes, Badaeva et al. 2021b) studied genome structure of 4x Ae. crassa. They confirmed that the D^c subgenome of allotetraploid Ae. crassa was contributed by Ae. tauschii, although the retention of minor NORs on chromosomes 1D^c and 6D^c indicated that Ae. crassa probably emerged prior to the loss of the respective loci in the diploid progenitor. Subgenome X^c might have originated from an ancestral S-genome species of subsection Emarginata.

9.5.4.3 Crosses with Other Species of the Wheat Group

Upon crossing tetraploid Ae. crassa with Ae. tauschii the F₁ triploid hybrids exhibited 7 bivalents (Kihara 1949) or 5.03 bivalents, 0.91 trivalents and 0.07 quadrivalents at first meiotic metaphase (Kimber and Zhao 1983). This high chromosomal pairing indicates that tetraploid Ae. crassa contains a subgenome that is related to the Ae. tauschii genome. Another triploid hybrid involving tetraploid Ae. crassa and Ae. bicornis showed 2–7 (mode of 5) bivalents, 0–2 trivalents, and 0–2 quadrivalents (Kihara 1949). A similar low level of pairing was also observed in the hybrid between Ae. crassa 6x and Ae. bicornis, namely, 3–6 bivalents (Kihara 1949), indicating that the two subgenomes of tetraploid and the three subgenomes of hexaploid Ae. crassa are not homologous to the S^b genome of Ae. bicornis. Chromosomal pairing between tetraploid Ae. crassa and Ae. uniaristata [3.89 bivalents, 0.61 trivalents, and 0.07 quadrivalent (Kimber et al. 1983)] and between hexaploid Ae. crassa and Ae. uniaristata [6–9 bivalents and 3 trivalents (Kihara 1949)], show that the genomes of the two species are not homologous. On the other hand, Kimber and Zhao (1983) studied chromosomal pairing in the hybrid between Ae. cylindrica and tetraploid Ae. crassa (Table 9.7), and hound that the two-allotetraploid species share one homologous subgenome, subgenome D.

Gupta and Fedak (1985) studied meiotic chromosomal pairing in F₁ hybrids between hexaploid Ae. crassa and species of Secale. The chiasmata frequency per cell ranged from 6.86 in hybrids with S. cereale to 9.93 in hybrids with S. strictum. These results provide evidence that a homoeologous-pairing control system operates in Ae. crassa. However, Cuñado (1992) and Cuñado et al. (2005) reported that chromosomal pairing at first meiotic metaphase of tetraploid Ae. crassa was as follows: 0.01 univalents, 3.02 rod bivalents, 10.88 ring bivalents and 24.78 chiasmata/cell. That of 6x Ae. crassa: 0.12 univalents, 4.27 rod bivalents, 15.78 ring bivalents, 0.39 multivalents and 36.98 chiasmata/cell. The multivalent pairing in 6x Ae. crassa presumably resulted from the partial homology between some chromosomes of D^c and D subgenomes, indicates that the cytologically-diploidizing genetic system is not fully effective in hexaploid crassa.

9.5.5 Aegilops vavilovii (Zhuk.) Chennav.

9.5.5.1 Morphological and Geographical Notes

Aegilops vavilovii (Zhuk.) Chennav. [Syn.: Ae. crassa Boiss. subsp. vavilovii Zhuk.; Ae. crassa var. palaestina Eig; Triticum syriacum Bowden; Gastropyrum vavilovii (Zhuk.) Á. Löve] is an annual robust plant, with thick 20–40-cm-high culms (excluding spikes). Spikes are 10–15-cm-long (excluding awns), with a cylindrical or slightly zigzag-shaped, tapering towards the tip, disarticulating into individual spikelets at maturity, each with its adjacent rachis segment (barrel-type dispersal unit). There are 5–10 spikelets, with 1–2, and usually 1, basal rudimentary spikelets. Spikelets are 12–14-mm-long, with 4–5 florets, linear, cylindrical to slightly inflated at the base, and slightly overlapping glume edges. Glumes are nearly truncate, covered with fine silvery hairs, and membranous at the tip, and are approximately 2/3 to ¾ as long as the lemmas, with 2–3 teeth (or weak awn) and a shallow sinus between the teeth and with veins equal in width. Lemmas are thickened in the upper part and usually keeled, ending with a tooth, which may be extended into a small awn. Lemmas of the terminal spikelet end with a strong, broad 5–6-cm-long awn, flanked with two small teeth, and has a prominent central nerve. The caryopsis adheres to lemma and palea (Fig. 9.2e).

Ae. vavilovii has limited morphological variation. It differs from Ae. crassa by characteristics derived from its S^s subgenome parent, i.e., Ae. searsii, including its long cylindrical and somewhat zig-zagged spike, that tapers in the upper half, and by the absence of awns on the lateral spikelets, while those of the uppermost spikelet are very long.

Ae. vavilovii is an Irano-Turanian (steppical) element. It has relatively limited distribution in the south-central part of the distribution area of the genus, namely, in the southeastern Mediterranean. Tetraploid Ae. crassa and Ae. searsii, the two parental species, may have contact in eastern Syria, where Ae. vavilovii presumably originated and spread southwards to high elevations of the southeastern Mediterranean steppes and dry and semi–desert habitats, where it is relatively isolated geographically from most of the other species of the group. Its distribution partially overlaps with that of its diploid parent Ae. searsii, and is south to the distribution of its tetraploid parent, Ae. crassa.

Ae. vavilovii grows in Egypt (Santa Katerina, Sinai Peninsula), south and southeastern Israel, Jordan, Syria, possibly also in eastern Lebanon, rarely in southeastern Turkey (close to the Syrian border) and in western Iraq (Table 9.1). It grows on grey calcareous, rendzina, alluvial or sandy soils, rarely on basalt, in the edges of dwarf-shrub steppe-like formations, wadis, stony slopes, fallows, grasslands, roadsides, edges of cultivation, and disturbed habitats. Populations of Ae. vavilovii may vary from small and scattered to large and dense stands, sometimes intermingled with Ae. searsii (in the higher elevations of the Israeli Negev and in southern Jordan) or with Ae. crassa in northeastern Syria. It grows at altitudes of 275–1550 m, in areas with 100–275 mm annual rainfall (and up to 550 mm in some higher locations). As such, it might be a drought-tolerant species. Ae. vavilovii grows sympatrically with Ae. searsii, Ae. longissima and Ae. kotschyi, and allopatrically with Ae. peregrina, Ae. crassa, and possibly Ae. triuncialis.

9.5.5.2 Cytology, Cytogenetics, and Evolution

Ae. vavilovii is an allohexaploid species (2n = 6x = 42: genome D^cD^cX^cX^cS^sS^s) (Dvorak 1998). The D^cD^cX^cX^c subgenomes derived from 4x Ae. crassa (Kihara 1957; Chennaveeraiah 1960; Chapman and Miller 1978; Kimber and Zhao 1983; Zhang and Dvorak 1992). Analysis of variation in 27 repeated nucleotide sequences and the 5S rRNA demonstrated that the D^c subgenome was contributed by ancient Ae. tauschii, and the X^c subgenome by an extinct species, possibly a species that was ancestral to the entire genus Aegilops. The origin of the third subgenome was a source of controversy; Kihara (1957), Kihara et al. (1959) and Nakai (1982) proposed that it derived from the D genome of Ae. tauschii, whereas Kihara (1963) Kihara and Tanaka (1970), Talbert et al. (1991) and Yen and Kimber (1992a) thought that it derived from Ae. longissima. However, recent molecular studies by Zhang and Dvorak (1992) and Dubcovsky and Dvorak (1995a) showed that the third subgenome of Ae. vavilovii derived from Ae. searsii. Similar conclusions were reached following C-banding analysis (Badaeva et al. 2002). In addition, FISH analysis confirmed the relationship between Ae. vavilovii, 4x Ae. crassa and Ae. searsii (Badaeva et al. 2002). The organellar genome is D², similar to that of Ae. crassa. (Ogihara and Tsunewaki 1988; Wang et al. 1997). Hence, Ae. vavilovii was formed through hybridization of 4x Ae. crassa (as female) and Ae. searsii (Zhang and Dvorak 1992; Dubcovsky and Dvorak 1995a; Badaeva et al. 2002). Slight variations in the C-banding pattern relative to that of the parental species Ae. crassa, were detected in Ae. vavilovii, as was the existence of intraspecific variation in Ae. vavilovii due to a translocation between chromosomes 3X^c and 3D^c (Badaeva et al. 2002). While Gong et al. (2006), using ISSR markers, reported that the D^c subgenome of Ae. vavilovii changed only slightly from the D genome of Ae. tauschii, studies of chromosomal pairing showed that the D^c subgenome of Ae. vavilovii may have been substantially modified from that of the D genome of Ae. tauschii (Zhao and Kimber 1984).

Karyotypically, Chennaveeraiah (1960) noticed that the morphology of the chromosomes of two subgenomes of Ae. vavilovii (formerly Ae. crassa subsp. vavilovii Zhuk. Or Ae. crassa var. palaestina Eig) resembled that of 4x Ae. crassa, and that that of the third subgenome was different from that of the D genome that exists in 6x Ae. crassa. Therefore, he separated this taxon from Ae. crassa and elevated it to the species rank, designated Ae. vavilovii. Kihara (1963) reported that Ae. vavilovii has features quite different from those of 6x Ae. crassa, and consequently, supported Chennaveeraiah’s decision to consider it as a separate species. A similar conclusion, based on cytological studies, was reached by Kihara and Tanaka (1970).

The Ae. vavilovii karyotype is symmetric, consisting of three pairs with median centromeres, and the rest with submedian centromeres (Chennaveeraiah 1960). While Sorokina (1928) noticed the presence of a single pair with a satellite, Senyaninova-Korchagina (1932) and Pathak (1940) observed three pairs with satellites. The presence of three pairs with satellites was also confirmed by Chennaveeraiah (1960). Two of the three satellite pairs are large and somewhat similar to each other, whereas those on the third pair are smaller and different. It is the smaller type that corresponds to the satellite of Ae. tauschii and is located on chromosome pair 5D^c.

Multicolor FISH analysis with probes pTA794 [410-bp-long sequence containing 5S rDNA from T. aestivum (Gerlach and Dyer 1980)] and pTa71 [9 kb of 18S-5.8S-26S rDNA from T. aestivum (Gerlach and Bedbrook 1979)] revealed six 5S rDNA loci and four 18S-5.8S-26S rDNA sites in Ae. vavilovii. Since three pairs of satellited chromosomes were observed in Ae. vavilovii (Chennaveeraiah 1960), the existence of only four 18S-5.8S-26S rDNA loci show that not all three pairs of NORs are active in organizing nucleoli (Badaeva et al. 2002). The reduction in the number of active NORs in Ae. vavilovii suggests that this locus was inactivated as a result of amphiplasty.

Usually, 21 bivalents per cell are formed at first meiotic metaphase, of which 4–5 are rod bivalents and the rest are ring bivalents. In some cells, two univalents were formed, one forming a ring univalent, indicative of an isochromosome (Chennaveeraiah 1960). Chapman and Miller (1978) observed 20.50 bivalents, of which 0–6 were rod bivalents, and a very low frequency of trivalents and quadrivalents. A similar frequency of bivalents and no multivalents were found in this species by Cuñado (1992).

9.5.5.3 Crosses with Other Species of the Wheat Group

The F₁ hybrid between Ae. vavilovii and Ae. tauschii (2n = 28; hybrid-genome D^cX^cS^sD) had at first meiotic metaphase 13.89 univalents, 5.03 bivalents, of which only 1.14 were ring bivalents, and several multivalents (0.75 trivalents, 0.37 quadrivalents and 0.05 pentavalents) (Table 9.6). This pattern of meiotic chromosome pairing confirms that the D^c subgenome of Ae. vavilovii is not completely homologous with the D genome of Ae. tauschii. Evidently, the D^c subgenome is a modified D genome. Likewise, the chromosomal pairing in the F₁ hybrid Ae. vavilovii x Ae. ventricosa (2n = 5x = 35; hybrid-genome D^cX^cS^sDN) and in the reciprocal hybrid, had 4.81 bivalents in one combination and 5.67 bivalents in another, of which only very few were ring bivalents, and several multivalents indicating again that D^cD subgenomes of the two species are not completely homologous (Table 9.7). A somewhat higher number of bivalents was observed in the F₁ hybrid Ae. vavilovii x Ae. cylindrica (2n = 5x = 35; hybrid-genome D^cX^cS^sCD), showing that the D^c subgenome of Ae. vavilovii has better pairing with the D subgenome of Ae. cylindrica than with D subgenome of Ae. ventricosa and the D genome of Ae. tauschii. On the other hand, the F₁ hybrid hexaploid Ae. crassa x Ae. vavilovii (2n = 6x = 42; hybrid-genome D^cD^cX^cX^cDS^s) had 11 bivalents and several multivalents (2.63 trivalents and 0.43 quadrivalents, indicating good homology between the D^cX^C subgenomes of the two species (Table 9.7). This type of chromosomal pairing justifies the conclusion that tetraploid Ae. crassa was one of the parents of Ae. vavilovii. The good homology between the D^cX^c subgenomes of the two species suggests that the formation of Ae. vavilovii from 4x Ae. crassa was relatively recent.

The subgenomes of Ae. vavilovii and those of tetraploid wheat, T. turgidum ssp. turgidum and ssp. durum (2n = 5x = 35; hybrid-genome D^cX^cS^sBA), have almost no chromosomal pairing in the F₁ hybrid between these two species (Chapman and Miller 1978; Melnyk and McGinnis 1962). Similar low chromosomal pairing was observed in the F₁ hybrid between Ae. vavilovii and T. aestivum (2n = 6x = 42; hybrid-genome D^cX^cS^sBAD) (Chapman and Miller 1978). In spite of the presence of the D^c subgenome of Ae. vavilovii and the D of T. aestivum, this low pairing is due to the action of the Ph1 gene of T. aestivum that suppresses pairing of homoeologous chromosomes.

9.5.6 Aegilops juvenalis (Thell.) Eig

9.5.6.1 Morphological and Geographical Notes

Aegilops juvenalis (Thell.) Eig [Syn.: Triticum juvenalis Thell.; Ae. turcomanica (Roshev.) Roshe.: Aegilonearum juvenale (Thell.) Á. Löve] is an annual robust plant, 15–40-cm tall (excluding spikes), jointed in the lower parts and then upright, and nearly or fully glabrous. Its leaves are broad. The spike is medium-sized, 3–7-cm-long (excluding awns), one rowed, cylindrical to slightly moniliform, becomes narrower toward the tip, is hairy, and has 3–7 (usually 5) spikelets. There are 1–3 and seldom no rudimentary spikelets. Spikelets disarticulate into individual spikelets, each with its adjacent rachis segment (barrel-type dispersal unit). Spikelets are elliptical, weakly inflated in the lower parts, and somewhat incised above. Glumes are hairy and overlapping in the upper parts. The tips of the glumes of lateral spikelets have 1–4 small, narrow and flat awns, separated from each other by an interval. Glumes of the terminal spikelet have a flat awn and two flanking teeth, or with 2 or 3 awns. The lemma is leathery, hairy at the top and about one-third longer than the glume, with 1 flat awn, 2–5-mm-long in the lower spikelets but up to 2–4 mm and two lateral teeth in the upper spikelet, or seldom with 3 awns with 2 lateral teeth, one of which may develop into a short awn. Lemma awns are more developed than glume awns. The caryopsis adheres to the lemma and palea (Fig. 9.2f).

The plant shows limited morphological variation mainly in length of spike, number of spikelets, glume and lemma. Thellung, who first described this species, thought it was a hybrid between Ae. crassa and Ae. triuncialis. It resembles its one parent Ae. crassa in many features, but differs from it by its many flat awns on the glumes and lemmas and by shorter and wider glumes, all of which are characteristics of its second parent, Ae. umbellulata.

It is a western and central Asiatic species occurring at rather dispersed locations in Turkmenistan, Uzbekistan, Iran, Iraq, north-east Syria and eastern Turkey. It grows on grey calcareous and alluvial soils, stony ground, gravel and open steppical habitats, edges of cultivation and roadsides. Ae. juvenalis is a weed of cultivation. It shows sporadic distribution, mainly in secondary disturbed habitats, throughout the warm steppes of central Asia. It is adventive in south France (Port Juvenal-from where it receives its name) and grows at altitudes of 150–1000 m. The plant is drought-tolerant in areas with 250–350 mm annual rainfall.

Ae. juvenalis has a medium-sized distribution in the eastern part of the distribution of the genus and is a steppical (Irano-Turanian) element. The two progenitors have contact in Iran, Northern Iraq, and eastern Turkey, where this species presumably originated. Ae. juvenalis is sympatric with the following species: Ae. columnaris, Ae. triuncialis, Ae. tauschii, Ae. cylindrica and Ae. crassa and allopatric with Ae. speltoides, Ae. caudata, T. monococcum subsp. aegilopoides, T. timopheevii subsp. armeniacum, Ae. umbellulata, Ae. geniculata, Ae. neglecta and Ae. biuncialis.

9.5.6.2 Cytology, Cytogenetics, and Evolution

Ae. juvenalis is an allohexaploid species (2n = 6x = 42; genome D^cD^cX^cX^cUU), (Dvorak 1998), originating from hybridization of tetraploid Ae. crassa (2n = 4x = 28; genome D^cD^cX^cX^c), as female, and Ae. umbellulata (2n = 2x = 14; genome UU) (Dubcovsky and Dvorak 1995a; Wang et al. 1997). Morphological analysis of Ae. juvenalis led to its classification into section Vertebrata. Some of the morphological characteristics that differentiate between it and tetraploid Ae. crassa are features derived from the U genome of Ae. umbellulata (Kihara et al. 1959), indicating that the U genome is the third subgenome of Ae. juvenalis, in addition to the two subgenomes of tetraploid Ae. crassa (Kihara et al. 1959).

Dubcovsky and Dvorak (1995a), using restriction fragments of nuclear repeated nucleotide sequences, found that Ae. juvenalis contains all restriction fragments of tetraploid Ae. crassa (except one) and six of the seven marker bands of Ae. umbellulata. All Ae. juvenalis bands of nuclear repeated nucleotide sequences were shared with either tetraploid Ae. crassa or Ae. umbellulata. However, the U subgenome of Ae. juvenalis has somewhat diverged from that of Ae. umbellulata. Divergence of a similar magnitude may have also occurred between Ae. juvenalis and tetraploid Ae. crassa. Differentiation of the U subgenome of Ae. juvenalis from the genome of Ae. umbellulata was also suggested by Kimber and Yen (1989) from their investigation of chromosome pairing in interspecific hybrids.

Ae. juvenalis was studied by C-banding and FISH using clones pTa71 (18S-5.8S-26S rDNA), pTa794 (5S rDNA), and pAs1 (non-coding repetitive DNA sequence) as probes (Badaeva et al. 2002). Their data confirm previous conclusions from genome analysis (Kihara 1957; Kihara et al. 1959) that tetraploid Ae. crassa and Ae. umbellulata are the parental species of Ae. juvenalis. Previously, the genomic constitution of Ae. juvenalis was DDM^jM^jC^uC^u (Kihara 1957; Kihara et al. 1959). The D subgenome is a modified D genome, designated D^c (Dvorak 1998), the C^u subgenome is currently designated U (Kimber and Tsunewaki 1988), and the M^j subgenome is not a modified M genome but rather close to the S genome of section Sitopsis and tentatively designated X^c (McGinnis and Melnyk 1956; Dubcovsky and Dvorak 1995a; Dvorak 1998). The presence of the D^c and U subgenomes was confirmed by genome analysis (McGinnis 1956; McGinnis and Melnyk 1962). The D^c subgenome of Ae. crassa, Ae. vavilovii and Ae. juvenalis are very similar since those of the latter two species derived from Ae. crassa. Indeed, studies of meiotic chromosome pairing showed that the D subgenome in Ae. juvenalis is substantially modified from the D genome of Ae. tauschii (Kimber and Zhao 1983; Zhao and Kimber 1984). Likewise, Bordbar et al. (2011), using allelic diversity at 25 nuclear microsatellite loci, nuclear rDNA ITS, and chloroplast trnL-F sequences, found a close phylogenetic relationship between the D^c subgenomes of Ae. crassa, Ae. vavilovii and Ae. juvenalis. In these three species, cloned sequences revealed high diversity at the nuclear rDNA ITS region (Bordbar et al. 2011). Using inter-simple sequence repeat (ISSR), Gong et al. (2006) also found that the D^c subgenome of Ae. juvenalis is a modified D genome of Ae. tauschii. McGinnis (1956) analyzed meiotic chromosome pairing in a number of interspecific and intergeneric F₁ hybrids involving species of Triticum and Aegilops, in order to determine the subgenomes present in Ae. juvenalis. It was concluded that Ae. juvenalis has the D genome of Ae. squarrosa, which is somewhat altered from the D of T. aestivum, and the U genome of Ae. umbellulata. The third genome was not determined. McGinnis and Melnyk (1962) presented evidence that the third subgenome of Ae. juvenalis is close to the S genome of section Sitopsis and not close to the M genome, as suggested by Kihara et al. (1959). Jaaska (1981), studying the electrophoretic genotype of the enzyme aspartate aminotransferase alcohol dehydrogenase in several D-genome species, concluded that Ae. tauschii subsp. strangulata contributed the D genome to hexaploid wheat, to tetraploids Ae. crassa, and Ae. ventricosa, and to hexaploid Ae. juvenalis. Its organellar genome is D², similar to that of its female parent (Ogihara and Tsunewaki 1988; Wang et al. 1997).

Kihara (1963) wrote that Tanaka (unpublished data) synthesized an allohexaploid from hybridization of tetraploid Ae. crassa x Ae. umbellulata and found that the synthetic hexaploid morphologically resembles Ae. juvenalis. Tanaka crossed the synthetic allohexaploid with Ae. juvenalis and chromosome pairing and fertility in the F₁ hybrid were as follows: the synthetic allohexaploid (2n = 6x = 42; genome D^cD^cX^cX^cUU) had 2.40 univalents, 17.00 bivalents, 0.70 trivalents, 0.90 quadrivalents and 0.10 pentavalents, and a 28.9% seed set (by selfing). The F₁ synthetic x natural Ae. juvenalis (2n = 6x = 42; genome D^cD^cX^cX^cUU) had 4.50 univalents, 14.30 bivalents, 0.50 trivalents, 1.00 quadrivalents and 0.40 pentavalents, and a 15.9% seed set. These data indicate that the synthesized allohexaploid and natural Ae. juvenalis are closely related, although not identical (Kihara 1963).

Parts of the distribution area of Ae. juvenalis in Iran and Iraq overlap with those of its two-parental species. This prompted Kihara et al. (1959) to suggest that these areas may be the place of origin of Ae. juvenalis. Its distribution overlaps that of its tetraploid female parent Ae. crassa, but is much more restricted.

The karyotypic study of Chennaveeraiah (1960) supported the reported existence of the D^c and U subgenomes in Ae. juvenalis. The karyotype of Ae. juvenalis consists of two types, the subgenomes D^c and X^c that derived from diploid species with symmetric karyotype, whereas the subgenome U derived from Ae. umbellulata is asymmetric. Chennaveeraiah (1960) observed three chromosome pairs with satellites, the pairs are of dissimilar sizes. In one satellited pair, there was an interstitial minute chromosome segment between the satellite and the short arm, probably caused by a supernumerary constriction. Others included one pair with extreme subterminal centromeres, four with submedian-subterminal centromeres, one with a median centromere, and the remaining with submedian centromeres. The chromosomes corresponding to the third subgenome did not have satellites and all the pairs had submedian centromeres (Chennaveeraiah 1960).

A comparative analysis of somatic metaphase chromosomes by phase contrast, C-banding and Ag-staining was performed to analyze the activity of the NORs (Cermeño et al. (1984b). It was found that the U subgenome completely suppresses the NOR activity of the D subgenome of Ae. juvenalis and that of one pair of the nucleolar organizer chromosomes of the X^c subgenome of Ae. juvenalis. This corresponds to the observation of only three pairs of satellited chromosomes in Ae. juvenalis Chennaveeraiah 1960) and to the fact that two active NORs belong to subgenome U and the third to subgenome X^c (Badaeva et al. 2002).

Ae. juvenalis had 21 bivalents at meiosis, of which 3–4 are rod bivalents (Chennaveeraiah 1960). In few cells, however, a quadrivalent was formed, indicating the presence of a reciprocal translocation (Chennaveeraiah 1960). C-banding analysis of chromosome pairing at first meiotic metaphase was studied in diploid and polyploid Aegilops species (Cuñado 1992). Most of the polyploid Aegilops species showed a diploid-like meiotic behavior, although multivalents involving homoeologous associations were occasionally observed in Ae. biuncialis, Ae. juvenalis and Ae. crassa(6x); therefore, the Aegilops diploidizing genetic system is not equally effective in all polyploid species (Cuñado 1992).

Badaeva et al. (2021b) recognized two karyotypic groups in Ae. juvenalis: juv-I and juv-II. All genomes of juv-I were significantly modified, whereas juv-II was karyotypically similar to 4x Ae. crassa and Ae. umbellulata. Probably, juv-II originated independently of juv-I, from more recent hybridization of the same parental species.

9.5.6.3 Crosses with Other Species of the Wheat Group

Chromosomal pairing in F₁ hybrids between Ae. juvenalis and its two diploid parents, Ae. tauschii and Ae. umbellulata confirm the suggestion made based on morphological characteristics, that Ae. juvenalis contains subgenomes that derived from these two diploids (Table 9.6). The F₁ hybrid Ae. juvenalis x Ae. tauschii had 6.55 bivalents, 0.92 trivalents and 0.10 quadrivalents (McGinnis and Melnyk 1962) or 5.11 bivalents, 1.07 trivalents and 0.26 quadrivalents (Kimber and Zhao 1983). The data from both hybrids show that Ae. juvenalis contains one subgenome that is closely related, although not completely homologous, to that of Ae. tauschii. This subgenome is a modified D genome and therefore, was designated D^c. On the other hand, the F₁ hybrid Ae. juvenalis x Ae. umbellulata had 8.05 bivalents, 1.42 trivalents, 0.28 quadrivalents and 0.03 hexavalents (McGinnis and Melnyk 1962), indicating that subgenome U of Ae. juvenalis underwent relatively few changes at the hexaploid level and still is homologous to that of its diploid parent.

Chromosomal pairing in F₁ hybrids between Ae. juvenalis and tetraploid species having the D genome, namely, Ae. ventricosa (genome DDNN) and Ae. cylindrica (genome DDCC), indicated that the D^c subgenome of Ae. juvenalis is not so close to the D subgenome of the two tetraploid species. The hybrid Ae. juvenalis x Ae. ventricosa) had 4.86 bivalents, 1.25 trivalents, 0.25 quadrivalents and 0.04 pentavalents (Kimber and Zhao 1983; Table 9.7) and the hybrid Ae. juvenalis x Ae. cylindrica had 6.57 bivalents, 2.03 trivalents, 0.77 quadrivalents, 0.17 pentavalents and 0.14 hexavalents (Kimber and Zhao 1983). The higher pairing in the latter hybrid results from pairing between the U subgenome of the hexaploid and the C subgenome of the tetraploid.

Chromosome pairing in F₁ hybrids between Ae. juvenalis and hexaploid species having the D subgenome, namely, Ae. crassa 6x (genome D^cD^cX^cX^cDD) and Triticum aestivum (genome BBAADD) show that the D^c subgenome of Ae. juvenalis is somewhat diverged from the D subgenome of either hexaploid Ae. crassa or T. aestivum (Table 9.7). The hybrid 6x Ae. crassa x Ae. juvenalis had 8.90 bivalents, 2.01 trivalents, 0.59 quadrivalents and 0.28 higher multivalents (McGinnis and Melnyk 1962) and the hybrid T. aestivum x Ae. juvenalis had 4.33 bivalents, 0.48 trivalents and 0.32 quadrivalents (Riley 1966a). The higher pairing in the former hybrid results from the presence of two homologous subgenomes (D^cD^cX^cX^c) in the hybrid.

Hybrids between Ae. juvenalis and several other Aegilops diploid species, namely, Ae. speltoides, Ae. sharonensis, Ae. longissima, Ae. caudata and Ae. uniaristata, showed reduced pairing (McGinnis and Melnyk 1962), indicating that all three subgenomes of Ae. juvenalis are homoeologous to the genomes of the diploid species. Similarly, hybrids between Ae. juvenalis and tetraploid species of section Aegilops having the U subgenome, namely, Ae. geniculata, Ae. columnaris and Ae. triuncialis, showed that the U subgenome of Ae. juvenalis is closely related to that of the tetraploid species, while the other subgenomes are homoeologous (McGinnis and Melnyk 1962; Kimber et al.1988).

Hybrids between Ae. juvenalis and Amblyopyrum muticum (2n = 2x = 14; genome TT) had 6.46 bivalents, 1.30 trivalents, 0.60 quadrivalents and 0.02 pentavalents (McGinnis and Melnyk 1962). The relatively higher pairing in this hybrid results from the activity of homoeologous pairing promoters that exists in A. muticum (Dover and Riley 1972).

9.6 Section Cylindropyrum (Jaub. & Spach) Zhuk

9.6.1 General Description

Section Cylindropyrum (Jaub. & Spach) Zhuk. (Syn.: Monoleptathera Eig) consists of annual and predominantly autogamous two species. The plants are slender with upright, 20–40-cm-high culms (excluding spikes), with narrow and cylindrical spikes, more than 10–20 times as long as wide but sometimes somewhat shorter. At maturity, the whole spike falls entire or disarticulates into individual spikelets, each falling with its segment beside it (a barrel-type dispersal unit). All spikelets have a similar shape but become smaller towards the tip of the spike. There are 4–10 spikelets per spike, all equal in length to the adjacent rachis segment. There are 1–2, and seldom no rudimentary spikelets. Each spikelet contains 3–5 florets, the upper 1–2 being sterile. Glumes of apical spikelets are with one or three awns, that are at least 3-cm-long, broad, with or without small lateral teeth at the base. Glumes of lateral spikelets may be awned with short awns or awnless. The caryopsis adheres to lemmas and palea.

Two monographs on the genus Aegilops that were published by Zhukovsky (1928) and Eig (1929a), morphologically classified the Aegilops species into several sections. Zhukovsky (1928) included only one species in section Cylindropyrum, Ae. cylindrica Host and put Ae. caudata in section Comopyrum together with Ae. comosa, Ae. heldreichii and Ae. uniaristata. Likewise, Eig (1929a) included only Ae. cylindrica in section Monoleptathera (=Cylindropyrum) and put Ae. caudata in section Macrathera together with Ae. comosa, Ae. heldreichii and Ae uniaristata. Yet, Kihara (1954), classifying the Aegilops species on the basis of genome analysis, noting that the allotetraploid Ae. cylindrica contains a subgenome that derived from Ae. caudata, included Ae. caudata in section Cylindropyrum, together with Ae. cylindrica. This classification was also adopted, on a morphological basis, by van Slageren (1994).

In light of the above, section Cylindropyrum contains two species—one diploid, Ae. caudata L. (2n = 2x = 14; genome CC) and one allotetraploid, Ae. cylindrica Host (2n = 4x = 28; genome DDCC). The Ae. tauschii contributed the D genome to the tetraploid, which was responsible for the barrel-shape disarticulation of many types in this species.

Species of this section distribute in the center of the distribution area of the genus, namely, Turkey, northern Iraq, western Iran, Syria and Lebanon, (Israel (rare) and the diploid species spread from this center to Cyprus and westwards into the Aegean Islands, Greece (incl. Crete), Albania and Bulgaria, and eastwards to Afghanistan. The tetraploid species spread westwards [Greece, Albania Bulgaria, Italy (incl. Sicily)], northwards (Macedonia, Serbia, Croatia, Hungary, Ukraine, Crimea, Georgia, Armenia, and Azerbaijan) and eastwards (Pakistan, Afghanistan, Kyrgyzstan, Tajikistan, Uzbekistan and Turkmenistan). The spread of the tetraploid species northwards and eastwards is presumably the influence of its D genome.

The diploid species grows mainly on the red Mediterranean terra rossa soil, while the tetraploid species grows on a variety of soils from terra rossa to grey-calcareous steppe soil as well as on stony slopes. Both species grow at the edge of and openings in deciduous and sclerophyllous oak forests and maquis, and open herbaceous park formations, and in secondary disturbed habitats such as abandoned fields, edges of cultivation and roadsides. The tetraploid species also grows in open dwarf shrub steppe-like formations, at altitudes of almost sea level to 1750 m above sea level. The two species are common throughout most of the distribution area of the section.

9.6.2 Aegilops caudata L.

9.6.2.1 Morphological and Geographical Notes

Aegilops caudata L. [Syn.: T. markgrafii Greuter in Greuter & Rechinger; Ae. markgrafii (Greuter) K. Hammer; T. caudatum (L.) Godr. & Gren. in Grenier & Godron; T. dichasians Bowden; Ae. dichasians (Bowden) Humphries; Orrhopygium caudatum (L.) Á. Löve] is annual, predominantly autogamous, and tufted, with many tillers, upright, 20–45-cm-tall culms (excluding spikes) and hairy leaves. The spike is linear, narrow, tapering a little toward the tip, 3–10-cm-long (excluding awns), and disarticulates as entire spike at maturity. There are 3–7 cylindrical spikelets, equal in length to the adjacent rachis segment, each with 3–4 florets, the upper 1–2 being sterile. There are 1–3 (usually 2) rudimentary spikelets at the base of the spike. The glumes are rough, with the upper parts overlapping. Glumes of lower spikelets have two teeth, or a sharp tooth and a short, thin awn, separated from the tooth by an acute angle. Glumes of the terminal spikelet gradually taper into a long (4–12-cm), broad awn, longer than the entire spike, with a small adjacent tooth separated by a gap (no angle). The awns diverge sharply from each other. The lemma is membranous, upper parts thickened, with 2–3 teeth, from which weak, short awns can develop in the terminal spikelet. The caryopsis adheres to the lemma and palea (Fig. 9.3a).

Fig. 9.3

Spikes of diploid Aegilops species of sections Cylindropyrum and Comopyrum; a Ae. caudata L.; b Ae. comosa Sm. in Sibth. & Sm. ssp. eucomosa; c Ae. uniaristata Vis.

Several taxonomists (e.g., Greuter, Bowden) included Ae. caudata in the genus Triticum while others maintained it in Aegilops (e.g., Zhukovsky, Eig, van Slageren). Consequently, the name of Ae. caudata has been a matter of controversy (Scholz and van Slageren 1994). Greuter (1976) included it in Triticum under the name T. markgrafii Greuter. But Hammer (1980) brought it back to Aegilops while endorsing the name Aegilops markgrafii (Greuter) Hammer. Bowden (1959) included the genus Aegilops in Triticum and named it T. dichasians, which later, when Aegilops was separated from Triticum, was changed to Aegilops dichasians (Bowden) Humphries. In spite of these suggestions, the name Ae. caudata has been widely used by taxonomists and geneticists, e.g., Zhukovsky (1928), Eig (1929a), Kihara (1954), Badaeva et al. (1996a, b), Ohta (2000), van Slageren (1994), Tsunewaki (1993), and many others. To maintain the common and widely used name Aegilops caudata L. in its traditional sense, Scholz and van Slageren (1994) proposed neotypification of this species endorsing to maintain the usage of the well-known name Ae. caudata.

Ae. caudata resembles Ae. cylindrica (2n = 4x = 28; genome DDCC) in its spike morphology. The main morphological differences between these two species of section Cylindropyrum are caused by the D genome in Ae. cylindrica. According to Scholz and van Slageren (1994), these differences are: (1) in Ae. caudata, the awns of the apical glumes are equal or usually longer than the entire spike and with a base which is a continuation of the apex of the glume, whereas in Ae. cylindrica, these awns are only one-third to one-half of the spike length; (2) absence of small lateral teeth at the base of the apical glume’s awn in Ae. caudata; (3) the apical lemma awns in Ae. caudata are short and narrowly linear, with one or two lateral teeth at the base, whereas they are well developed, equal in length but usually longer than the apical glume awns in Ae. cylindrica. The presence of well-developed awns on the lateral glumes in some forms of Ae. caudata, namely in var. polyathera, superficially makes this species look similar to Ae. cylindrica.

In some spike characteristics, Ae. caudata resembles Ae. comosa. However, the spike of Ae. caudata is more primitive, less specialized and shows a greater resemblance to the basic spike structure of the group. Symeonidis et al. (1979) studied the esterase and peroxidase patterns in five varieties of Aegilops caudata and Ae. comosa in order to elucidate the phylogenetic relationships within and between the two groups. In spite of considerable isozyme polymorphism, closer relationships in the banding patterns were found between different varieties of a single species than between varieties of the two different species. Esterase and peroxidase patterns of the two Ae. caudata varieties, typica and polyathera, were very similar and prove their close phylogenetic relationship. Overall, the electrophoretic data agree well with morphological and cytological findings (Zhukovsky 1928; Eig 1929a; Chennaveeraiah 1960; Kihara 1954).

Ae. caudata has limited morphological variation, mainly involving spike length, number and size of spikelets, awn length and development on the lateral spikelets. On a morphological basis, Ae. caudata was subdivided by Eig (1929a) into two varieties, var. typica (=var. dichasians; var. margrafii), which has awnless lateral spikelets, and var. polyathera Boiss., which has awned lateral spikelets. Var. typica is found in Greece, western and central Turkey, and neighboring Aegean Islands, while var. polyathera extends over the whole area reaching Syria and North Mesopotamia (Kihara 1954).

Ae. caudata is an East Mediterranean and Western Asiatic element. It has a relatively large distribution in the central and eastern regions of the distribution of the genus. It is a bi-regional species of the Mediterranean and steppical (Irano-Turanian) regions. Ae. caudata grows in Croatia, Serbia, Bosnia-Hercegovina, Macedonia, Albania, South Bulgaria, Greece (incl. Crete, Rhodes, and the Aegean region), Turkey, Cyprus, Lebanon, Syria, N. Iraq, Iran and Afghanistan (rare). Ae. caudata occupies a variety of primary and secondary habitats. It grows on a variety of soils from terra rossa to grey-calcareous steppe soil, as well as on stony slopes at the edge of and openings in sclerophyllous and deciduous oak forests and maquis, open herbaceous park formations, open dwarf shrub steppe-like formations, abandoned fields, edges of cultivation and roadsides. It is common throughout most of its distribution area. It grows from almost sea level to 1850 m, in areas with annual rainfall that range from 399–799 mm. Ae. caudata is adventive in the Genoa region and in Sardinia in Italy. It is also found near Marseille, France, and in Scotland.

Ae. caudata can form dense stands, often together with other Aegilops species. It grows sympatrically with A. muticum, Ae. speltoides, Ae. comosa, Ae. uniaristata, Ae. umbellulata, Ae. tauschii, T. monococcum subsp. aegilopoides, T. urartu, T. timopheevii subsp. armeniacum, T, turgidum subsp. dicoccoides, Ae. geniculata, Ae. neglecta, Ae. recta, Ae. biuncialis, Ae. columnaris, Ae. triuncialis and Ae. cylindrica, and allopatrically with Ae. searsii, Ae. peregrina, Ae. crassa, and Ae. juvenalis. Ae. caudata is involved in the parentage of Ae. cylindrica and Ae. triuncialis (section Aegilops).

Ae. caudata possesses genes that confer resistance to leaf rust, stem rust, and powdery mildew, as well as genes for higher grain protein and lysine content (Baldauf et al. 1992; Gong et al. 2017).

9.6.2.2 Cytology, Cytogenetics and Evolution

Ae. caudata is a diploid (2n = 2x = 14), bearing the C genome (Kihara and Lilienfeld 1932; Kihara 1949; Kimber and Tsunewaki 1988; Dvorak 1998). Its organellar genome was designated C by Tsunewaki (1993, 2009) and Wang et al. (1997). This species has the smallest genome in the genus (1C DNA = 4.84 ± 0.089 pg) (Eilam et al. 2007; Table 9.3). Eilam et al. (2007) determined DNA content in seven different accessions of Ae. caudata collected from different regions of the distribution area of the species and very little, if any, intraspecific variation in DNA content was noticed. The karyotype of Ae. caudata is highly asymmetric. Sorokina (1928), Senyaninova-Korchagina (1932), and Chennaveeraiah (1960) described the karyotype of Ae. caudata. They recognized two pairs with satellites on short arms, one pair with sub-median centromere, one pair with a sub-median-sub-terminal centromere and three pairs with subterminal centromeres. Chennaveeraiah (1960) found that the three pairs with sub-terminal centromeres differ from one another with respect to their long arms, while their short arms were not different. Ae. caudata was considered by Eig (1929a) to be a more advanced species than the Sitopsis species and the question is if the formation of this advance species was accompanied by a decrease in DNA content and in the change to an asymmetric karyotype.

In in situ hybridization experiments using a noncoding repetitive DNA sequence derived from T. aestivum (designated TC22b) on genomes of diploid Aegilops species, Teoh et al. (1983) found that Ae. caudata had much less heterochromatin than other species, as shown by a moderate amount of labeling, as well as the intensity of the hybridization sites amount on some chromosomes. The relatively small amount of heterochromatin is in agreement with the low DNA content in this species. In addition, Teoh et al. (1983) and Badaeva et al. (1996b), using a cloned repetitive DNA that consists of 18S and 25S rRNA genes, found that has two chromosome pairs had nucleolar organizer regions. This finding is in complete correspondence with the number of satellites in this species. Badaeva et al. (1996b) found that the two pairs of satellited chromosomes in Ae. caudata belong to homoeologous groups 1 and 5.

Teoh and Hutchinson (1983), using an improved C-banding technique, described a characteristic C-banding pattern in Ae. caudata, which enables the identification of individual chromosomes. All chromosomes have at least one intensely stained centromeric band and most have variable numbers of interstitial bands, while prominent telomeric bands are absent. Likewise, Badaeva et al. (1996a), studying the C-banding pattern of Ae. caudata, found that C-bands in this species are mainly medium or small in size with predominantly interstitial locations. They, like Teoh and Hutchinson (1983), concluded that Ae. caudata possesses an intermediate amount of C-heterochromatin. In situ hybridization with the highly repetitive DNA sequences pSc119 (from S. cereale) and pAsl (from Ae. tauschii), showed that only a few minor pAs1 sites were observed in Ae. caudata, whereas the pSc119 probe hybridized with telomeres of one or both arms of all chromosomes and several interstitial sites (Badaeva et al. (1996a).

Friebe et al. (1992b) analyzed the C-banding patterns of 19 different accessions of Ae. caudata from Turkey, Greece and west Asia, and established a generalized C-banded karyotype. Chromosome-specific C-bands are present in all C-genome chromosomes, allowing the identification of each of the seven chromosome pairs of Ae. caudata. While only minor variations in the C-banding pattern were observed within the accessions, a large amount of polymorphic variation was found between different accessions (Friebe et al. 1992b). C-banding analysis was also carried out to identify Ae. caudata chromosomes in six chromosome addition lines to common wheat. C-banding patterns of the added Ae. caudata chromosomes were identical to those of the ancestor species, indicating that these chromosomes were not structurally rearranged.

In an attempt to produce addition lines of Ae. caudata in common wheat, one chromosome was found to have been selectively retained in common wheat (Endo and Katayama 1978). The gametocidal chromosome was sub-telocentric and related to homoeologous group 3 (Endo 1990; See Sect. 9.3.3.).

Kihara and Lilienfeld (1935), Kihara (1959) and Upadhya (1966) studied chromosomal pairing in F₁ hybrids between common wheat and Ae. caudata and found somewhat higher pairing than in other hybrids between common wheat and diploid species, e.g., with Ae. longissima or Secale cereale. The number of bivalents in common wheat x Ae. caudata hybrids ranged from 3 to 5, with a mean of 4 bivalents/cell and trivalents varying ranged from 0 to 2 (Kihara and Lilienfeld 1935), 0–5 bivalents (mode 3) and trivalents (Kihara 1959), and the number of paired chromosomes ranged from 2 to 13, with a mean of 5.74 chromosomes per cell; the mean chiasma frequency per cell was found to be 2.98 (Upadhya 1966). Evidently, the Ae. caudata genome partially counteracts the suppression of pairing between homoeologous chromosomes in meiosis of common wheat x diploid hybrids, which is driven by the Ph1 gene of common wheat. The Ae. caudata genome seems to modify the potency of the Ph1 gene, resulting in a lower threshold of homoeologous chromosome pairing inhibition (Upadhya 1966). This altered threshold is not of the same magnitude as that brought about by Ae. speltoides or A. muticum genotypes. Kihara’s (1959) report of the occurrence of higher frequencies of multivalents in the Ae. caudata x common wheat hybrids as compared to in the reciprocal common wheat x Ae. caudata hybrids, indicates that the effect of the caudata genome on the Ph1 gene in the F₁ hybrids is enhanced by the caudata cytoplasm.

Kihara and Lilienfeld (1932) thought that the genomes of Ae. caudata and Ae. umbellulata are related and therefore grouped them in the same genomic group, i.e., the C group, and designated them C and C^u, respectively. The opinion that Ae. triuncialis (2n = 4x = 28; genome CCC^uC^u) is nearly autotetraploid, has been expressed by several authors (e.g., von Berg 1931; Karpechenko and Sorokina 1929; Kihara 1929, 1937). It was based on the fact that the hybrid between Secale cereale and Ae. triuncialis has 3–7 bivalents (or sometimes even 6–7), which should be ascribed to autosyndesis of the triuncialis chromosomes. This high pairing led (Kihara 1937) to assume that Ae. triuncialis was formed when the two genomes of the diploid parents, C and C^u, were not significantly differentiated (Kihara and Lilienfeld 1932).

However, Sears (1948), as well as Kihara (1954) himself, felt the inadequacy of placing the genomes of these two species in one genomic group. The two species are morphologically different and are placed in different sections by taxonomists, e.g., Zhukovsky (1928) and Eig (1929a), and by the cytologists Senyaninova-Korchagina (1932) and Chennaveeraiah (1960), who found that the karyotypes of Ae. caudata and Ae. umbellulata are vastly different. Likewise, Teoh and Hutchinson (1983) and Badaeva et al. (1996a) described differences in the C-banding patterns of Ae. caudata and Ae. umbellulata, thus providing additional evidence supporting separation of these two species to different genomic group. The C and C^u genomes also differ in heterochromatin content (Friebe et al. 1992b), which lies in agreement with the low chromosomal pairing observed in meiosis of F₁ hybrids between these two species (Kimber and Abu-Baker 1981). Consequently, Chennaveeraiah (1960) and Kimber and Abu-Baker (1981) suggested to separate Ae. umbellulata’s genome from the C group and to designate a new genomic symbol, U.

Molnár et al. (2016) described the distribution of GAA and ACG microsatellite repeats on chromosomes of the U, M, S and C genomes of Aegilops, and the use of microsatellite probes to label the chromosomes in suspension by fluorescence in situ hybridization (FISH). Purified chromosome fractions enabled them to investigate the structure and evolution of the Aegilops genomes, by comparing the positions of conserved orthologous set (COS) markers on the purified Aegilops chromosomes with known positions on common wheat A, B and D subgenomes. Such comparisons revealed that the distribution of GAA and ACG hybridization signals differs within the U, M, S and C genomes, namely, significant rearrangements had occurred in the U and C genomes, while the M and S genomes exhibited structure similar to wheat.

At the whole-genome level, the structures of the S genome chromosomes of Ae. speltoides and the M genome chromosomes of Ae. comosa were the most similar to wheat, followed by the U genome of Ae. umbellulata, while the structure of the C genome in Ae. caudata differed considerably (Molnár et al. 2016). These results are in line with the findings of previous phylogenetic studies in which Ae. umbellulata and Ae. caudata formed a closer sub-cluster on the Aegilops-Triticum clade, indicating greater genetic similarity, relative to Ae. comosa and Ae. speltoides (Petersen et al. 2006; Mahelka et al. 2011).

Danilova et al. (2017) used molecular cytogenetic techniques and next-generation sequencing to explore the genome organization of Ae. caudata. Fluorescence in situ hybridization with a set of common wheat cDNAs showed that only two chromosomes of Ae. caudata maintained collinearity with wheat, whereas the remaining were highly rearranged as a result of inversions and inter- and intra-chromosomal translocations. Danilova et al. (2017) used sets of barley and wheat orthologous gene sequences to compare discrete parts of the Ae. caudata genome involved in the rearrangements. Analysis of sequence identity profiles and phylogenic relationships grouped chromosome blocks into two distinct clusters. Chromosome painting revealed the distribution of transposable elements and differentiated chromosome blocks into two groups consistent with the sequence analyses. Danilova et al. (2017) suggested that introgressive hybridization accompanied by gross chromosome rearrangements might have had an impact on karyotype evolution and speciation in Ae. caudata.

Tanaka et al. (1967) discovered the occurrence of intra-specific sterility in Ae. caudata, and Ohta (1992, 1995b, 2000; Ohta and Yasukawa 2015) further studied this phenomenon in crossing experiments. In two series of crossing experiments, Ohta (1992) first made reciprocal crosses between the two varieties, var. typica and var. polyathera, collected from sympatric populations and found that all F₁ hybrids showed normal fertility. Second, he crossed accessions collected from different geographical regions, namely, from the western distribution of the species (Aegean area and Greece) and the eastern distribution areas (eastern Turkey, Syria and northern Iraq) and found that the F₁ hybrids from these crosses were completely sterile. In contrast, F₁ hybrids from crosses between the Aegean accessions and the accession from Greece showed normal fertility, and F₁ hybrids from crosses between the accession from northern Iraq and those from Syria and northern Turkey showed high fertility. Accordingly, Ohta (1992) concluded that the intra-specific hybrid sterility in Ae. caudata did not correlate with morphological differences, i.e., it is not related to the varietal subdivision of the species, but rather, to geographical differentiation.

A similar conclusion was reached by Ohta (2000), who further studied the intra-specific hybrid sterility in Ae. caudata. Based on his results, the distribution area of Ae. caudata was divided into two geographical regions effectively isolated by the mountainous region lying between West Anatolia and Central Anatolia. The western region includes Greece, the Aegean Islands, and West Anatolia, while the eastern region consists of Central Anatolia, South Anatolia, East Anatolia, Syria, and Northern Iraq.

The results of Ohta (2000) and the findings from recent palaeopalynological works suggest that during the maximum glacial period from 18,000 BP to 16,000 BP, Ae. caudata occurred in the two isolated regions, i.e., the region surrounding the Aegean Sea and the western Levant or some sheltered habitats in the East Taurus/Zagros mountains arc. It then migrated into Central and East Anatolia from the latter regions, as the climate became warmer (Ohta 2000). Furthermore, it is also suggested that the Levant populations now occur in the eastern region of the distribution, while those occurring in the Aegean Sea region during the last glacial period now occupy the western region of the distribution (Ohta 2000).

Further elucidation of the geographical differentiation pattern of Ae. caudata, was reported by Ohta and Yasukawa (2015). They crossed 35 accessions derived from the entire distribution area, with four tester lines. It became clear that the present distribution area of Ae. caudata can be divided into the western and eastern regions, with the border in the mountains lying between West Anatolia and Central Anatolia: the western and eastern accessions are isolated not only geographically but also reproductively by genetic hybrid sterility.

Ohta (1995b), studied chromosome pairing and segregation at meiosis as well as fertility in sterile F₁ hybrids, a tetraploid derivative induced from one of the sterile hybrids, and their parental lines. The F₁ hybrids showed normal configurations and frequency of chromosome pairing at first meiotic metaphase, but was completely sterile. At first anaphase, chromosomes consisting of two sister chromatids of different lengths were observed. The induced tetraploid was shown to be an autotetraploid based on the configuration and frequency of chromosome pairing at first metaphase, and it showed partial restoration of fertility. Ohta (1995b) suggested that the intraspecific hybrid sterility observed in Ae. caudata is caused by chromosomal cryptic structural hybridity. The differences in chromosomal structure between the parental lines are presumably not large enough to cause preferential pairing in the induced autotetraploid. However, Ae. caudata represents intraspecific divergence that might be an initial step towards speciation due cryptic chromosomal rearrangements (Ohta 1995b).

9.6.2.3 Crosses with Other Species of the Wheat Group

Chromosomal pairing at first meiotic metaphase of Ae. caudata exhibited 7 bivalents, of which 2.87 were rod and 4.13 ring (Cuñado 1992). The relatively high number of rod bivalents in this species results from lack of chiasmata in the short arms of the three pairs of sub-telocentric chromosomes (Chennaveeraiah 1960).

Chapman and Riley (1964) reported chromosomal pairing at meiosis of a haploid individual of Ae. caudata. They observed that all the chromosomes were univalents in most cells, but one bivalent was occasionally formed. The low frequency of this bivalent was interpreted to mean that there is a small duplication of genetic material within the genome of Ae. caudata. This information may be of value to those concerned with the interpretation of meiotic chromosome pairing in hybrids involving Ae. caudata, in genome analysis and evolutionary studies.

Analysis of chromosome pairing at first meiotic metaphase of the F₁ triploid hybrid Ae. caudata x Ae. cylindrica showed the presence of 6–7 bivalents and 0–1 trivalent (Kihara 1954 and Table 9.5), indicating that Ae. cylindrica contains a subgenome that derived from Ae. caudata. [The second subgenome of this allotetraploid derived from Ae. tauschii (Kihara 1949)].

Data of meiotic chromosomal pairing in F₁ hybrids between Ae. caudata and other diploid Aegilops species are presented in Table 9.4. Sears (1941b) crossed two different lines of Ae. speltoides var. ligustica with Ae. caudata, one of which (designated speltoides I) is a high-pairing line that induces high homoeologous pairing in interspecific hybrids and the second (designated speltoides II) is an intermediate-pairing line. In the hybrid Ae. caudata x speltoides I, Sears (1941b) found 3.74 bivalents, of which only 0.48 were ring bivalents, and 1.26 trivalents. The reciprocal hybrids had a similar level of pairing. The Ae. caudata x speltoides II hybrid had 2.90 bivalents, of which only 0.12 were ring bivalents, and 0.50 trivalents. A similar level of chromosomal pairing was observed in F₁ hybrids Ae. caudata x Ae. sharonensis, and somewhat lower level of pairing was observed in the Ae. caudata x Ae. longissima hybrid. These data indicate that the genome of Ae. caudata and those of the studied Sitopsis species have diverged considerably from one another and are homoeologous. Hybrids between Ae. caudata x Ae. tauschii had 3.60 bivalents, of which only 0.60 were ring bivalents, 1.06 trivalents and 0.04 quadrivalents (Sears 1941b; Kihara 1949). This low level of chromosomal pairing shows that the genomes of Ae. caudata and Ae. tauschii also diverged considerably from one another. Somewhat higher chromosomal pairing was observed in F₁ hybrids between Ae. caudata and the species of section Comopyrum, Ae. comosa and Ae. uniaristata. The Ae. caudata x Ae. comosa hybrid had 4–6 bivalents with a mode of 5 and 0–1 trivalents (Kihara 1949), and the Ae. caudata x Ae. uniaristata hybrid had 3.94 bivalents, of which 0.44 were ring bivalents, 0.46 trivalents and 0.06 quadrivalents (Sears 1941b) or 2–5 bivalents (Kihara 1949). A similar level of chromosomal pairing was observed in the F₁ hybrid Ae. caudata x Ae. umbellulata, namely, 3.58–3.68 bivalents, of which 0.12–0.28 were ring bivalents, and 0.40–1.12 trivalents (Sears 1941b) or 3–6 bivalents with a mode of 5 bivalents and 0–1 trivalents (Kihara 1949). Evidently, the genomes of Ae. caudata and those of the Comopyrum species and Ae. umbellulata have diverged from each other, but less than that of Ae. caudata and the Sitopsis species.

Chromosome pairing at first meiotic metaphase of F₁ hybrids Ae. caudata x Amblyopyrum muticum, lacking B chromosomes, showed 3.03 univalents, 2.83 rod bivalents, 0.70 ring bivalents, 1.30 trivalents and 7.70 chiasmata/cell (Ohta 1990; Table 8.2). This low pairing, in spite of the presence of homoeologous-pairing promoters in A. muticum (Dover and Riley 1972), indicates little homology between the chromosomes of the two species. Moreover, hybrids with two B chromosomes of A. muticum showed a drastically low frequency of A chromosome pairing (Table 8.2). The F₁ hybrids with or without B chromosomes, were completely sterile (Ohta 1990).

Hybrids between Ae. caudata and Ae. triuncialis had 7 bivalents, mostly ring bivalents (Kihara 1949), indicating that this allotetraploid contains a subgenome that derived from Ae. caudata [the second subgenome of Ae. triuncialis derived from Ae. umbellulata (Kihara 1954)]. The hybrids between other tetraploids of section Aegilops, i.e., Ae. biuncialis and Ae. columnaris, with Ae. caudata, had approximately 7 bivalents (Kihara 1949) as a result of pairing between the chromosomes of Ae. caudata with either those of the U subgenome of the allotetraploids or the second subgenome, M^b or Xⁿ, as well as autosyndetic pairing between the two subgenomes of the allotetraploids.

The hybrid Ae. caudata x diploid wheat, T. monococcum, had 3.72 bivalents, of which only 0.03 were ring bivalents, and 0.59 trivalents (Table 9.8). The hybrid Ae. caudata x tetraploid wheat, T. timopheevii, had 0–5 bivalents (Kihara 1949), and the hybrid Ae. caudata x hexaploid wheat, T. aestivum ssp. aestivum, had 1.82 bivalents and no trivalents (Table 9.10). Evidently, there is very little pairing between Ae. caudata and the wheat species. Interestingly, there is more pairing with diploid wheat than with polyploid wheats, presumably due to the suppressive effect of the Ph1 gene of polyploid wheat.

9.6.3 Aegilops cylindrica Host

9.6.3.1 Morphological and Geographical Notes

Ae. cylindrica Host, jointed goat grass, [Syn.: Triticum cylindricum (Host) Ces., Pass & Gibelli; Triticum caudatum (L.) Godr. & Gren.; Cylindropyrum cylindricum (Host) Á. Löve] is a predominantly autogamous, annual species, tufted with many tillers, with a 20–40(-80)-cm tall (excluding spikes) culm is prostrate near the ground and then upright. Leaves are narrow, linear, smooth or hairy. The spike is 5–8(-12)-cm-long (excluding awns), cylindrical, one-rowed, and more or less awned. There are 6–10(-12) spikelets, that become smaller toward the tip of the spike. The spike disarticulates into individual spikelets, each with the adjacent rachis segment (barrel-type dispersal unit) or falls entire from the culm. The number of rudimentary spikelets is 1–2, seldom zero. Spikelets are cylindrical, three or more times longer than broad, mostly equal in length and appressed to the rachis segment. Glumes have a short triangular awn (up to 3.5-cm-long) and an associated short, broad, blunt tooth forming an obtuse or acute angle with the awn. Awns on glumes of lower spikelets are shorter than awns on glumes of upper spikelets, which are 3–6-cm-long. Lemmas are solid in their upper parts, nerved, 2–3-toothed, always with 3 teeth in the terminal spikelet, with the middle tooth elongated into an awn thicker and longer (4–8-cm-long) than the awns on the glumes. Awns of lemmas of lower spikelets are shorter (or absent) than associated glume awns. Apical glume and lemma awns are always shorter than the length of the spike. The caryopsis adheres to the lemmas and palea (Fig. 9.2g).

Ae. cylindrica exhibits very wide variation in the length of culms, length of spike, number, size, hairiness and color of spikelets, and in development of the lateral glume awns. There are more awns on the glumes but the lemma awns are always more developed. This wide morphological variation led Zhukovsky (1928), Eig (1929a) and Hammer 1980) to subdivide the species into several intraspecific categories. Hammer (1980) subdivided Ae. cylindrica into four varieties: var. cylindrica, var. aristulata (Zhuk.) Tzvel., var. pauciaristata Eig, and var. prokhanovii Tzvel. Var. cylindrica was further divided by Hammer (1980) into three forma: f. cylindrica, f. ferruginea (Popova) Hammer, and f. brunnea (Popova) Hammer. Var. prokhanovii was also further divided to three formas: f. prokhanovii (Tsvel.) Hammer, f. rubiginosa (Popova) Hammer, and f. fuliginosa (Popova) Hammer. Chennaveeraiah (1960), based on karyotypic studies, elevated the varieties to subspecies and recognized only two subspecies in Ae. cylindrica, subsp. cylindrica and subsp. pauciaristata (Eig) Chennav.

Genetic variation of Ae. cylindrica was assessed on various levels and using different methodological approaches, morphological studies, isozyme analyses, storage protein comparisons and molecular markers. Using quantitative and qualitative morphological traits, Arabbeigi et al. (2015) evaluated genetic variation of 66 Aegilops cylindrica genotypes, collected from western and northwestern Iran. They noted high genetic variation in the quantitative and qualitative traits of Iranian Ae. cylindrica, with lemma length and glume color showing the highest variation. Cluster analysis divided the studied genotypes into three groups, each occupying a different region of Iran.

Isozymes analyses showed little variation between accessions of Ae. cylindrica collected from various locations in the distribution area of the species. Thus, Nakai (1981), studying isoelectric focusing patterns of esterase isozymes, Watanabe et al. (1994), studying alpha-amylase isozymes, and Hedge et al. (2002), studying 10 isozymes, revealed high uniformity in the isozyme patterns of the studied accessions. Likewise, analysis of glutenin subunits in different accessions of Ae. cylindrica did not disclose variation in the high-molecular weight glutenin subunits (Farkhari et al. 2007; Khabiri et al. 2012), whereas some variation was noticed in the low-molecular weight (LMW) glutenin subunits (Khabiri et al. 2012). While glutenin subunits exhibit low diversity, gliadin subunits are much more polymorphic in Ae. cylindrica. Khabiri et al. (2013), using polyacrylamide gel electrophoresis, assessed genetic diversity in banding patterns of gliadin protein in seventeen populations of Aegilops cylindrica from northwestern Iran. Their results showed that most bands were related to the ω type of gliadins, whereas the smallest number of bands pertained to the β type gliadins. Genetic diversity between populations was greater than within populations. Assessment of total variation for the three gliadin types indicated that the highest total variation was related to the β type, while the lowest belonged to the ω type. Cluster analysis using the complete linkage method divided populations into two separate groups, in which genetic diversity does not follow geographical distribution.

Molecular markers such as random amplified polymorphic DNA (RAPD) assays (Okuno et al. 1998; Pester et al. 2003; Goryunova et al. 2004; Farkhari et al. 2007), and amplified fragment length polymorphism (AFLP) (Pester et al. 2003) revealed a certain degree of variation. RAPD analysis of Ae. cylindrica accessions collected in Central Asia and North Caucasia showed that while accessions of this species from north Caucasia were genetically uniform, those from Central Asia were slightly more diverse (Okuno et al. 1998). Yet the authors did not find associations between altitudinal variation and variability of RAPD markers. Pester et al. (2003) used RAPD and AFLP to study genetic variation in Ae. cylindrica and found that, although AFLP produced more scorable bands as compared to RAPD, both methods revealed limited genetic diversity in Ae. cylindrica. Similarly, Goryunova et al. (2004), using RAPD, found that the intraspecific level of genetic variation in Ae. cylindrica was considerably lower than in either parent, Ae. tauschii or Ae. caudata. Yet, in contrast to the above findings, Farkhari et al. (2007), using RAPD, evaluated genetic variation of 28 populations of Ae. cylindrica, collected from different parts of Iran, and found that primers of RAPD generated 133 reproducible fragments, 69% of which were polymorphic. There was little relationship between genetic divergence and geographical origin (Farkhari et al. 2007).

The results of Farkhari et al. (2007) showed that RAPD is suitable for genetic diversity assessment in Ae. cylindrica populations. This is expected, as protein markers reflect only variation in the coding parts of the genome, which is, by nature, more conservative and thus less polymorphic. RAPDs, on the other hand, can detect variation in both coding and non-coding sequences, and the length of the primers allows the amplification of a large number of fragments with a single primer (Guadagnuolo et al., 2001). This technique has the advantage of requiring very small quantities of template DNA and no prior sequencing knowledge of the target genome (Pester et al. 2003).

Simple sequence repeats (SSRs) are another type of molecular marker used to study intraspecific genetic diversity. Naghavi et al. (2008. 2009b), using SSRs to assess genetic diversity in Ae. cylindrica accession, collected from 13 different sites in Iran, found that intraspecific genetic diversity in this species is not as high as it was in Ae. tauschii and Ae. crassa. Moreover, SSR markers showed that the genetic distance between Ae. tauschii and Ae. cylindrica is very small. Mohammadi et al. (2014), using 17 inter-simple sequence repeat (ISSR) primers, investigated genetic diversity among 35 accessions of Ae. cylindrica, collected from different Iranian locations. Out of 190 alleles that were amplified, 188 (98.95%) were polymorphic. Cluster and principal coordinate analysis (PCA) showed no association between molecular diversity and geographic diversity of genotypes, indicating that there is high genetic diversity within populations (Mohammadi et al. 2014). Based on their findings, Mohammadi et al. (2014) concluded that the center of diversity and origin of Ae. cylindrica might be the western region of Iran and this species migrated from this region to other parts of its distribution area.

Ae. cylindrica is a Mediterranean and steppical (Irano-Turanian) element, which distributes in the north Mediterranean and central Asiatic parts of the distribution of the genus. It grows in the west in Greece (rare), including Crete (rare), Albania, Macedonia, Serbia, Croatia, Hungary, Romania, Bulgaria, south Ukraine, Crimea, Ciscaucasia, Georgia, Armenia, Azerbaijan, southern Turkmenistan, southern Uzbekistan, northern Tajikistan, western Kyrgyzstan, Afghanistan (rare), Iran, northern Iraq (rare), Turkey, Syria, Lebanon (rare), Jordan (rare) and Israel (rare).

Ae. cylindrica inhabits a wide variety of primary and secondary habitats. It grows at the edges and in openings of deciduous Mediterranean and steppical oak forest and maquis, dwarf shrub formations, dwarf shrub steppe-like shrub formations, dry hill and mountain slopes, plains, pastures, abandoned fields, ruderal and disturbed sites, wastelands, edges of cultivation, disturbed habitats and roadsides, railway sides, grasslands, and close by or within cultivation, such as orchards, vineyards, and wheat fields. Near common wheat fields, Ae. cylindrica forms natural hybrids with wheat. It grows mainly on calcareous and basaltic soils, less frequently on sands, in areas with annual rainfall from 450 to 800 mm, indicating a preference for more humid environments than most Aegilops species. Ae. cylindrica is usually found at altitudes from 300 to 2000 m, rarely at lower attitudes (around the Caspian Sea).

The species is common throughout most of its range and locally abundant. Ae. cylindrica, more than most species of the genus Aegilops, shows weedy behavior, occupying large stands after recent disturbances, i.e., it is a successful colonizing species (van Slageren 1994). It grows in mixed populations with many Aegilops species, with which it may introgress. Its two diploid parents overlap in east Turkey, and Iran, where Ae. cylindrica may have originated and then spread both westwards and northwards. Its distribution area is larger than those of its diploid parents. It grows sympatrically with Ae. caudata, Ae. tauschii, Ae. umbellulata, Ae. geniculata, Ae. neglecta, Ae. biuncialis, Ae. columnaris, Ae. triuncialis, Ae. ventricosa, Ae. crassa, Ae. juvenalis, Amblyopyrum muticum, wild Triticum monococcum ssp. aegilopoides, T. urartu, and wild T. timopheevii, and allopatrically with Ae. comosa, Ae. uniaristata, Ae. speltoides, Ae. peregrina, Ae. vavilovii and wild T. turgidum.

Ae. cylindrica is an adventive in north Italy, France, Germany, Switzerland, and in many other countries of central, northwestern, northern and eastern Europe. It was first introduced to Kansas, USA, in the late nineteenth century, as a contaminant in Turkey winter wheat seed, and since then, it was introduced again and again and spread to many different additional states, mainly to western and northwestern states where it has become very common (Hitchcock 1951; Barkley 1986; Donald and Ogg 1991). Moreover, changing wheat production practices during the second half of the twentieth century have encouraged the spread and increase of this weed (Donald and Ogg 1991).

No variation was revealed when isozyme studies were carried out on Ae. cylindrica introduced to North America and Eurasian accessions (Watanabe et al. 1994; Hedge et al. 2002). Likewise, Pester et al. (2003), using two DNA molecular marker techniques, RAPD and AFLP, found only low variation between North American and Eurasian accessions. Cluster analysis showed small genetic distances between all Ae. cylindrica accessions from North America, with the greatest distance observed between an accession from Washington and a group of three other U.S. accessions from Nebraska, Oklahoma and Utah. These results suggest either-multiple introductions of Ae. cylindrica into the United States or some genetic divergence within U.S. populations since their introduction. These introductions may have originated from different Eurasian geographic locations and included different Ae. cylindrica genotypes. The Eurasian accessions, which, although somewhat more genetically varied than those from the United States, are still much less diverse than might be expected in populations from the center of origin of the species (Okuno et al. 1998; Watanabe et al. 1994).

Ae. cylindrica has similar growth habits as hexaploid winter wheat and consequently, has developed into a serious pest in winter wheat fields, especially in northwestern states, where it can lower winter wheat yield by competing for growth requirements, thereby reducing harvesting efficiency, and lowering crop quality by contaminating harvested grain (Donald and Ogg 1991). In 1999, it was reported that Ae. cylindrica infested an estimated 2 million hectares in the US alone, and caused annual losses of $145 million in crop yield and quality, and that the infested area increases annually at a rate of about 20,000 hectares (Kennedy and Stubbs 2007). In addition, it can easily hybridize with winter wheat and form viable hybrids that can produce seeds via backcrossing to either parent (Gandilyan and Jaaska 1980; Guadagnuolo et al. 2001; Hegde and Waines 2004; Galaev and Sivolap 2005; Gandhi et al. 2006). Because Ae. cylindrica contains the D subgenome, which is homologous to the D subgenome of common wheat, it is very difficult to control it in commercial fields. Aside from its spikelets (the dispersal units) that are similar in shape and size to the grains of winter wheat, making it difficult to separate them from wheat using conventional methods of grain cleaning, the shared genetics makes it practically impossible to selectively kill off Ae. cylindrica without harming the winter wheat. This poses problems for farmers who have to suffer through reduced yields and poorer quality winter wheat (Donald and Ogg 1991). However, it is hoped that breeding of winter wheat lines resistant to imidazolinone will allow the use of imazamox to selectively kills Ae. cylindrica. Yet, Seefeldt et al. (1998) mentioned the concerns about the possibility of transferring the resistant genes from common wheat to Ae. cylindrica through hybridization and backcrossing to the wild parent. Indeed, Guadagnuolo et al. (2001) found that introgression of wheat DNA into Ae. cylindrica is possible. Gandhi et al. (2006) obtained similar results when studying the occurrence of wheat nuclear microsatellite markers in BC₁ plants obtained from Ae. cylindrica x common wheat backcrossed to the wild parent.

Ae. cylindrica possesses a large number of desirable genes that can be used for wheat improvement. It contains genes for resistance to various fungal diseases (e.g., Bai et al. 1995; Singh et al. 2004) for cold hardiness and salt tolerance (e.g., Farooq et al. 1992; Arabbeigi et al. 2014; Kiani et al. 2015), and for storage protein genes (Wan et al. 2000).

9.6.3.2 Cytology, Cytogenetics, and Evolution

Ae. cylindrica is an allotetraploid species (2n = 4x = 28; genome DDCC). Its genome constitution was determined by the analyses of chromosome pairing (Kihara 1930; Kihara and Matsumura 1941; Sears 1944b; McFadden and Sears 1946; Kimber and Zhao 1983; Zhao and Kimber 1984), karyotype structure (Chennaveeraiah 1960), storage proteins (Johnson 1967; Masci et al. 1992), isozymes (Jaaska 1981; Nakai 1981), and differences in restriction length patterns of repeated nucleotide sequences (Dubcovsky and Dvorak 1994). All these studies identified the diploid species Ae. caudata as the donor of the C subgenome and Ae. tauschii as the donor of the D subgenome. Hence, Ae. cylindrica originated from spontaneous chromosome doubling of an F₁ hybrid between Ae. tauschii and Ae. caudata (Kihara 1944), the former being the plasmon donor to Ae. cylindrica (Tsunewaki 1996).

Amphiploid Ae. caudata-Ae. tauschii (2n = 4x = 28; genome CCDD) was synthesized from its putative parents (Sears 1941b). The synthetic amphiploid closely morphologically resembled the natural Ae. cylindrica and chromosomal pairing at meiosis of the F₁ hybrid between the synthetic and the natural Ae. cylindrica was almost regular, i.e., 0.54 univalents, 13.14 bivalents, 0.02 trivalents, and 0.28 quadrivalents per cell (Sears 1941b). It thus appears that the synthetic amphiploid has no significant chromosome differences from natural Ae. cylindrica, except possibly a reciprocal translocation. Concerning seed set, the hybrid was less fertile (44%) than the natural Ae. cylindrica (76%), but more fertile than the synthetic amphiploid (27%).

Johnson (1967) showed that Ae. tauschii subsp. tauschii (former subsp. eusquarrosa Eig) var. tauschii (former var. typica Eig) + Ae. caudata var. polyathera had similar seed protein pattern as that of Ae. cylindrica. Based on isozyme studies, Jaaska (1981) also proposed subsp. tauschii of Ae. tauschii as the donor of the D genome to Ae. cylindrical, and subsp. strangulata as the D subgenome donor to the hexaploid wheats. Nakai (1981) found no zymogram variation in studies of the isoelectric focusing patterns of esterase isozymes in 30 strains of Ae. caudata. Consequently, it was impossible to identify the line that contributed the C subgenome to Ae. cylindrica. Since Ae. cylindrica had an isozyme pattern which corresponded to a mixture of esterases from Ae. caudata and one genotype of Ae. tauschii, Nakai (1981) concluded that Ae. cylindrica originated with a single amphiploidy event, and that the C and D subgenomes have remained remarkably constant regarding esterase isozyme composition. Similar to Jaaska (1981), Nakai (1981) also found that the D subgenome of common wheat derived from a genotype of Ae. tauschii different than the one donated to Ae. cylindrica. Masci et al. (1992) confirmed the presence of the alpha-gliadin genes of Ae. caudata in Ae. cylindrica, in addition to those of Ae. tauschii.

Maan (1976) obtained fully fertile plants of normal vigor, with a normal chromosome number, and normal meiotic pairing in alloplasmic lines of common wheat bearing the cytoplasm of Ae. tauschii or Ae. cylindrica. The similar nucleo-cytoplasmic interactions indicate cytoplasmic homology between Ae. tauschii and Ae. cylindrica. Thus, Maan (1976) concluded that the cytoplasm of Ae. cylindrica was contributed by Ae. tauschii, a conclusion that was later verified by Tsunewaki (1989, 1996). Ae. tauschii is the maternal parent of Ae. cylindrica and of two other allotetraploids, Ae. ventricosa and Ae. crassa. The average distances between pairs of the D-genome tetraploids and Ae. tauschii are 0.015 with Ae. cylindrica, 0.050 with Ae. crassa and 0.006 with Ae. ventricosa (Wang et al. 1997). This suggests that the allotetraploids with the D genome originated at three different times from Ae. tauschii as female, in the following order: Ae. crassa, Ae. cylindrica, and Ae. ventricosa (Wang et al. 1997).

Tsunewaki et al. (2014) produced an alloplasmic Ae. cylindrica with the plasmon of Ae. caudata and found that the genetic effect of the Ae. caudata plasmon on the manifestation of Ae. cylindrica genomes was limited to male fertility: the alien plasmon caused pollen and self-pollinated seed sterility, without exerting any detectable effects on the other morphological and physiological characters investigated. Further studies (Tsunewaki et al. 2014) indicated that the male sterility expressed by the alloplasmic line was due to genetic incompatibility between the Ae. cylindrica genome and Ae. caudata plasmon. Previously, Tsunewaki et al. (2002) investigated the genetic effects of the Ae. caudata plasmon on the genome manifestation of 12 genotypes of common wheat and found that the caudata plasmon caused pollen and seed sterility in nine of the 12 genotypes.

Yet, a recent investigation comparing chloroplast and nuclear microsatellite loci of 36 Ae. cylindrica accessions with those of seven accessions of Ae. caudata and 17 accessions of Ae. tauschii, revealed that one of the examined Ae. cylindrica accessions possessed a plastom of the C type, the caudata plastom, whereas all others had the D type, the tauschii plastom (Gandhi et al. 2009). The accession with the C cytoplasm was originally collected in Turkey. Their result suggests that Ae. cylindrica arose from multiple hybridizations between Ae. caudata and Ae. tauschii, presumably along the Fertile Crescent, where the geographic distributions of its diploid progenitors overlap. In most cases, Ae. cylindrica originated from a cross between Ae. tauschii as female and Ae. caudata as male, as proposed by Kihara (1944) and Tsunewaki (1996). However, their result also suggests a diphyletic origin of Ae. cylindrica from the reciprocal crosses between Ae. tauschii and Ae. caudata, although its diphyletic origin must be confirmed by other means (Tsunewaki 2009). In this case, caudata cytoplasm did not cause male sterility in the allotetraploid.

There is 9.59 pg nuclear 1C DNA in Ae. cylindrica (Eilam et al. 2008). This value deviates by −4.20% from the expected cumulative amount of DNA of the two diploid parents, 4.84 pg 1C DNA of Ae. caudata and 5.17 pg of Ae. tauschii (Eilam et al. 2007). Bakhshi et al. (2010) determined the DNA peak mode of 100 accessions of Ae. cylindrica using flow cytometry and found variations among accessions from different locations. There was no significant difference in morphological traits of accessions with high and low DNA peak mode. Accessions with lower and higher DNA content had shorter and longer chromosome lengths, respectively (Bakhshi et al. 2010). The observed intraspecific differences in DNA size in Ae. cylindrica are predominantly associated with differences in the amounts of repetitive sequences. Changes in copy number of certain DNA sequences, as a response to different environments, may be responsible for the observed changes in DNA size.

The karyotype of Ae. cylindrica is asymmetric. Senyaninova-Korchagina (1932) reported that the karyotype of Ae. cylindrica contains four pairs with satellites on short arms, three pairs with extreme subterminal centromeres, two pairs with submedian-subterminal centromeres, one pair with an almost median centromere, and the rest with submedian centromeres. Pathak (1940) observed one pair with satellites and another pair with secondary constrictions and four nucleoli. In other words, he observed two pairs with satellites. Chennaveeraiah (1960) studied the karyotype in two subspecies of Ae. cylindrica, ssp. cylindrica and ssp. pauciaristata. In neither of them did he observe four satellited pairs. Ae. cylindrica ssp. cylindrica showed a karyotype including three satellite pairs, three pairs with extreme subterminal centromeres, one pair with a submedian-subterminal centromere, and the rest with submedian centromeres. The karyotype of Ae. cylindrica ssp. pauciaristata differs from that of subsp. cylindrica in that it has only one pair with satellites.

Senyaninova-Korchagina (1932), based on her karyotypic studies, thought that one subgenome of Ae. cylindrica corresponds to the karyotype of Ae. caudata, whereas the other subgenome is closely related to the karyotype of Ae. bicornis. She made another wrong decision by depicting four satellited pairs in Ae. cylindrica. In contrast, Chennaveeraiah (1960) observed only three satellited pairs in ssp. cylindrica, two on the C subgenome and one on the D subgenome. According to him, one subgenome of Ae. cylindrica resembles the karyotype of Ae. caudata whereas the other subgenome resembles the karyotype of Ae. tauschii. The single pair of satellited chromosomes in Ae. cylindrica ssp. pauciaristata corresponds to the satellited pair in Ae. tauschii. The second subgenome of this subspecies, the C subgenome, does not have the two satellited pairs. It is possible that the differences between the two subspecies reflect some variation in the karyotype of the parental species, Ae. caudata (Chennaveeraiah 1960).

Teoh et al. (1983) used a repetitive DNA sequence, derived from T. aestivum, coding for ribosomal RNA, as a probe in an in-situ hybridization experiment on genome of Ae. cylindrica. They found only two pairs of rRNA sites, one less than the expected three from the presence of three pairs of satellited chromosomes (Chennaveeraiah 1960).

The genomic constitution of Ae. cylindrica was analyzed by C-banding, genomic in situ hybridization (GISH), and fluorescence in situ hybridization (FISH), using the DNA clones pSc119, pAs1, pTa71 and pTA794 (Linc et al. 1999). The C-banding patterns of the D and C subgenome chromosomes were similar to those of the D and C genome chromosomes of the diploid progenitor species Ae. tauschii and Ae. caudata, respectively. These similarities permitted the identification at first meiotic metaphase of the chromosomes of the D and the C subgenomes of Ae. cylindrica (Cuñado 1992). The chromosomes of the D subgenome were almost without C-banding, just like the chromosomes of Ae. tauschii, whereas the chromosomes of the C subgenome were smaller than those of the D subgenome and showed thin interstitial bands (Cuñado 1992).

FISH analysis detected one major 18S-5.8S-25S rDNA locus in the short arm of chromosome 1C (Linc et al. 1999). Minor 18S-5.8S-25S rDNA loci were mapped in the short arms of 5D and 5C. 5S rDNA loci were identified in the short arm of chromosomes 1C, 5D, 5C and 1D. GISH analysis detected inter-genomic translocation in three of the five Ae. cylindrica accessions studied. The breakpoints in all translocations were non-centromeric, with similar-sized segment exchanges (Linc et al. 1999).

Cermeño et al. (1984b) analyzed the activity of the nucleolar organizer regions (NORs) in somatic metaphase chromosomes of Ae. cylindrica by phase contrast, C-banding and Ag-staining. They reported that the nucleolar activity of the D subgenome is completely suppressed by the C subgenome.

A recent FISH analysis, using repetitive DNA probes (Afa family, pSc119.2, and 18S rDNA) on root tip mitotic metaphase spreads, enabled identification of the entire set of chromosomes in Ae. cylindrica (Molnar et al. 2015). In addition, use of two-color GISH that differentially labeled total genomic DNA from Ae. caudata and Ae. tauschii, discriminated the constituent C and D subgenomes of Ae. cylindrica. Yet, certain differences were found between the C genome of Ae. caudata and that of Ae. cylindrica (Molnar et al. 2015).

Chromosome 2C has a gametocidal effect when added as a monosome to common wheat cv. Chinese Spring (Endo 1996,). Yet the effect of this gametocidal chromosome is relatively mild and there are viable progeny that lack the Ae. cylindrica chromosome. Approximately half of these progeny exhibit structural changes, such as deletions and translocations in various regions of all the wheat chromosomes. Thus, the ability of the gametocidal chromosome 2C to induce deletions of a variety of chromosomal segments observed in the viable progeny of the monosomic addition line that lacks the gametocidal chromosome, has been exploited to induce deletions in various wheat stocks (Endo 2007). The deletions that were stably transmitted to the offspring that became homozygous lines, were identified by C-banding (Endo and Gill 1996). The deletion stocks showed variations in morphological, physiological and biochemical traits, depending on the size of their chromosomal deficiency, and constitute powerful tools for physical mapping of wheat chromosomes (Endo and Gill 1996).

9.6.3.3 Crosses with Other Species of the Wheat Group

Chromosome pairing at first meiotic metaphase of Ae. cylindrica was regular, i.e., 0.09 univalents, 3.61 rod bivalents, 10.30 ring bivalents and 24.21 chiasmata per cell (Cuñado 1992). With the ability to identify the chromosomes of the D and C subgenomes at first meiotic metaphase, Cuñado (1992) determined chromosomal pairing of each subgenome. Average pairing of D subgenome chromosomes was 0.03 univalents, 1.58 rod bivalents, 5.39 ring bivalents and 12.31 chiasmata. That of the C subgenome chromosomes was 0.65 rod bivalents, 6.34 ring bivalents and 13.34 chiasmata. Interestingly, in spite of the occurrence of three pairs of chromosomes with extremely subterminal centromeres in the C subgenome (Chennaveeraiah 1960), there are more ring bivalents in this subgenome than in the D subgenome.

Chromosomal pairing at first meiotic metaphase was analyzed in F₁ hybrids between Ae. cylindrica and each of its two parental diploids. The triploid hybrid Ae. cylindrica x Ae. caudata (genome DCC) had 6–7 bivalents and 0–1 trivalents (Kihara 1954) and the triploid hybrid Ae. cylindrica x Ae. tauschii (genome DCD) had 8–9 bivalents and 0–5 trivalents (Kihara 1949). Evidently, one of the two subgenomes of Ae. cylindrica, subgenome C is homologous to the genome of Ae. caudata while the second, subgenome D, is homologous to that of Ae. tauschii. The genomes of the two diploids are not homologous, as evident from the average chromosomal pairing in the hybrid between them, that had only 3.60 bivalents, of which only 0.60 were ring bivalents, and 1.06 trivalents (Sears 1941b; Table 9.4).

Data of chromosomal pairing in F₁ hybrids between Ae cylindrica and allopolyploids of Section Vertebrata are presented in Table 9.7. The hybrid Ae. cylindrica x tetraploid Ae. crassa exhibited 6.00 bivalents and several multivalents and the reciprocal hybrid had 7.30 bivalents (Kimber and Zhao 1983). The hybrid Ae. ventricosa x Ae. cylindrica also had seven bivalents (Kimber and Zhao 1983). Likewise, the hybrids between Ae. cylindrica and the hexaploid species Ae. juvenalis x Ae. cylindrica, and the hybrid Ae. vavilovii x Ae. cylindrica, and hexaploid Ae. crassa x Ae. cylindrica, had about 7 bivalents at meiosis (McGinnis and Melnyk 1962; Melnyk and McGinnis 1962; Kimber and Zhao 1983). These data indicate that the D subgenomes in all these species are still closely related.

Average chromosome pairing in the hybrid T. turgidum ssp. durum x Ae. cylindrica (genome BADC) was 27.50 univalents and 0.50 bivalents (Sears 1944b), indicating that these two species do not share a common subgenome. In contrast the T. aestivum x Ae. cylindrica hybrid (genome BADDC) had about 7 bivalents (Riley 1966a; Kimber and Zhao 1983), indicating that the third subgenome of hexaploid wheat is homologous to one of the subgenomes of Ae. cylindrica.

9.7 Section Comopyrum (Jaub. & Spach) Zhuk.

9.7.1 General Description

Section Comopyrum (Jaub. & Spach) Zhuk. (Syn.: Macrathera Eig) consists of annual, predominantly autogamous two species. Plants are many-tillered with thin, short (10–30-cm-tall; excluding spikes) culms, with narrow, usually hairy leaves, and narrow, cylindrical or moniliform linear-lanceolate or narrow-elliptical, short (1.5–7.0-cm-long) spikes that gradually taper toward the tip. At maturity, the entire spike falls as one unit. There are 2–3 long narrow to oval spikelets that are shorter than the adjacent rachis segment, with the terminal spikelet sometimes being sterile. There are 1–3 rudimentary spikelets at the base. Spikelets contain 3–4 florets, the upper 1–2 being sterile. Glumes of apical spikelets have either 3 awns (a well-developed central one and 2 more slender, lateral ones which are sometimes reduced to teeth), or only 1 awn. Glumes of lateral spikelets are rough or hairy, with the upper parts overlapping, with two teeth with an angle between; sometimes one of the teeth develops into a small awn. Lemmas are seldom awned in the upper spikelet. The caryopsis adheres to lemmas and palea (Fig. 9.3).

Section Comopyrum contains two diploid species, Ae. comosa Sm. in Sibth. & Sm. (genome M), and Ae. uniaristata Vis. (genome N). The former has a cylindrical and the latter a moniliform spike. Section Comopyrum was first described by Jaubert and Spach in 1850–1851, who included in it only one species, Ae. comosa, that contained one variety, var. subventricosa. Zhukovsky (1928), based on taxonomical criteria, classified Ae. caudata L., Ae. comosa Sm. in Sibth. & Sm., Ae. heldreichii Holzm. and Ae. uniaristata Vis. in section Comopyrum. Eig (1929a) also included Ae. caudata, together with Ae. comosa and Ae. uniaristata, in section Macrathera (currently Comopyrum), but included Ae. heldreichii in Ae. comosa. Based on studies of chromosomal pairing in the interspecific hybrids, Kihara (1937, 1940b, 1954) transferred Ae. caudata from section Comopyrum to section Cylindropyrum. Moreover, Kihara (1937, 1940b) thought that Ae. comosa and Ae. uniaristata are very closely related species and designated their genomes M and M^u, respectively. However, Chennaveeraiah (1960), based on a karyotypic analysis, suggested to designate the genome of Ae. uniaristata N. This suggestion was accepted by Kimber and Sears (1987), Kimber and Feldman (1987), Kimber and Tsunewaki (1988), and Dvorak (1998). Wang et al. (2000) claimed that the classification of section Comopyrum should be reconsidered based on their study of internal transcribed spacers (ITS) of nuclear ribosomal DNA sequences. Badaeva et al. (1996b) also postulated that Ae. uniaristata should be removed from section Comopyrum based on in situ hybridization and chromosome morphology data. However, Yamane and Kawahara (2005) reported that the unrooted strict consensus tree based on BPS + indels + microsatellites showed that Ae. uniaristata and Ae. comosa are clustered together. This finding is in agreement with previous molecular studies, based on chloroplast DNA RFLP analysis, that showed monophyletic origin of the two species of section Comopyrum (Terachi et al. 1984; Ogihara and Tsunewaki 1988; Tsunewaki 2009).

The species of this section grow in the western part of the distribution area of the diploid Aegilops species, i.e., western Turkey, Greece (incl. Crete) Albania, Macedonia, Serbia, Bosnia-Herzegovina, and Croatia. They grow on terra rossa soil in the edges of sclerophyllous and deciduous oak forests and maquis, in open and degraded dwarf shrub formations, pastures, abandoned fields, edges of cultivation and roadsides. It is common and forms dense stands in many localities and grows as altitudes of almost sea level to 1000 m.

9.7.2 Aegilops comosa Sm. in Sibth. & Sm.

9.7.2.1 Morphological and Geographical Notes

Ae. comosa Sm. in Sibth & Sm. [Syn.: Ae. subventricosa Jaub & Spach ex Bornm.; Triticum heldreichii (Holzm. ex Boiss.) K. Richt.; Ae. turcica Azn.; Triticum comosum (Sm.) K. Richt.; Ae. ambigua Hausskn.; Comopyrum comosum (Sm.) Á. Löve] is a predominantly autogamous, annual, tufted, multi-tillered plant. It has thin 10–35-cm-tall (excluding spikes) culms, that geniculate at the base then rising upright, with narrow, and usually hairy leaves. Its spike is linear-lanceolate to narrow-elliptical to narrow-oval, short (2–3.5-cm-long), usually rough or hairy, and gradually tapering towards the tip. The entire spike disarticulates at maturity (umbrella-type of dispersal unit). There are 3–4(-5), long, narrow to oval spikelets, that are shorter than the adjacent rachis segment. There are 1 and rarely 2 rudimentary spikelets at base. Each spikelet contains 3–4 florets, with the upper 1–2 being sterile. Glumes are more or less rough or hairy, with the upper parts overlapping. Glumes of lateral spikelets have two teeth, with an angle between them; sometimes one of the teeth develops into a short awn. The glumes of the terminal spikelet have 3 (seldom 1) large, 4–11-cm-long awns. The awns on each glume spread laterally from each other and the central awn of each glume diverges laterally from the central awn of the other glume of the spikelet. Lemmas are membranous, cartilaginous in the upper parts, and seldom awned in the upper spikelets. The caryopsis adheres to lemma and palea (Fig. 9.3b).

Ae. comosa is a highly polymorphic species. Variation involves spike and spikelet structure and number and length of awns. Eig (1929a) divided the species into two subspecies, ssp. eucomosa (currently comosa) and ssp. heldreichii (Boiss.) Eig. Ssp. eucomosa contains three varieties, var. comosa (=var. typica Eig), var. thessalica Eig, and var. ambigua Eig [=Ae. ambigua (Hausskn.) Eig]. Ssp. heldreichii contains four varieties: var. subventricosa Boiss., var. achaica Eig, var. biaristata Eig, and var. polyathera Hausskn. Hammer (1980), accepted Eig’s division, but changed the name of subsp. eucomosa to subsp. comosa.

The question if the taxa comosa and heldreichii is composed of two subspecies of Ae. comosa or of two different species, Ae. comosa and Ae. heldreichii, was a matter of dispute. Zhukovsky (1928), on a taxonomical ground, and Kihara (1940b), on a cytogenetic basis, regarded ssp. heldreichii as a separate species, Ae. heldreichii (Holzm.) Halac. Badaeva et al. (1999) supported this view of separation of these two taxa into two separate species, arguing that Ae. comosa and Ae. heldreichii were well-differentiated (though similar genomes) and consequently, must be considered separate species. This argument was based on the finding that the two taxa differ in heterochromatin content, morphology and C-banding pattern of several chromosomes, as well as in the number of the 5S rRNA gene loci (Badaeva et al. 1996a, 1999).

Teoh et al. (1983) also observed differences in the C-banding patterns of the two taxa: comosa had predominantly interstitial C-bands, whereas heldreichii had centromeric and telomeric bands. Yet, since this sort of change has no effect on chromosome pairing and fertility of the F₁ hybrids between the two taxa, they saw no justification to regard them as two separate species. The two taxa had similar karyotype structure, similar pattern of hybridization with pSc119 and pAs1, and similar distribution of major and minor NORs (Badaeva et al. 1999). Moreover, based on the observation of Kihara (1940b), that five bivalents and one quadrivalent are regularly formed at meiosis of the F₁ hybrid between the two subspecies and that the hybrid is fertile, Lilienfeld (1951) concluded that the two taxa should be at the intraspecific rank rather than at the specific one. In agreement to this, Chennaveeraiah (1960) reported only minor karyotypic differences between the two subspecies. Wang et al. (2000) analyzed nucleotide sequences of the internal transcribed spacer (ITS) of nuclear ribosomal DNA in Ae. comosa, and found that ssp. comosa and ssp. heldreichii were closely related and formed a sister-group relationship. The two subspecies grow sympatrically in the same geographic area and in the same ecological niches (van Slageren 1994). Since there is no noteworthy ecogeographical difference between them, the existence of a reciprocal translocation between the two taxa is not sufficient to separate them into two species.

Van Slageren (1994) considered the two intraspecific taxa as varieties, var. comosa and var. subventricosa Boiss. [syn: subsp. heldreichii Eig; Ae. subventricosa Jaub. & Spach ex Bornm.; Triticum heldreichii (Holzm. ex Boiss.) K. Richt.; Ae. turcica Azn.], which followed Art 35.3 of the International Code of Botanical Nomenclature (ICBN), adopted by the 14th International—Botanical Congress of Berlin 1987 (Greuter et al. 1988). According to this code, when the two taxa are classified at the variety level, the final epithet must be subventricosa, and when classified at the subspecific level, the epithet must be heldreichii. Since the epithet heldreichii is usually used by researchers of this group, it is adopted in this book and presented as ssp. heldreichii.

According to van Slageren (1994), the two taxa differ from one another by the following morphological characteristics: comosa has a cylindrical spike, is not inflated, and bears 3–4(-5) spikelets. The apical glume has 1 strongly developed, diverging, 4–11-cm-long central awn, and 2 lateral awns, more-slender, 3–7.5(-10) long awns. Heldreichii is shorter than comosa, with a monoliform spike that contains 1–2(-3) fertile spikelets; glumes of the apical spikelet have 1 strongly developed, diverging, 3–5.5-cm-long central awn, and 2 lateral, more-slender, 2–2.5-cm-long awns that are frequently reduced to teeth or absent. The spikelets of heldreichii are more inflated than those of comosa, and heldreichii has 1 + 3 awns on the terminal spikelet, while comosa has 3 + 3. The awns of heldreichii are usually shorter than those of comosa and the uppermost spikelet is smaller. Intermediate forms between the two taxa are frequent. The two taxa grow sympatrically in Greece, Albania and in western Turkey. Heldreichii does not grow in Cyprus and Bulgaria and is rare throughout its range, but possibly more common in Greece.

Ae. comosa distributes in the northeastern Mediterranean: in western Turkey, northern Cyprus, Greece (including Aegean Islands and Crete), Albania and southern Bulgaria. Greece is its main area of distribution. It grows on terra rossa soil and on dry, sandy, grassy and rocky places, in the edges of sclerophyllous and deciduous oak forests and maquis, in edges of and opening in garrigue, open and degraded dwarf shrub formations, cleared areas, pastures, abandoned fields, edges of cultivation and roadsides. It is common in Greece, frequently abundant and forms dense stands with restricted ranges. It is uncommon to rare in other regions of its distribution area. It grows at altitudes of almost sea level to 500 m, rarely 800 m.

Ae. comosa has a relatively limited distribution in the central-western region of the distribution of the genus. It is a Mediterranean element. Together with Ae. uniaristata, the other species of section Comopyrum, it occupies the western part of the distribution region of the diploid species of this group. It grows sympatrically with Ae. caudata, Ae. uniaristata, Ae. umbellulata, Ae. geniculata, Ae. biuncialis, Ae. neglecta, Ae. recta and Ae. triuncialis, and allopatrically with Ae. speltoides, Ae. peregrina, Ae. cylindrica and wild T. monococcum.

Ae. comosa is resistant to all the physiologic races of yellow rust (Puccinia striiformis) for which it has been tested (Riley et al. 1968).

9.7.2.2 Cytology, Cytogenetics, and Evolution

Ae. comosa is a diploid species (2n = 2x = 14), bearing the M genome (Kihara 1937, 1940a, b, 1954; Kimber and Tsunewaki 1988; Dvorak 1998). The organellar genome of its two subspecies, subsp. comosa and subsp. heldreichii, are designated M and M^h, respectively (Tsunewaki 1993; Ogihara and Tsunewaki 1988). The nuclear genome of Ae. comosa is small (1C DNA = 5.53 ± 0.052 pg) (Eilam et al. 2007 and Table 9.3). Eilam et al. (2007) determined DNA content in 3 different accessions of Ae. comosa, two belonging to subsp. comosa and one to subsp. heldreichii, that were collected from different regions of the distribution area of the species. Very little intraspecific variation in DNA content was found between the accessions.

According to Senyaninova-Korchagina (1932), the karyotype of Ae. comosa consists of 5 types of chromosomes including one pair with a large satellite on the short arm, one pair with an almost median centromere, and the rest with submedian centromeres. The karyotype described by Chennaveeraiah (1960) is asymmetric and differs in some respects from that described by Senyaninova-Korchagina (1932). It has two pairs with secondary constrictions, one pair with almost median centromere, three pairs with submedian centromeres and one pair with a somewhat submedian-subterminal centromere. One of the two pairs with the secondary constrictions has a large satellite, while the second pair has a small satellite; the satellites in both pairs are on the short arm.

Chennaveeraiah (1960) reported that the karyotype of ssp. heldreichii is almost similar to that of ssp. comosa. The differences in the karyotypes of these two subspecies are that ssp. heldreichii has two pairs with a submedian-subterminal centromere in contrast to one in ssp. comosa. In addition, two pairs in heldreichii versus one in comosa have almost median centromeres, and only one pair in heldreichii has a submedian centromere. The difference between the two subspecies lies mainly in the arm ratio of these pairs.

Similar results were obtained by Karataglis (1975), who studied the karyotypes of three varieties of Aegilops comosa from Greece, two (var. biaristata and var. subventricosa) of ssp. heldreichii, and one (var. thessalica) of ssp comosa. He found that the two varieties of ssp. heldreichii had the same chromosome morphology, but differed in their total chromosome length, in the length of the long arms and in the length of the satellites. The karyotype of var. thessalica was slightly different from those of heldreichii; the longest satellite of heldreichii occurred in the longest SAT chromosome, whereas the contrary was observed in the shortest SAT in the case of thessalica. From the two varieties of heldreichii, var. biaristata stands closer to var. thessalica than var. thessalica does to var. subventricosa. The great similarity between the karyotypes of the two varieties of heldreichii indicates their close phylogenetic relationship whereas the slight differences between the karyotypes of the varieties of heldreichii and that of comosa show some remoteness between the two subspecies, albeit insufficient to elevate them to the specific rank.

Teoh et al. (1983) used a repetitive DNA sequence, pTa71, that codes for ribosomal RNA, and which was derived from T. aestivum by Gerlach and Bedbrook (1979), as a probe in an in-situ hybridization experiment on the genome of Ae. comosa. They observed two pairs of rRNA sites in Ae. comosa, as expected from the presence of two pairs of SAT chromosomes (Chennaveeraiah 1960). An extra pair of rRNA gene cluster located terminally in some cells, but not on others, was also observed (Teoh et al. 1983). This third pair is not in correspondence with the number of satellites in this species.

C-banding studies also showed that the karyotypes of ssp. comosa and ssp. heldreichii are asymmetric, possessing metacentric, submetacentric and sub-telocentric chromosomes (Badaeva et al. 1996a). The distribution of the 18S-26S and 5S ribosomal RNA gene families on chromosomes of Ae. comosa was studied by in situ hybridization with pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) DNA clones (Badaeva et al. 1996b). Two major 18S-26S rDNA loci were found in the nucleolar organizer region (NOR) of chromosomes 1 M and 6 M. In addition to the major NORs, minor loci were observed in all the chromosomes, except for 6 M of ssp. comosa and 7 M of ssp. heldreichii (Badaeva et al. 1996b). Some minor loci were polymorphic, whereas others were conserved. All major and minor 18S-26S rDNA loci were on the short arms, except for chromosome 4 M (in both subspecies) and 3 M (in ssp. heldreichii), where the minor loci were on the long arm. One major 5S rDNA was located on the short arm of chromosome 1 M and a minor locus on the short arm of 5 M. The minor 5S rDNA on 5 M was absent in ssp. heldreichii.

The genomes of ssp. comosa and ssp. heldreichii had similar highly repetitive DNA sequence (Badaeva et al. 1996a) and NOR probe labelling patterns (1996b). However, they differed from each other in the amount and distribution of heterochromatin, the number of 5S rDNA loci, and the chromosomal locations of some minor 18S-26S rDNA hybridization sites (Friebe et al. 1996; Badaeva et al. 1996b).

The chromosomal locations of the major NORs were similar in Amblyopyrum muticum, in Ae. speltoides and in both subspecies of Ae. comosa (Badaeva et al. 1996b). However, Ae. comosa differs from A. muticum by the presence of several minor NORs, C-banding patterns, and the distribution of pSc119 and pAs1 DNA clones (Badaeva et al. 1996b). The phylogenetic proximity between A. muticum and Ae. comosa is evident from the data obtained from a comparative analysis of repeated nucleotide sequences in the Triticum–Aegilops group (Dvorak and Zhang 1992b). Thus, it seems that A. muticum contributed to the development of Ae. comosa via hybridization.

Cermeño et al. (1984b) analyzed the nucleolar activity in Ae. comosa via comparative analysis of somatic metaphase chromosomes by phase contrast, C-banding and Ag-staining. There were four Ag-NORs in Ae. comosa, and the number of nucleoli at interphase registered were one in 38, two in 121, three in 42, and four in 7 cells. Accordingly, the maximum number of nucleoli fits the expectation from the number of pairs of Ag-NORS and SAT chromosomes.

Teoh and Hutchinson (1983), using an improved C-banding technique, found that Ae. comosa has very faint and small centromeric bands in all chromosomes. The NORs can be clearly seen but are not banded. Moreover, there is an intraspecific variation in C-banding patterns differentiating the two subspecies of Ae. comosa, i.e., comosa and heldreichii (Teoh et al. 1983). Ssp. comosa has predominantly interstitial C-bands, in contrast to centromeric and telomeric bands in ssp. heldreichii. Yet, these polymorphic differences between the two subspecies have no effect on chromosome pairing or fertility of the F₁ hybrids between these two taxa (Kihara 1940a). Polymorphic differences in C-bands patterns appear to be widespread within and between the two subspecies of Ae. comosa and their non-random distribution seems to suggest that these differences could be intimately associated with the processes of subspeciation as suggested by Teoh et al. (1983).

Friebe et al. (1996) analyzed the C-banding pattern and polymorphisms in several accessions of the two subspecies of Ae. comosa and established the standard karyotypes of these subspecies. Although variation in C-band size and location was observed between different accessions, the different chromosomes were still distinct. The homoeologous relationships of these chromosomes were established by comparison of chromosome morphologies and C-banding patterns of comosa chromosomes to those of other diploid Aegilops species with known chromosome homoeology (Friebe et al. 1996). In addition, in situ hybridization analysis with a 5S rDNA probe was used to identify homoeologous groups 1 and 5 chromosomes. The present analysis enabled assignment of all the chromosomes of ssp. comosa and ssp. heldreichii to their homoeologous groups.

Teoh et al. (1983) used a non-coding repetitive DNA sequence, TC22b, which was derived from the B subgenome of hexaploid wheat by HaeIII restriction endonuclease digest (Hutchinson and Lonsdale 1982). Ae. comosa exhibited an intermediate amount of labeling with this probe. Comparisons between the in-situ hybridization and C-banding results indicated that, Ae. comosa, which has little heterochromatin, also shows little labeling with the TC22b probe.

Badaeva et al. (1996a), using in-situ hybridization with the highly repetitive DNA sequences pSc119 and pAs1, combined with C-banding, found that Ae. comosa possessed an intermediate amount of C-heterochromatin. The genome of ssp. heldreichii was characterized by a very low C-heterochromatin content, distinguishing it from ssp. comosa. Only few small C-bands were detected in this subspecies and they were located mainly at the telomeric regions.

Five chromosomes of Ae. comosa had hybridization sites with pSc119, whereas all chromosomes were labeled with pAs1. One to four pAs1 hybridization sites per chromosome were observed in Ae. comosa and each chromosome had a distinctive labeling pattern (Badaeva et al. 1996a). Yet, moderate levels of labeling were detected in ssp. comosa and ssp. heldreichii and the distribution of hybridization sites of both the pSc119 and pAS1 probes on chromosomes of these two subspecies was similar but not identical; the distribution of pAS1 sites in several chromosomes of ssp. heldreichii was different from the distribution in their homologous chromosomes of ssp. comosa.

The two species of section Comopyrum, Ae. comosa and Ae. uniaristata, display very different C-banding patterns (Teoh and Hutchinson 1983; Badaeva et al. 1996a). Kihara (1937, 1940a, b) designated the genome of Ae. uniaristata M^u because, based on chromosome pairing in the F₁ hybrid between Ae. comosa and Ae. uniaristata, he thought that the genome of Ae. uniaristata is related to genome M of Ae. comosa. However, Chennaveeraiah (1960) found major karyotypic differences between the two species and suggested to designate the genome of Ae. uniaristata N. Support for this suggestion came from the observation, made by Maan and Sasakuma (1978), of high univalent frequencies in amphiploid combinations containing these two species, indicating the homoeology level of their genomes. The different C-banded patterns of Ae. comosa and Ae. uniaristata reinforces this conclusion and the adoption of genome symbol N for Ae. uniaristata ((Teoh and Hutchinson 1983). Indeed, Kimber et al. (1983), based on meiotic chromosome pairing configurations in triploid, tetraploid and pentaploid hybrids involving Ae. comosa and Ae. uniaristata, found different genomes between the two, and consequently, proposed the genome symbol Uⁿ for Ae. uniaristata. Later, Kimber and Sears (1987) and Kimber and Feldman (1987) adopted the symbol N for the genome of Ae. uniaristata. In accord with the above, Yamane and Kawahara (2005) clustered Ae. comosa and Ae. uniaristata as sister species in one clade in the trees based on SSRs and cpDNA, and Haider et al. (2010) constructed two phylogenetic trees based on DNA analysis using RAPDs and ISSRs and found that Ae. comosa and Ae. uniaristata are sister species. Earlier studies had also revealed such a close genetic relationship between the two species based on cpDNA analysis (Terachi et al. 1984; Ogihara and Tsunewaki 1988; Haider 2003) and RFLPs (Dvorak and Zhang 1992b).

Ogihara and Tsunewaki (1988) and Haider and Nabulsi (2008) mentioned that the cpDNA in the two species showed identical restriction profiles using restriction enzymes. Similarly, the two species were clustered together based on the analysis of the 5S rDNA sequence (Appels et al. 1992) and restriction of repeated nucleotide sequences (Ogihara and Tsunewaki 1988). In another study carried out by Cuñado and Santos (1999), authors confirmed that there is a close relationship between the genomes of both species based on chromosome pairing in a hybrid of the two species. All these data agree with the classification of Eig (1929a), who treated these two species as sister species in the section Comopyrum.

Genome analysis studies (Kihara 1949) have shown that Ae. comosa was involved in the parentage of the allotetraploid species, Ae. geniculata (genome UUM^oM^o) and Ae. biuncialis (genome UUM^bM^b) of section Aegilops.

9.7.2.3 Crosses with Other Species of the Wheat Group

Meiosis in Ae. comosa was found to be very regular (Chennaveeraiah 1960); seven bivalents, mostly ring bivalents, were found in all 67 PMCs studied. Similar data were obtained by Cuñado (1992) who, using C-banding to study chromosome pairing at first meiotic metaphase of Ae. comosa ssp. comosa, observed an average of 0.89 rod bivalents and 6.11 ring bivalents (13.11 chiasmata/cell) and in ssp. heldreichii, an average of 0.02 univalents, 1.19 rod bivalents and 5.79 ring bivalents (12.77 chiasmata/cell).

Yamada and Suzuki (1941) studied chromosome pairing in F₁ hybrids between ssp. heldreichii and ssp. comosa and observed in most (about 90%) PMCs, an average of 5.00 bivalents and 1.00 quadrivalent. Metaphase cells with 7.00 bivalents or a univalent, 5.00 bivalents and 1 trivalent were also observed in about 10% of the PMCs. F₂ segregated to plants with two chromosome types in a ratio of 1:1, one with 7.00 bivalents and the other with 5.00 bivalents and 1 quadrivalent. The formation of a quadrivalent in these generations indicates the occurrence of reciprocal translocation between the two subspecies.

Kihara and Yamada (1942) and Kihara (1949) observed 1.00 univalent, 5.39 bivalents and 1.00 trivalent in the F₁ hybrid of Ae. comosa ssp. comosa x Ae. uniaristata, and a similar level of pairing in the ssp. heldreichii x Ae. uniaristata hybrid. Similar data in Ae. comosa x Ae. uniaristata hybrids were obtained by Sears (1941b) and Cuñado and Santos (1999) (Table 9.4), validating that the two species of section Comopyrum are closely related.

In the F₁ hybrid between Ae. speltoides x Ae. comosa Kihara (1949) reported on the presence of 1–7 bivalents (a mode of 6 bivalents), and 0–1 trivalent, whereas in the Ae. comosa x Ae. longissima hybrid, he found lower pairing, i.e., 0–6 bivalents (a mode of 3 bivalents), and 0–1 trivalent. The higher pairing in the hybrids that involve Ae. speltoides is presumably due to the action of the pairing promoters of Ae. speltoides. Nevertheless, the above pairing data show that the genome of Ae. comosa are homoeologous rather than homologous to those of the Sitopsis species. Similar conclusion was obtained from chromosome pairing in the F₁ hybrid between Ae. comosa or Ae. uniaristata and Sitopsis species (Table 9.4). Sears (1941b) observed higher pairing in the hybrid Ae speltoides (HP type) x Ae. comosa than in the hybrid Ae. speltoides (IP type) x Ae. uniaristata. Similar reduced level of pairing was observed in the hybrid Ae. sharonensis x Ae. uniaristata. The hybrid Ae. caudata x Ae. comosa had 4–6 bivalents (with a mode of 5 bivalents), and 0–1 trivalents, while the hybrid Ae. umbellulata x Ae. comosa had a lower level of pairing, i.e., 2–5 bivalents and 0–2 trivalents (Kihara 1949).

The hybrids between the allotetraploid species of section Aegilops bearing a modified M genome, namely, Ae. geniculata (genome M^oM^oUU) and Ae. biuncialis (genome UUM^bM^b) with Ae. comosa (genome MM), had 7 bivalents (Kihara 1949; Kimber et al. 1988) or 6 bivalents and a trivalent (Kimber et al. 1988), indicating that one of the subgenomes of these allotetraploids, subgenome M^o, is very closely related to the genome of Ae. comosa. The hybrid between Ae. comosa and another allotetraploid species of section Aegilops, i.e., Ae. columnaris (genome UUXⁿXⁿ), had much less pairing [3–7 bivalents and 0–3 trivalents (Kihara 1949)], indicating that none of the subgenomes of Ae. columnaris are closely related to the genome of Ae. comosa.

The hybrids with diploid wheat T. monococcum, either domesticated ssp. monococcum or wild ssp. aegilopoides, exhibited relatively low chromosomal pairing (Table 29). The hybrid with hexaploid wheat T. aestivum ssp. aestivum x Ae. comosa also showed very little chromosome pairing (Table 9.10). Evidently, the genome of Ae. comosa is only homoeologous to the subgenomes of allopolyploid wheats.

The cytogenetic relationships between Ae. comosa and Amblyopyrum muticum were studied by Ohta (1990). The F₁ hybrid Ae. comosa x A. muticum without B chromosomes of A. muticum showed a high frequency of chromosome pairing, whereas pairing in the hybrid with B chromosomes was much reduced (Table 8.2). All the hybrids with 0, 1, and 2 B chromosomes were male sterile (Ohta 1990).

B chromosomes of A. muticum effectively suppress pairing between homoeologous chromosomes but do not affect pairing of fully homologous ones (Mochizuki 1964; Dover and Riley 1972; Vardi and Dover 1972). Thus, the pairing in hybrids with to the genome of Ae. comosa. The hybrids Ae. caudata, Ae. uniaristata and Ae. umbellulata x A. muticum showed a lower frequency of A chromosome pairing than those from the hybrids Ae. comosa x A. muticum. This may indicate that the genome of A. muticum is closer to that of Ae. comosa than to those of these three species.

The hybrids between each of the two subspecies of Ae. comosa and A. muticum (without B chromosomes) showed high frequency of pairing (Ohta 1990). Yet, they differed in the frequency of multivalents. The hybrids involving ssp. comosa showed few or no quadrivalents, while those involving ssp. heldreichii frequently formed a quadrivalent or a trivalent. The occurrence of a reciprocal translocation between the two subspecies of Ae. comosa [Kihara (1937) and Yamada and Suzuki (1941)], causes the difference in the frequency of multivalents in hybrids involving these two subspecies. The high frequency of multivalents in the hybrid with ssp. heldreichii may indicate that this translocation was formed in ssp. heldreichii.

9.7.3 Aegilops uniaristata Vis.

9.7.3.1 Morphological and Geographical Notes

Aegilops uniaristata Vis. [Syn.: Triticum uniaristatum (Vis.) K. Richt.: Chennpyrum uniaristatum (Vis.) Á. Löve] is predominantly an allogamous, annual plant, tufted with many culms, which are 15–35-cm-tall (excluding spikes), and usually prostrate before turning upwards. Leaves are narrow, linear and usually hairy in the upper pars. The spike is moniliform, short, 1.5–4.0-cm-long (excluding awns), lanceolate to oval-lanceolate, glabrous, and tapering rapidly to the tip. The spike disarticulates entirely at maturity (umbrella-type dispersal unit). There are 3–5 number of spikelets, the terminal one usually being sterile, and 2–3 basal rudimentary spikelets. There are 4 florets, the upper 1–2 sterile. Glumes of the lateral spikelets have a conspicuous triangular tooth separated from an awn (up to 4-cm-long) by an acute angle. Glumes of the terminal spikelet end stepwise or abruptly in a broad, flat awn (3–5-cm-long), sometimes with an accompanying tooth. The middle nerve of the awn of the terminal glume is strongly projecting and is a continuation of the projecting nerve of the glume. Lemmas of the lateral spikelets have small teeth which sometimes elongate into small awns. The lemmas of the terminal spikelet have a weakly developed awn (up to 2-cm-long) and 1–2 adjacent teeth. The caryopsis adheres to lemma and palea (Fig. 9.3c).

Ae. uniaristata has limited morphological variation, primarily manifesting in spike and spikelet size. Kawahara (2000) analyzed genetic variation at 21 enzyme loci of Ae. comosa and Ae. uniaristata, the two species exhibited different levels of genetic variation; in Ae. comosa, the mean number of alleles per locus was 2.00 and the proportion of polymorphic loci was 0.667, while in Ae. uniaristata, they were 1.19 and 0.143, respectively. No heterozygotes were found, confirming that Ae. uniaristata and Ae. comosa are self-pollinating species, as was also reported by Hammer (1980). Since Ae. uniaristata occupies a relatively limited geographical area and is uncommon or rare throughout its distribution area (van Slageren 1994), it is probable that this species consists of small populations that are susceptible to the loss of genetic variation due to genetic drift during its evolution (Kawahara 2000).

Morphologically, Ae. uniaristata is close to Ae. comosa, particularly to several forms of Ae. comosa ssp. heldreichii, from which it differs in the following characters: the glumes of the terminal spikelet carry only one awn and those of the lateral spikelets have an awn and a large triangular tooth.

The taxonomists Zhukovsky (1928) and Eig (1929a), classified Ae. uniaristata and Ae. comosa as sister species: in section Comopyrum by Zhukovsky and in section Macrathera by Eig. Hammer (1980), and van Slageren (1994) gave priority, on the basis of earliest designation, to Comopyrum. The genome analysis studies of Kihara (1937, 1949, 1954) also arranged Ae. uniaristata and Ae. comosa in section Comopyrum. This classification was confirmed by the study of Yamane and Kawahara (2005) on the phylogenetic relationships among the diploid Aegilops-Triticum species. They found that SSRs and cpDNA clustered Ae. comosa and Ae. uniaristata as sister species in one clade in the phylogenetic trees. Likewise, the two phylogenetic trees that were constructed by Haider et al. (2010), based on DNA analysis using RAPDs and ISSRs, categorized Ae. uniaristata and Ae. comosa as sister species. Previous cpDNA analyses (Terachi et al. 1984; Ogihara and Tsunewaki 1988; Haider and Nabulsi 2008), and RFLPs (Dvorak and Zhang 1992b) also reported such close genetic relationship between these species. Ogihara and Tsunewaki (1988) and Haider and Nabulsi (2008) mentioned that the cpDNA of these species showed identical restriction profiles using restriction enzymes. Similarly, the two species were clustered together based on an analysis of the 5S rDNA sequence (Appels et al. 1992). In another study, Cuñado and Santos (1999) confirmed that there is a close relationship between the genomes of both species based on chromosome pairing in hybrids between the two species.

Ae. uniaristata has limited distribution in the central-western part of the distribution of the genus. It is a Mediterranean element that grows in European Turkey, Greece (including Crete and the Aegean Islands), Albania, Bosnia-Herzegovina, Serbia, and Croatia, and rarely in southeastern Italy. It is rare in European Turkey, sporadic in Greece, and more common in the Adriatic region of Bosnia-Herzegovina and Croatia.

Ae. uniaristata grows in grasslands and bushy slopes, mainly on rocky, calcareous soils (terra rossa), more rarely on sandstone, in edges of sclerophyllous Mediterranean oak forest and maquis, open or degraded dwarf shrub formations, pasture, disturbed habitats, edges of cultivation and roadsides, at altitudes of 0–750 m.

Ae. uniaristata occupies the northwestern corner of the distribution area of the diploid species of Aegilops. It also occupies the most mesophytic of all the habitats of the diploid species. It grows sympatrically with Ae. geniculata, Ae. biuncialis, Ae. neglecta, Ae. recta, Ae. triuncialis, and Ae. comosa (in Greece and European Turkey). It grows allopatrically with Ae. caudata, Ae. umbellulata and Ae. cylindrica.

Ae. uniaristata is tolerant to high levels of soil aluminum (Berzonsky and Kimber 1986, 1989), which remains effective when transferred to wheat (Miller et al. 1995, 1997). It is highly resistant to wheat stem and leaf rusts (Valkoun et al. 1985), and to some other fungal diseases (Gong et al. 2014).

9.7.3.2 Cytology, Cytogenetics, and Evolution

Ae. uniaristata is a diploid species (2n = 2x = 14), with the nuclear genome N (Kimber and Tsunewaki 1988; Dvorak 1998) and organellar genome N (Ogihara and Tsunewaki 1988; Wang et al. 1997). Haider and Nabulsi (2008) showed that Ae. uniaristata and Ae. comosa have very similar chloroplasts. This finding is in accord with Ogihara and Tsunewaki (1988), who reported that the cytoplasm of both species displayed identical restriction patterns when using 13 endonucleases. In contrast, there has been dispute concerning the nuclear genome of the two species. Originally, Kihara (1937, 1940a, b, 1949), based on genome analysis, gave to Ae. uniaristata the genomic symbol M^u, implying a genome modified from the M genome of Ae. comosa. Yet, Chennaveeraiah (1960) observed that the karyotype of Ae. uniaristata is unique and considerably different from that of Ae. comosa and, consequently, suggested to change its genome symbol to N. This conclusion was further supported by the data of Maan and Sasakuma (1978), who observed high univalent frequencies in amphiploid combinations containing Ae. uniaristata and Ae. comosa, indicating non-homology of their genomes. Kimber et al. (1983) examined meiotic chromosome pairing in triploid, tetraploid, and pentaploid hybrids involving the genomes of these two species, looking for values of relative affinity, and concluded that there is no preferential pairing between the chromosomes of these genomes. Consequently, they suggested that the M^u genome of Ae. uniaristata be changed to Un, and later, it was changed to N (Kimber and Sears 1987; Yen and Kimber 1992b). C-banding and FISH patterns of chromosomes (Teoh and Hutchinson 1983; Teoh et al. 1983; Badaeva et al. 1996a, b; Friebe et al. 1996; and Iqbal et al. 2000a, b) and isozyme studies (Kawahara 2000) confirmed that genome N is different from M. Ae. uniaristata is involved in the parentage of the allotetraploid Ae. ventricosa (genome DDNN) and the allohexaploid Ae. recta (genome UUXⁿXⁿNN).

Ae. uniaristata has the largest genome in section Comopyrum; its 1C nuclear DNA content = 5.82 ± 0.105 pg is significantly larger than that of Ae. comosa (1C DNA = 5.53 ± 0.052 pg) (Eilam et al. 2007). The karyotype of Ae. uniaristata was first described by Chennaveeraiah (1960) and was compared with that of Ae. comosa. It is an asymmetric karyotype containing only one pair with satellites, whereas in Ae. comosa there are two pairs with satellites. Of its six other pairs, two have sub-median centromeres, two have sub-median-sub-terminal centromeres, and the other two have sub-terminal centromeres. None of the distinctive karyotypic characteristics of Ae. comosa are seen in Ae. uniaristata (Chennaveeraiah 1960). A pair with a secondary constriction and an arm ratio of 1:1:1.5 characteristic to Ae. comosa was never found in Ae. uniaristata. In Ae. uniaristata, there are two pairs with sub-terminal centromeres which are not present in Ae. comosa. On the whole, the karyotype of Ae. uniaristata differs from, more than it resembles, the karyotypes of the two subspecies of Ae. comosa. The karyotypic changes in Ae. uniaristata may have occurred by inversions (Chennaveeraiah 1960).

The C-banding pattern at mitotic metaphase of Ae. uniaristata chromosomes was first described by Teoh and Hutchinson (1983), and was found to be unique and characteristically different from that of Ae. comosa. However, one common feature shared by both species is the absence of prominent telomeric bands. In Ae. uniaristata, all the centromeric regions have large and intensely stained bands. There are very few interstitial bands in both arms, which makes identification of chromosomes easier. Badaeva et al. (1996a) also found that large proximal C bands and some medium-sized interstitial bands were typical of the Ae. uniaristata N genome.

C-band findings in eight different accessions of Ae. uniaristata reported by Friebe et al. (1996) supported the observation of Teoh and Hutchinson (1983). Friebe et al. (1996) observed intraspecific variation in C-band size and location between different accessions, but this did not prevent chromosome identification and establishment of standard karyotypes. Similar to the report by Chennaveeraiah (1960), Friebe et al. (1996) found that the karyotype of Ae. uniaristata consisted of two sub-metacentric and five more or less acrocentric chromosome pairs, all with diagnostic markers.

Teoh et al. (1983) used two different cloned repetitive DNA sequences as probes in in-situ hybridization experiments on the genome of Ae. uniaristata. The TC22b probe is a highly repeated, noncoding 260-bp DNA segment, derived from the B subgenome of hexaploid wheat (Hutchinson and Lonsdale 1982). The rRNA probe they used, described by Gerlach and Bedbrook (1979), consists of 18S and 25S rDNA with an associated spacer DNA. Ae. uniaristata chromosomes exhibited little labeling with TC22b and the large centromeric heterochromatic bands showed hardly any labeling with this highly repetitive probe. In-situ hybridization with the rDNA probe revealed only one pair of rRNA sites. The location of these sites was on the one pair of satellited chromosomes that was described by Chennaveeraiah (1960).

Badaeva et al. (1996a) performed in-situ hybridization with the highly repetitive noncoding DNA sequences, pSc119, derived from Secale cereale, and pAsl, derived from Ae. tauschii, and observed strong labeling with both pSc119 and pAs1, indicating that the karyotype of this species was highly heterochromatic. The pSc119 probe hybridized to the telomeres of all chromosomes and the distribution of its hybridization sites was similar to that seen in Ae. umbellulata and Ae. caudata, whereas the strong and unique hybridization sites with pAs1 differed from labeling in Ae. umbellulata, Ae. caudata and Ae. comosa (Badaeva et al. 1996a). This difference, combined with differences in chromosome morphology, karyotypic features, and C-banding, indicates again that Ae. uniaristata and Ae. comosa have different genomes.

The distribution of the 5S and 18S-26S ribosomal RNA gene families on Ae. uniaristata chromosomes was studied by in-situ hybridization with pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) DNA clones (Badaeva et al. 1996b). Similar to the finding of Teoh et al. (1983), only one major 18S-26S rDNA locus was found in the single pair of the nucleolar organizer region (NOR). This chromosome pair was identified as chromosomes 5N by hybridization with rDNA. In addition to the major NORs, several minor loci were also observed in Ae. uniaristata, some of which were polymorphic, while others were conserved. The presence of only one pair of chromosomes with major NORs in Ae. uniaristata (and in Ae. tauschii), and their location on the short arms of homoeologous group 5, is in sharp contrast to the presence of two such pairs, on homoeologous groups 1 and 6, in all other diploid Aegilops species.

Two 5S rRNA loci were observed in Ae. uniaristata, one major locus located in the distal part of the long arm of chromosome 1N and one minor locus located in the short arm of chromosome 5N. The 5S rDNA loci were not associated with NORs. The different distribution of 5S rRNA loci in Ae. uniaristata and Ae. comosa also confirm that these two species have distinct genomes.

The nucleolar activity of the NORs in somatic chromosomes of Ae. uniaristata was analyzed using a highly reproducible silver-staining procedure (Cermeño et al. 1984b. Two Ag-NORs were identified, one nucleolus was seen in 262 cells at interphase and two nucleoli were seen in 269 cells. All diploid Aegilops species analyzed, except for Ae. uniaristata and Ae. tauschii, showed 4 nucleolus organizer chromosomes, further supporting the difference of Ae. uniaristata and Ae. tauschii in this respect from the rest of the diploid Aegilops species.

The homoeologous relationships of the chromosomes of Ae. uniaristata were established by comparison of their morphologies and C-banding patterns to chromosomes of other diploid Aegilops species with known chromosome homoeology (Friebe et al. 1996). In addition, in-situ hybridization analysis with a 5S rDNA probe was used to identify chromosomes of homoeologous groups 1 and 5. This analysis permitted the assignment of three Ae. uniaristata chromosomes to their homoeologous groups. The homoeology of the remaining four acrocentric chromosomes could not be determined. Chromosome 5N is sub-metacentric with a secondary constriction and a small satellite in the short arm. The middle of the short arm of this chromosome has a 5S rDNA in-situ hybridization site. The other sub-metacentric chromosome pair was identified as 4N. The least sub-telocentric chromosome pair had a 5S rDNA in situ hybridization site in a distal region of the long arm, suggesting that this chromosome is at least partially homoeologous to group 1 chromosomes, and was designated accordingly. One accession of Ae. uniaristata was homozygous for a whole-arm translocation involving chromosomes 1N and 5N.

Iqbal et al. (2000b) performed RFLP analyses on wheat-Aegilops uniaristata addition lines and translocation lines to confirm the identity of the added N-genome chromosomes. Complete 1N, 3N, 4N, 5N and 7N chromosome additions were identified, while the complete long arm and only part of the short arm was identified for chromosome 2N. There were no wheat-like 4/5 and 4/7 translocations in the Ae. uniaristata chromosomes (Iqbal et al. 2000a, b). Chromosome 3N carried an asymmetric pericentric inversion. Chromosome-specific RAPD and microsatellite markers were also identified all the added Ae. uniaristata chromosomes available in this set of addition lines.

Hybridization sites of the repetitive DNA sequences pAs1, pSc119.2 and pTa71 were identified on the N-genome chromosomes of Ae. uniaristata using the FISH technique (Iqbal et al. 2000a, b). Like Badaeva et al. (1996a), Iqbal et al. (2000a, b) also found six pairs of Ae. uniaristata chromosomes showing strong hybridization signals with pAs1, while an additional pair showed weak signals with this probe. pAs1 is a D genome specific probe isolated from Ae. tauschii (Rayburn and Gill 1986) and shows strong hybridization sites with the D subgenome of common wheat, indicating the relatedness of the N genome to the D subgenome of wheat. For pSC119.2, three pairs of chromosomes showed one signal at the telomeres of their short arms, while two pairs had signals at both the short- and the long-arm telomeres. Two pairs of chromosomes showed three sites on each chromosome, one at the telomere of the short arm and two on the long arm.

Construction of the genetic map of A. uniaristata chromosome 3N revealed the important role that asymmetric pericentric inversions played in the evolution of the N-genome (Iqbal et al. 2000a). This mechanism led to the formation of all four sub-telocentric chromosomes of A. uniaristata and caused transposition of the 5S rDNA locus from the short arm of chromosome 1N to its long arm.

Gong et al. (2014) performed a C-banding analysis on Ae. uniaristata chromosomes present in the amphiploid Triticum turgidum–Ae. uniaristata and on the available set of T. aestivum cv. Chinese Spring–Ae. uniaristata addition lines (6N addition was lacking in this set). Their study showed easily recognizable C-banding patterns for chromosomes 1N–5N and 7N that were distinguishable from wheat chromosomes based on chromosome size and the position of the C-bands. Moreover, chromosome 6N was easily recognized by comparing the C-bands of the six N-genome chromosomes (1N–5N and 7N) to the seven N-genome chromosomes in the amphiploid Triticum turgidum–Ae. uniaristata. Similar to the results of Teoh and Hutchinson (1983) and those of Badaeva et al. 1996a), Gong et al. (2014) found that the C-bands mainly exist in the centromeric regions of Ae. uniaristata and rarely at the distal ends.

Moreover, FISH on mitotic metaphase chromosomes of the amphiploid T. turgidum–Ae. uniaristata, using SSR (GAA)₈ as a probe, showed that the hybridization signals of all Ae. uniaristata chromosomes differ from those of T. turgidum. Thus, (GAA)₈ can be used to identify all Ae. uniaristata chromosomes in wheat background simultaneously (Gong et al. 2014). In addition, a total of 42 molecular markers specific for Ae. uniaristata chromosomes were developed by screening expressed sequence tag—sequence tagged site (EST-STS), expressed sequence tag—simple sequence repeat (EST-SSR), and PCR-based landmark unique gene (PLUG) primers (Gong et al. 2014) These markers were subsequently localized using the T. aestivum–Ae. uniaristata addition lines and different wheat cultivars as controls (Gong et al. 2014).

Evolution of the N-genome has also been associated with amplification, elimination and re-distribution of the different families of repetitive DNA sequences (Badaeva et al. 2011). The most interesting example is the emergence of pericentromeric heterochromatin bands on Ae. uniaristata chromosomes, which was probably caused by massive amplification of certain classes of highly repetitive DNA sequences. Large pericentromeric C-bands are also found on the chromosomes of A. speltoides; however, the results of a comparative study of several Aegilops species using a TC22b probe, (Teoh et al. 1983), suggest that the molecular composition of pericentromeric heterochromatin differ between Ae. uniaristata and Ae. speltoides species. Bardsley et al. 1999) also found that probes for the SSR repeats (AAC)₅, (ACG)₁₀ and (CGT)₁₀, which are the major components of C-bands in wheat and many Aegilops species, hybridized poorly to the N-genome chromosomes. Pericentromeric C-bands on A. uniaristata chromosomes also did not contain the pAs1 sequence. Moreover, the long arm of chromosome 7N, which displayed the highest amount of hybridization with the pAs1 sequence, contained only few very faint interstitial C-bands, and conversely, the most heterochromatic chromosome, 4N, possessed only two very small pAs1 sites. Thus, the molecular composition and organization of heterochromatin in the A. uniaristata genome is distinct from other species and remains to be characterized.

9.7.3.3 Crosses with Other Species of the Wheat Group

Chromosome pairing at first meiotic metaphase of Ae. uniaristata was studied by Cuñado (1992), using the C-banding technique. Chromosome pairing was regular with average configurations per PMC of 0.03 univalents, 4.07 rod bivalents, 2.90 ring bivalents and 9.88 chiasmata. Seven bivalents existed in most cells; the relatively large frequency of rod bivalents resulted from the presence of four chromosome pairs with sub-median–subterminal and sub-terminal centromeres (Chennaveeraiah 1960).

Chromosome pairing in F₁ hybrids between the two species of section Comopyrum, Ae. uniaristata and Ae. comosa, showed a high number of bivalents, with most being rod bivalents (Table 25). This can result partly from the presence of several chromosome pairs with sub-median–subterminal and sub-terminal centromeres in Ae. uniaristata and partly from reduced homology. Cuñado and Santos (1999) studied chromosome pairing in this hybrid by electron microscopy in surface-spread-first meiotic prophase nuclei and compared the results with light-microscopic observations of first metaphase cells after C-banding and FISH. At first prophase, the hybrid showed extensive synapsis and complex multivalents, involving up to 14 chromosomes, but at first metaphase, most associations were in the form of bivalents between homoeologous chromosomes as follows: the average pairing between the M and the N genomes was 5.08 rod bivalents, 0.33 ring bivalents and 0.08 trivalents. Yet, there was also autosyndesis pairing between chromosomes of genome M (0.01 rod bivalents and 0.03 trivalents). In the hybrid comosa x uniaristata, the mean bivalent frequency at first prophase (3.07) was lower than that observed at first metaphase (5.41). Therefore, a considerable number of bivalents formed by homoeologous chromosomes must be involved in the complex multivalent associations observed at first prophase (Cuñado and Santos 1999).

Chromosomal pairing in hybrids was more or less the same in hybrids between Ae. uniaristata and each of the two subspecies of Ae. comosa. Average chromosome pairing in the hybrid involving ssp. comosa contained 1.00 univalent, 5.00 bivalent and 1.0 trivalents and with ssp. heldreichii, 5.48 bivalents (Kihara and Yamada 1942). Percival (1932) observed 0–4 univalents, 5–7 bivalents (mostly rod), and an occasional trivalent in the hybrid Ae. uniaristata x ssp. heldreichii. The above data show that ssp. comosa differs by a reciprocal translocation from Ae. uniaristata, whereas such translocation does not exist between ssp. heldreichii and Ae. uniaristata, indicating that the latter may evolved from ssp. heldreichii.

Chromosome pairing in hybrids involving two Sitopsis species, Ae. speltoides (IP type) and Ae. sharonensis, with Ae. uniaristata (Table 9.4) showed fa reduced number of bivalents, most of which were rod, indicating that the genomes of these species have diverged considerably. Likewise, Ae. uniaristata x Ae. bicornis had 1–6 bivalents and 0–2 trivalents (Kihara 1949), Ae. longissima x Ae. uniaristata had 0–6 bivalents with a mode of 2, and 0–1 trivalents (Kihara 1949). The diploid hybrid Ae. tauschii x Ae. uniaristata had 5.94 univalents, 3.63 rod bivalents, 0.08 ring bivalents and 0.14 trivalents (Sears 1941b). In the reciprocal hybrid, Cuñado and Santos (1999) observed a similar level of pairing, i.e., 3.08 rod bivalents, 0.08 ring bivalents and 0.14 trivalents. Evidently, the genomes of these two species have diverged considerably from one another. Chromosomal pairing in hybrids of Ae. uniaristata with the diploid species Ae. caudata and Ae. umbellulata was also relatively low, indicating divergence of the genome of Ae. uniaristata from those of Ae. caudata and Ae. umbellulata. More specifically, the pairing in the hybrid Ae. caudata x Ae. uniaristata had 4.50 univalents, 3.50 rod bivalents, 0.44 ring bivalents, 0.46 trivalents and 0.06 quadrivalents (Sears 1941b), or 2–5 bivalents (Kihara 1949). In the hybrid with Ae. umbellulata, Percival (1932) observed 6–14 univalents and 0–4 bivalents (all rod) and Sears (1941b) found 8.30 univalents, 2.70 bivalents (all rod) and 0.10 trivalents.

Chromosome pairing with the allotetraploid Ae. crassa (genome D^cD^cX^cX^c) was similar to the amount of pairing with Ae. tauschii, but had a somewhat higher frequency of trivalents (Table 9.5). On the other hand, chromosomal pairing with Ae. ventricosa (genome DDNN), that possesses subgenome N from Ae. uniaristata, was significantly higher (Table 9.5). Chromosome pairing in hybrids with the allohexaploid cytotype of Ae. crassa (genome D^cD^cX^cX^cDD) showed higher pairing, most of which was presumably autosyndesis between chromosomes of the two D subgenomes of hexaploid Ae. crassa. In contrast, the hybrid with the allohexaploid species Ae. juvenalis (genome D^cD^cX^cX^cUU), that contains only one D subgenome, showed a significantly low level of pairing (McGinnis and Melnyk 1962).

Similarly, chromosome pairing in the hybrid Ae. geniculata (genome M^oM^oUU) x Ae. uniaristata (genome NN) indicated low homology and great divergence between the genomes of these species (Table 9.5). The low level of pairing in this hybrid is in spite of the presence of the modified M subgenome of Ae. geniculata and the N genome of Ae. uniaristata. A similarly low level of chromosome pairing was observed in the hybrids of Ae. columnaris (genome UUXⁿXⁿ) x Ae. uniaristata (4–6 bivalents and 0–1 trivalents) and of Ae. neglecta (genome UUX^tX^t) x Ae. uniaristata (2–6 bivalents with mode of 4) (Kihara 1949).

Sears (1941b) observed a low pairing in the hybrids between T. monococcum, either domesticated or wild, and Ae. uniaristata (Table 9.8), indicating great divergence between the genomes of these two diploid species. Likewise, chromosome pairing in F₁ hybrids Ae. uniaristata x Amblyopyrum muticum without B chromosomes was relatively low, indicating a considerable divergence of the genomes of the two species (Table 8.1). The hybrids were sterile (Ohta 1990).

9.8 Section Aegilops

9.8.1 General Description

Section Aegilops (Syn.: sect. Surcullosa Zhuk.; sect. Polyides Zhuk.; sect. Pleionathera Eig) contains annual, predominantly autogamous species. The plants are slender, with short spikes, which are more or less ovate, elliptic or lanceolate in outline, rarely elongate-linear, mostly awned, and fall as a unit at maturity (umbrella-type dispersal unit). There are 2–5 spikelets, which are ventricose or elliptic, and rarely linear. mostly awned; Spikelets are reduced and not inflated at the upper part of the spike, and usually less than 10 times long as wide, rarely more. The glumes of apical spikelets have 3–4 awns, those of lateral spikelets with 2–4 (-5) awns and or teeth. Glume awns are always stronger than lemma awns. There are 1–4 rudimentary spikelets at the base of the spike. The caryopsis is free or adhered to lemmas and palea (Fig. 9.4).

Fig. 9.4

Plants and spikes of Aegilops species carrying the U genome; a Ae. umbellulata Zhuk.; b Ae. geniculata Roth; c Ae. biuncialis Vis.; d Ae. neglecta Req. ex Bertol.; e Ae. recta (Zhuk.) Chennv.(from Kimber and Feldman 1987); f Ae. columnaris Zhuk.; g Ae. triuncialis L.; h Ae. kotschyi Boiss. i Ae. peregrina (Hack. in J. Fraser) Maire & Weiler ssp. euvariabilis Eig; and j Ae. peregrina ssp. cylindrostachys Eig

Section Aegilops contains nine species: one diploid [Ae. umbellulata Zhuk. (genome UU), seven allotetraploids [Ae. geniculata Roth (genome M^oM^oUU), Ae. biuncialis Vis. (genome UUM^bM^b), Ae. neglecta Req. ex Bertol. (Genome UUXⁿXⁿ), Ae. columnaris Zhuk. (Genome UUXⁿXⁿ), Ae. triuncialis L. (genome UUCC), Ae. kotschyi Boiss. (genome S^lS^lUU) and Ae. peregrina (Hack. in J. Fraser) Maire & Weiler (genome S^lS^lUU)] and one allohexaploid, [Ae. recta (Zhuk.) Chennv. (Genome UUXⁿXⁿNN)]. The spike of all the allopolyploid species resembles that of Ae. umbellulata, the U genome donor (Fig. 9.4).

Section Aegilops was divided by Eig into two subsections: subsection Libera Eig, which contains all species with free caryopsis, and subsection Adhaerens Eig, which contains the two species, Ae. kotschyi and Ae. peregrina, which have an adherent caryopsis.

The distribution of the species of section Aegilops is very wide, one of them, Ae. triuncialis, distributes in almost all the distribution area of the genus whereas several others distribute from western Mediterranean to West Asia. The distribution of the single diploid species in the section, Ae. umbellulata, is relatively wide, falling in the central part of the genus distribution area, namely, from Greece to west and north Iran. The distribution of the allopolyploid species, on the other hand, is, in most cases, very wide, extending beyond that of their diploid progenitors, and covering a large part of the genus distribution area. While the distribution of Ae. triuncialis is extremely large almost overlaps the genus distribution area, that of the three species, Ae. geniculata, Ae. biuncialis and Ae. neglecta, is more in the Mediterranean part of the genus distribution area. The distribution of Ae. columnaris extends from the center eastwards up to Iran, Ae. peregrina grows in the south and east Mediterranean parts, whereas Ae. kotschyi, the steppical species, grows from the south (Libya) toward the east, up to Pakistan. The distribution of the single allohexaploid species, Ae. recta, is more limited than that of its allotetraploid progenitor, Ae. neglecta; it was found in Portugal, Spain, France, Italy, Croatia, Greece and Turkey. All species of this section (except Ae. kotschyi) grow on a great variety of soils (e.g., terra rossa, basalt, alluvial, stand sands and sandy loams), Ae. kotschyi grows on loess and grey calcareous steppe soil. All species occupy a large number of habitats and often infest cultivated fields as weeds. The allotetraploids are great colonizers, strive well in disturbed habitats, and often form very dense, mixed populations.

9.8.2 Aegilops umbellulata Zhuk.

9.8.2.1 Morphological and Geographical Notes

Aegilops umbellulata Zhuk. [Syn.: Triticum umbellulatum (Zhuk.) Bowden; Ae. ovata var. anatolica Eig; Kiharopyrum umbellulatum (Zhuk.) Á. Löve] is a predominantly autogamous, annual plant, tufted with many, upward-bent, 10–30-cm-tall (excluding spikes) culms. Leaves are linear, 2–5-cm-long, and more or less hairy. The spike is lanceolate-ovoid, 1.5–4-cm-long, and typically rough. The entire spike disarticulates at maturity (umbrella-type dispersal unit). Spikelets 5–6, rarely 3, the upper 1–3 being sterile, so the ear suddenly becomes narrow, rudimentary basal spikelets 3, rarely 2. The rachis of the lower spikelets is much shorter than the adjacent spikelet. There are four florets, the upper two being sterile. The rachis segment of the upper spikelets is much longer than the adjacent spikelet causing the narrow upper part of the spike to protrude from the lower wider part. Glumes are similar, shorter than the spikelets, and suddenly inflated above the middle, above which they narrow to a deeply incised margin. Glumes of the lower spikelets have 4–5 (3–6) awns, while glumes of the upper spikelets have 3–5 awns, all of which are similar in shape. Lemmas have 1–3 awns, all resembling the glume awns. All awns diverge at maturity, producing a characteristic umbel shape. The lemma of the lower spikelets has 8–12 veins near the upper margin. The caryopsis is free (Fig. 9.4a).

Morphological variation involves mainly spike size, color and hairiness. There are two forms which differ in spike size, and sometimes grow in mixed populations and in such stands, intermediates may be found. According to Hammer (1980), Ae. umbellulata contains two subspecies, subsp. umbellulata (glumes with 5–7 awns) and subsp. transcaucasua Dorof. et Migusch. (glumes with 3–4 awns), the former contains two varieties, var. umbellulata (glabrous glumes) and var. pilosa Eig (hairy glumes).

Ae. umbellulata is an East Mediterranean/Western Asiatic element extending into the steppical (Irano-Turanian) region. It is predominantly occurring in Turkey, but also present in Greece (including Rhodes and the Aegean Islands), Syria, Lebanon, North Iraq, West and Northwest Iran, Armenia, and Azerbaijan. It is common and locally abundant, growing at altitudes from sea level to 1800 m, on terra rossa, basalt, alluvial and grey calcareous steppe soils in the edges and openings of sclerophyllous or deciduous oak forest or maquis, degraded dwarf shrub formations, open dwarf shrub semi-steppe and steppe-like formations, abandoned fields, edges of cultivation and roadsides. It often grows as a weed in cultivated areas. Ae umbellulata prefers more humid conditions than many other Aegilops species, with annual rainfall from 350 to 700 mm.

Ae. umbellulata has a wide distribution in the central region of the distribution of the genus. It occupies a large variety of primary and secondary habitats. In the central part of the genus distribution area, Ae. umbellulata occurs in mixed populations with many other species of Aegilops and has sporadic contact with others. It grows sympatrically with Amblyopyrum muticum, Ae. speltoides, Ae. caudata, Ae. comosa, Ae. tauschii, wild T. monococcum, wild T. timopheevii, Ae. geniculata, Ae. biuncialis, Ae. neglecta, Ae. recta, Ae. columnaris, Ae. triuncialis, Ae. peregrina, and Ae. cylindrica, and allopatrically with Ae. longissima, Ae. searsii, Ae. uniaristata, wild T. turgidum, Ae. kotschyi, Ae. crassa, and Ae. juvenalis.

Ae. umbellulata is resistant to leaf rust (Sears 1956), stem rust (Ozgen et al. 2004), stripe rust (Bansal et al. 2017), powdery mildew, hessian fly and green bug (Gill et al.1985). Sears (1956), using a radiation treatment, pioneered the transfer of a gene resistant to leaf rust, Lr9, from chromosome 6U of Ae. umbellulata to common wheat chromosome 6B (Sears 1941a).

9.8.2.2 Cytology, Cytogenetics and Evolution

Ae. umbellulata is the only diploid species in the section Aegilops having the basic chromosome number of the section (2n = 2x = 14). Both its nuclear genome and organellar genome were designated U (Kimber and Tsunewaki 1988; Dvorak 1998). It is involved in the parentage of the seven allotetraploids of section Aegilops, namely, Ae. geniculata (genome M^oM^oUU), Ae. biuncialis (genome UUM^bM^b), Ae. neglecta (genome UUXⁿXⁿ), Ae. columnaris (genome UUXⁿXⁿ), Ae. triuncialis (genome UUCC), Ae. peregrina (genome S^lS^lUU), and Ae. kotschyi (genome S^lS^lUU), and of two allohexaploids (Ae. recta (genome UUXⁿXⁿNN) from section Aegilops and Ae. juvenalis (genome D^cD^cX^cX^cUU) from section Vertebrata (Kihara 1954, 1957; Kihara and Tanaka 1970; Kimber and Feldman 1987; Kimber and Tsunewaki 1988; Dvorak 1998). As the donor of the U genome to all the allopolyploids of the section, Ae. umbellulata contributed many of its morphological features (glume awns, sudden narrowing of the spike, free caryopsis, inflation of the glumes and whole-spike disarticulation) to its related allopolyploids.

Kihara and Lilienfeld (1932) designated the genome of Ae. umbellulata C^u and considered it to be a modified genome derived from the C genome of Ae. caudata Consequently, Kihara (1954) placed it, together with Ae. caudata, in the C-genomic group, based on the assumption that Ae. triuncialis (2n = 4x = 28; genome UUCC), containing the genomes of Ae. umbellulata and Ae. caudata, was considered to be an autotetraploid by von Berg (1931), who observed the autosyndetic behavior of the C and C^u genomes of Ae. triuncialis in a hybrid with Secale cereale. Kihara (1954) also based his grouping of Ae. umbellulata in the same genomic group with Ae. caudata, on the formation of some multivalents in a synthetic amphiploid of Ae. triuncialis (Karpechenko and Sorokina (1929). Yet, Kihara (1954) himself expressed the inaccuracy of placing these two species in one group, since the view of autopolyploidy of Ae. triuncialis was rejected by several cytogenetic studies. Also, there was little or no support of the assumption of Kihara and Lilienfeld (1932) that the two genomes were not widely differentiated at the time of their incorporation into the tetraploid Ae. triuncialis. Sears (1948) questioned the genomic similarity of the C and C^u genomes on both cytological and morphological grounds, pointing out that both the C- and C^u-genome chromosomes paired with M-genome chromosomes of Ae. comosa at about the same frequency as they paired with each other, and further, that the spike morphology of the two species is very different. Such weak chromosomal affinity in meiosis of hybrids between these two species was also found by Sears (1941b) and Kihara (1949). Kimber and Abu-Baker (1981) reached the same conclusion by investigating chromosome pairing in a series of hybrids involving Ae. umbellulata and Ae. caudata. These findings agree with the taxonomic treatment of Ae. umbellulata and Ae. caudata that, based on their morphology, placed the two species in two different sections (Zhukovsky 1928), Eig (1929a), Hammer (1980), and van Slageren (1994). In addition, the karyotypes of Ae. umbellulata and Ae. caudata are widely different (Chennaveeraiah 1960). Consequently, it was concluded that the C^u genome of Ae. umbellulata is not a modified C genome of Ae. caudata and should, therefore, be given a separate genomic status (Chennaveeraiah 1960; Kimber and Abu-Baker 1981). Thus, the symbol U was assigned to the genome of Ae. umbellulata and is also used in the genomic descriptions of allopolyploid species with Ae. umbellulata in their parentage (Chennaveeraiah 1960; Kimber and Abu-Baker 1981). The major differences in the C-banding patterns of Ae. caudata and Ae. umbellulata provide additional support for the differential designation of the Ae. umbellulata and Ae. caudata genomes (Teoh and Hutchinson 1983).

The nuclear genome of Ae. umbellulata is significantly larger than that of Ae. caudata, equal to that of Ae. comosa and smaller than the genome of Ae. uniaristata (Eilam et al. 2007; Table 9.3). Its 1C DNA size, determined in 9 accessions that were collected from different regions of the distribution area of the species, is 5.53 ± 0.052 pg (Eilam et al. 2007). Very little intraspecific variation in DNA content was found between the accessions.

The karyotype of Ae. umbellulata is asymmetric. Senyaninova-Korchagina (1932) recognized several types of chromosomes that included one pair with large satellites on the short arm, one pair with a median centromere, four pairs with sub-median-sub-terminal centromeres, and one pair with a characteristic knob due to an extremely sub-terminal centromere. The karyotype described by Chennaveeraiah (1960), although essentially similar to that observed by Senyaninova-Korchagina (1932), differed from it in one respect. One of the pairs with a sub-median-sub-terminal centromere also featured a satellite on the shorter arm, but the satellite was considerably smaller, approximately 1/3 of the large satellite. Thus, the karyotype consists of 6 types of chromosomes, one pair with a large satellite on the short arm, one pair with a small satellite on the short arm, one pair with a sub-median centromere, two pairs with sub-median-sub-terminal centromeres, another smaller pair with a sub-median centromere and one pair with the characteristic knob due to an extreme sub-terminal centromere. The shortest chromosome is about ¾ the size of the largest one. A similar description of the Ae. umbellulata karyotype was presented by Al-Mashhadani et al. (1978), who analyzed the karyotype of several Aegilops species native to Iraq.

Ae. umbellulata has a complex C-banding pattern with intensely stained pericentromeric, telomeric and interstitial bands (Teoh and Hutchinson 1983). Telomeric bands occur on most of the long arms. Chromosomes 2U, 4U, and 5U according to the designation of Friebe et al. (1995b), [chromosomes D, F, and C according to Kimber’s (1967) designation] are easily distinguishable by their C-banding patterns, but the other chromosomes also exhibit specific C-banding patterns. Chromosomes 1U and 5U are the SAT chromosomes that are clearly differentiated by their banding patterns (Teoh and Hutchinson 1983). Badaeva et al. (1996a) also found that Ae. umbellulata is among the most heterochromatic species with large centromeric, intercalary and terminal C-bands present on all chromosomes.

While Teoh and Hutchinson (1983) analyzed only one accession of Ae. umbellulata, Friebe et al. (1995b) studied ten different accessions, collected from different geographic regions, and established a standard karyotype and a generalized ideogram of Ae. umbellulata, based on C-banding of these accessions. In addition, these authors identified, by C-banding and GISH, the individual Ae. umbellulata chromosomes in T. aestivum cv. CS– Ae. umbellulata chromosome addition lines, produced by Kimber (1967). These six umbellulata chromosomes were designated by Kimber (1967) as follows: A (subterminal chromosome), B (SAT chromosome), C (SAT chromosome), D (subterminal chromosome), E (subterminal chromosome), G (subterminal chromosome). Friebe et al. (1995a, b, c) determined the homoeology of these six umbellulata chromosomes in the addition lines as follows: 1U = B, 2U = D, 4U = F, 5U = C, 6U = A, and 7U = E). [The missing chromosome, F (=3U), is sub-median; Kimber (1967)].

Ae. umbellulata has two pairs of SAT-chromosomes, 1U and 5U, that were described earlier by the presence of secondary constrictions (Chennaveeraiah 1960). These pairs were shown by in-situ hybridization (ISH) using a ribosomal DNA probe (Teoh et al. 1983), and by Ag-Nor banding (Cermeño et al. 1984b), to possess NORs. The NOR in 1U is usually more prominent than that in 5U. Chromosome 6U is the shortest chromosome with the extreme sub-terminal centromere. Some polymorphism for C-band size and position was observed between the different accessions. However, this did not prevent chromosome identification (Friebe et al. 1995b). No large structural rearrangements detectable by C-banding analysis were found in any of the accessions. Yet, Badaeva et al. (1996b) reported that line TA1965 of Ae. umbellulata from Iran has a reciprocal translocation of chromosomes 1U and 5U.

The homoeology of chromosome 6U of Ae. umbellulata was also determined by studying its pairing relationships with wheat telocentric lines in hybrids having the chromosome constitution of 20 wheat chromosomes + one wheat telocentric chromosome + seven Ae. speltoides chromosomes that promote homoeologous pairing + chromosome 6U (Athwal and Kimber 1972). It has been found that chromosome 6U is homoeologous to the group-6 chromosomes of wheat. Castilho and Heslop-Harrison (1995) performed ISH with the 5S and the 18S–5.8S–25S rRNA genes and the repetitive DNA sequence pSc119.2, on Ae. umbellulata chromosomes. The pSc119.2 probe hybridized with all Ae. umbellulata chromosomes at the telomeres, except for the short arm of chromosome 6U, and showed intercalary sites on the long arms of chromosomes 6U and 7U. The 5S and 18S–25S rDNA only mapped physically on the short arms of chromosomes 1U and 5U. On chromosome 1U, 5S rDNA was shown to be sub-terminal and 18S–25S rDNA more proximal, while on chromosome 5U, the position of the genes was reversed. Similar result was obtained by Badaeva et al. (1996b), using in situ hybridization with pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) DNA probes. Also, Martini et al. (1982), studying common wheat plants into which 1U and 5U chromosomes of Ae. umbellulata had been separately introduced, found that these two chromosomes possess ribosomal RNA genes. ISH analysis showed that the two addition lines with 1U and 5U have more ribosomal RNA genes and rDNA clusters than the recipient wheat plants. The repeating rDNA unit in Ae. umbellulata is longer than most of the units in the standard laboratory wheat cultivar Chinese Spring (Martini et al. 1982). The additional DNA is probably in the non-transcribed spacer, as probably suggested by restriction endonuclease maps of rDNA. In Chinese Spring plants possessing 1U or 5U chromosomes, the largest nucleoli formed on 1U or 5U chromosomes and the wheat NORs formed only micronucleoli. This is not because the NORs on chromosomes 1U and 5U have many more rRNA genes than the wheat NORs, rather, Martini et al. (1982) suggested that they compete more effectively for some limiting factor. The partial inactivation of the wheat NORs by chromosomes 1U or 5U does not result in reduced total nucleolus volume in root tip or PMCs, because of the compensation by the NORs of chromosomes 1U or 5U.

Lacadena and Cermeño (1985) analyzed the influence of U genome chromosomes on NOR activity of common wheat, in the complete set of the chromosome addition lines of Ae. umbellulata to common wheat. Chromosomes 1U and 5U induced partial inactivation of wheat NORs of chromosome 6B, 1B and 5D. Chromosomes 2U and 3U, which are not SAT-chromosomes, also influenced the activity of wheat NORs. The predominant status of the U genome with respect to nucleolar competition in common wheat was in accord with its predominance in the allopolyploid species of Aegilops that contains the U genome (Cermeño et al. 1984b).

Teoh et al. (1983) probed the Ae. umbellulata genome with two different cloned repetitive DNA sequences derived from common wheat, one, TC22b, being a noncoding sequence and the other coding for ribosomal RNA. Chromosomes exhibited very little labelling with TC22b. Like in Ae. uniaristata, Ae. umbellulata also had large centromeric heterochromatic bands which did not correlate with heavy labelling by the TC22b probe. On the other hand, ISH with the repetitive DNA sequence that codes for ribosomal RNA, confirmed the presence of two rDNA sites in Ae. umbellulata.

Using the probe pAs1 from Ae. tauschii in ISH experiments with Ae. umbellulata, revealed only a few minor pAs1 sites on chromosomes 1U and 6U (Badeva et al. 1996a). Faint inconsistent signals were detected in several other U-genome chromosomes. In contrast, the probe pSc119 from Secale cereale hybridized to telomeric regions and some interstitial sites of most Ae. umbellulata chromosomes (Badaeva et al. 1996a).

The C genome of Ae. caudata and the U genome of Ae. umbellulata differ significantly in heterochromatin content (Friebe et al. 1995b), but minimally in C-band patterns (Badaeva et al. 1996a). The number and chromosomal location of hybridization sites with the pAs1 and pSc119 probes were similar in the two species (Badaeva et al. 1996a). This kind of similarity in ISH patterns suggests that the C and U genomes are related. Yet, both genomes were significantly rearranged during speciation and their chromosomes show little affinity in meiosis of hybrids between the two species (Kimber and Abu-Baker 1981).

Zhang et al. (1998), using RFLP probes that detect homoeoloci previously mapped in hexaploid wheat, constructed a comparative genetic map of Ae. umbellulata with wheat. It was found that all seven Ae. umbellulata chromosomes displayed one or more rearrangements relative to wheat, changes that are consistent with the sub-terminal morphology of chromosomes 2U, 3U, 6U and 7U. Comparison of the chromosomal locations assigned by mapping and those obtained by hybridization to wheat –Ae. umbellulata single chromosome addition lines verified the composition of the added Ae. umbellulata chromosomes and indicated that no further cytological rearrangements had taken place during the production of the alien-wheat aneuploid lines. Relationships between Ae. umbellulata and wheat chromosomes were also confirmed, based on homoeology of the centromeric regions.

Edae et al. (2017) developed a framework consensus genetic map for Ae. umbellulata comprising 3009 genotype-by-sequence SNPs with a total map size of 948.72 cM. On average, there were three SNPs per centimorgan for each chromosome. Chromosome 1U was the shortest (66.5 cM), with only 81 SNPs, whereas the remaining chromosomes had between 391 and 591 SNP markers. A total of 2395 unmapped SNPs were added to the linkage maps through a recombination frequency approach, and brought the number of SNPs placed on the consensus map to a total of 5404 markers. Segregation distortion was disproportionally high for chromosome 1U for both populations used to construct component linkage maps, and thus, segregation distortion may be one of the reasons for the exceptionally reduced linkage size of chromosome 1U. From comparative analysis, all Ae. umbellulata chromosomes, with the exception of 4U, showed moderate to strong collinearity with corresponding homoeologous chromosomes of hexaploid wheat and barley. The present consensus map may serve as a reference map in QTL mapping and validation projects, and in genome assembly of a reference genome sequence for Ae. umbellulata.

Zymogram analysis was used to identify the Ae. umbellulata chromosomes that carry the structural genes for particular isozymes (Benito et al. 1987). It was found that umbellulata chromosome 6U carries a structural gene for 6-phosphogluconate dehydrogenase, chromosome 1U carries structural genes for glucose phosphate isomerase and phosphoglucose mutase, chromosome 2U carries genes for leaf peroxidases, chromosome 7U carries structural genes for endosperm peroxidases, acid phosphatases and leaf esterases, chromosome 4U carries a gene for embryo plus scutellum peroxidases, and chromosome 3U carries structural genes for endosperm alkaline phosphatases, leaf alkaline phosphatases and leaf esterases. The results obtained indicate that chromosome 1U is partially homoeologous to the common wheat chromosomes of group 1 and 4, and chromosome 7U is partially homoeologous to common wheat chromosomes of groups 7 and 4.

9.8.2.3 Crosses with Other Species of the Wheat Group

Meiosis in Ae. umbellulata is regular; seven bivalents were formed in every pollen mother cell (PMC) of which 3–4 were rod and 3–4 ring (Chennaveeraiah 1960). Likewise, Cuñado (1992) observed an average of 3.07 rod bivalents, 3.93 ring bivalents, (total of seven bivalents), and 10.94 chiasmata per cell. Obviously, Ae. umbellulata, having several sub-telocentric chromosomes, displays less ring bivalents than species with symmetric karyotype.

Data on average chromosome pairing between allotetraploid species of section Aegilops containing a U subgenome with Ae. umbellulata, signifies the homology between this subgenome and that of the diploid (Table 9.5). Average chromosome pairing in meiotic metaphase of F₁ hybrids between Sitopsis Species and Ae. umbellulata was low. Thus, hybrids involving two Sitopsis species, Ae. speltoides and Ae. sharonensis, with Ae. umbellulata shows that their genomes are very distantly related (Table 9.4). More specifically, the hybrid Ae. speltoides var. ligustica I (HP type) x Ae. umbellulata had 3.22 univalents, 2.98 rod bivalents, 0.0 ring bivalents, 1.58 trivalents and 0.03 quadrivalents, whereas the hybrid Ae. speltoides var. ligustica (IP type) x Ae. umbellulata had 9.08 univalents, 1.98 rod bivalents, 0.0 ring bivalents, and 0.32 trivalents. Similar to the hybrid with the HP type of Ae. speltoides, also the hybrid Ae. sharonensis x Ae. umbellulata had low pairing. Similarly, Tanaka (1955a) observed 5–11 univalents, 1–5 bivalents with a mode of 3, 0–1 trivalents and 0–1 quadrivalents in the hybrid Ae. sharonensis x Ae. umbellulata. Chromosome pairing in the hybrid Ae. bicornis x Ae. umbellulata had 0–5 bivalents (with a mode of 3) and 0–3 trivalents (Kihara 1949), and in the hybrid Ae. longissima x Ae. umbellulata 0–4 bivalents (with mode of 2) and 0–1 trivalents (Kihara 1949).

Average chromosomal pairing in the F₁ hybrid Ae. umbellulata x Ae. tauschii displayed 3.36 rod bivalents, 0.0 ring bivalents and 0.66 trivalents (Cuñado and Santos 1999). Similar data were obtained by Kihara (1949). Average chromosome pairing in hybrids between Ae. caudata and Ae. umbellulata included 5.64 univalents, 3.46 rod bivalents, 0.12 ring bivalents and 0.40 trivalents, and in another hybrid of Ae. caudata x Ae. umbellulata 5.64 univalents, 3.46 rod bivalents, 0.12 ring bivalents and 0.40 trivalents (Sears 1941b). Somewhat higher pairing in the hybrid Ae. caudata x Ae. umbellulata was reported by Kihara (1949), with 3–6 bivalents (with mode of 5) and 0–1 trivalents. Nevertheless, the above data indicate low affinity between the genomes of these two species. In the F₁ hybrid Ae. umbellulata x Ae. comosa, Kihara (1949) observed 2–5 bivalents and 0–2 trivalents. In the hybrid Ae. uniaristata x Ae. umbellulata, Percival (1932) observed 0–4 bivalents, while Sears (1941b) observed 8.30 univalents, 2.70 rod bivalents, 0.0 ring bivalents and 0.10 trivalents). Evidently, the genome of Ae. umbellulata is closer, although not very close, to the genome of Ae. caudata than to those of Ae. comosa and Ae. uniaristata.

Chromosomal pairing between hexaploid Ae. crassa (genome D^cD^cX^cX^cDD) x Ae. umbellulata included 13.94 univalents, 4.06 rod bivalents, 1.46 ring bivalents (5.52 total), 0.81 trivalents, 0.09 quadrivalents and 0.03 hexavalents (Melnyk and McGinnis 1962). Most of the pairing is presumably autosyndetic between chromosomes of genomes D^c and D of Ae. crassa. The rest of the pairing indicate very little affinity between the U genome and the subgenomes of hexaploid Ae. crassa. In contrast, two reports of chromosome pairing in the hybrid Ae. juvenalis (genome D^cD^cX^cX^cUU) and Ae. umbellulata had 6.39 univalents, 3.59 rod bivalents, 4.46 ring bivalents (8.05 total), 1.42 trivalents, 0.28 quadrivalents and 0.03 hexavalents (McGinnis and Melnyk1962) and 6.93 univalents, 4.80 rod bivalents, 4.42 ring bivalents (9.22 total), 0.27 trivalents and 0.08 quadrivalents (Kimber and Abu-Baker 1981), indicating that a subgenome in Ae. juvenalis is homologous to the genome of Ae. umbellulata.

Average chromosomal pairing in the F₁ hybrid between T. monococcum subsp. aegilopoides and Ae. umbellulata had low pairing (Table 9.8). Evidently, the genomes of these two species are very distantly related. The hybrid between hexaploid wheat, T. aestivum ssp. aestivum and Ae. umbellulata was very low (Table 9.10) Obviously, the subgenomes of hexaploid wheat are not closely related to the genome of Ae. umbellulata.

In certain lines of domesticated diploid wheat, T. monococcum subsp. monococcum, Sears (1944a) identified two alleles which act as dominant-lethal that are responsible for unviability of hybrids with Ae. umbellulata, but without an effect in T. monococcum itself. The alleles differ in the time at which they cause death. A third normal allele is present in the wild subspecies of T. monococcum, i.e., subsp. aegilopoides.

Chromosome pairing in the F₁ hybrid Ae. umbellulata x Amblyopyrum muticum without muticum B chromosomes, showed intermediate level of pairing and many multivalents (Table 8.1). Hence, pairing was low in spite of the presence of homoeologous pairing promoter genes in A. muticum. The range of chromosome pairing in hybrids with two B chromosomes, which suppress homoeologous pairing, was much lower. Evidently, the genome of Ae. umbellulata is only distantly related to that of A. muticum.

9.8.3 Aegilops geniculata Roth

9.8.3.1 Morphological and Geographical Notes

Ae. geniculata Roth, commonly known as ovate goat grass, [Syn.: Ae. ovata L.; T. ovatum (L.) Raspail:] is a predominantly autogamous, annual, tufted, multi-tillered plant. It has thin, 10–40-cm-tall (excluding spikes) culms, that geniculate at the base and then rise upright, with narrow, hairy or glabrous leaves. The upper 1/3 or 1/4 of the culms is defoliated. Leaf blades are usually short, 2–5-cm-long. Its spike is broad-oval to narrow-elliptical, short (1.5–3.0-cm-long), does not become suddenly narrow, and is usually awned. The entire spike disarticulates at maturity (umbrella-type dispersal unit). There are 3(2–4) oval or urceolate spikelets that are appressed to the rachis, all potentially fertile, aside from the terminal, which is usually sterile. The lower rachis segment is usually shorter than the adjacent spikelet. There is one basal rudimentary spikelet, and rarely 2 or none. There are 5 florets in each spikelet, with the upper 3 being sterile. Glumes are 7–8-mm-long, with veins (5–7) that are unequal in width and unequally spaced, and are inflated at the middle, usually with 3–4(-5) awns (2.0–3.5-cm-long), equal in width. Upper spikelets usually have more awns (4–5), which are as long as or shorter than lateral spikelet awns. There are usually two lemma awns on lower spikelets, and three on upper spikelets. All awns are widely spread at maturity. There are 5–7 veins near the upper margin of the lemma of the lower spikelets. The caryopsis is free (Fig. 9.4b).

Ae. geniculata exhibit very large variation in spike shape, size, hairiness, and compactness, spikelet shape, site of glume inflation, and awn count, length and structure. In the southwestern part of its distribution, there is a compact form with irregular awns. The wide morphological variation led taxonomists to subdivide the species into two subspecies, each containing several varieties, albeit, with some disagreement on the sub-specific classification. Thus, Zhukovsky (1928) subdivided Ae. geniculata into four subspecies, namely, gibberosa, umbonata, globulosa and planiuscula. Eig (1929a) divided Ae. geniculata into two subspecies, euovata (currently subsp. geniculata), that contains four varieties, and subsp. atlantica, that contains three varieties. Hammer (1980) also subdivided the species into two subspecies: geniculata and gibberosa. Ssp. geniculata includes six varieties and is characterized by loose arrangement of spikelets in the ear, awns that have the same width, and often by more than 3 spikelets per spike, whereas subsp. gibberosa includes three varieties and is characterized by more compact arrangement of spikelets in the ear, and rarely with more than 3 spikelets per spike.

Medouri et al. (2015) evaluated morphological polymorphism among Ae. geniculata accessions from Algeria and found a great variation in most of the studied traits. A weak relationship between morphological traits and ecological factors was found. They also revealed high polymorphism among accessions in the high molecular weight (HMW) glutenin subunits and discovered several new subunits. Mahjoub et al. (2009, 2010), used morphological traits and RAPD markers to assess genetic diversity in Ae. geniculata from North and Central Tunisia. Both morphological traits and RAPD markers showed a high degree of variation within and between populations. Yet, gene diversity was attributable mostly to diversity within populations. The morphological variation was associated with environmental (climatic) change. A non-significant correlation was found between morphological and RAPD variations. Similar results of high variation in European and Tunisian Ae. geniculata accessions, determined using RAPD, was also reported by Zhang et al. (1996) and Mahjoub et al. (2016), respectively. On the other hand, Monte et al. (2001) reported only slight variation in Ae. geniculata accessions from Spain, whereas, when using RAPD and ISSR, Thomas and Bebell (2010) found great genetic diversity in accessions of this species from Greece, the center of its distribution. Similarly, AFLP analysis of Ae. geniculata accessions from Turkey revealed high polymorphism in this species (Kaya et al. 2011).

Ae. geniculata is a widespread Mediterranean and Western Asiatic element. It grows in Macaronesia (Madeira, Tenerife), Portugal, Spain, South France (including Corsica), Italy (including Sardinia and Sicily), Malta, Slovenia, Croatia, Serbia, Bosnia-Herzegovina, North Macedonia, Albania, Bulgaria, Greece, (including Crete and the Aegean Islands), South Ukraine, South Russia (including Crimea and Ciscaucasia), rare in Transcaucasia (Georgia, Armenia, and Azerbaijan), Turkey (Northwest and Mediterranean), Northern Iraq, Western Iran, Syria, Lebanon, Cyprus, Israel, Jordan, Egypt, Libya, Tunisia, Algeria and Morocco. In several regions of North-Africa, it penetrates into the Sahara. Ae. geniculata grows on terra-rossa, basalt, rendzina, calcareous sandstone and alluvial soils. It is found in edges and openings of sclerophyllous oak forests, garrigue and maquis, shrub and herbaceous formations, fallow fields, roadsides, disturbed habitats, and often on the edges of and within cultivated plantations and wheat fields (with which it may form natural hybrids). It grows at altitudes from almost sea level to 1750 m. Ae. geniculata is adventive in parts of central and northwestern Europe (van Slageren 1994) and was introduced into parts of United States, e.g., California, where it is a noxious weed invading dry land pastures (Hitchcock 1935).

Ae. geniculata grows with its two diploid parents in Turkey and Greece. However, its distribution is larger than that of its diploid parents and it occupies more types of habitats that its diploid parents. It is common, locally abundant, forming dense stands throughout its range in both primary and secondary habitats. Yet, it is sporadic in marginal and semi-steppical regions. Like other Aegilops allotetraploids, Ae. geniculata is a typical colonizer species. The two diploid parents grow together in western Turkey and in Greece and this is presumably the center of origin of the species.

Ae. geniculata is very widely distributed in the central and western part of the distribution of the genus. With the exclusion of Ae. bicornis and Ae. vavilovii, both of which grow in xeric habitats, and Ae. juvenalis, Ae. geniculata has contact with all the species of the wheat group. Ae. geniculata grows sympatrically with most of the specpes of Aegilops and wild, and allopatrically with A. muticum, Ae. bicornis, Ae. tauschii, Ae. kotschyi, and Ae. juvenalis.

Ae. geniculata usually grows in mixed stands with other Aegilops species, particularly with those belonging to the U-genome cluster, i.e., species of section Aegilops, with which it may form natural hybrids and introgressed derivatives. In many mixed populations of Ae. geniculata, Ae. peregrina and Ae. biuncialis in Israel, natural hybrids, hybrid derivatives and many highly introgressed types involving Ae. geniculata have been found, indicating a wide occurrence of gene flow between these three species, resulting in increased variation (Zohary and Feldman 1962; Feldman 1965a, b, c). It is assumed, therefore, that in many parts of the Ae. geniculata distribution area, mixed populations of Ae. geniculata and other allopolyploid species of section Aegilops, such interspecific genetic connections leading to introgression that blurs, to some extent, the specific boundaries. Evidence indicating gene flow that occurred between Ae. geniculata and common wheat, when the former grew near or in wheat fields, was presented by Zaharieva and Monneveux (2006), Loureiro et al. (2006), Loureiro and Escorial (2007), and by Arrigo et al. (2011). Spontaneous hybridization between tetraploid wheat (T. turgidum ssp. durum) and Ae. geniculata was also regularly observed in sympatric populations, resulting in spontaneous formation of durum-geniculata fertile amphiploids that arose through unreduced gametes of the F₁ hybrids (David et al. 2004).

Ae. geniculata contains genes that confer resistance to several fungal diseases, such as powdery mildew (Gill et al. 1985; Stoilova and Spetsov 2006), leaf rust (Gill et al. 1985; Zaharieva et al. 2001b; Aghaee-Sarbarzeh et al. 2002; Anikster et al. 2005), stem rust (Zaharieva et al. 2001b; Liu et al. 2011a, b), and yellow (stripe) rust (Zaharieva et al. 2001b; Aghaee-Sarbarzeh et al. 2002; Anikster et al. 2005), resistance to barley yellow dwarf virus (BYDV) (Zaharieva et al. 2001b), Hessian fly (Gill et al. 1985), common root rot (Bailey et al. 1993), and cereal cyst nematodes (CCN) (Zaharieva et al. 2001b). It also contains a gene that may improve tolerance to abiotic stresses related to water status, chlorophyll content and plant thermal regulation under Mediterranean field conditions (Zaharieva et al. 2001a). The genetic potential of Ae. geniculata to improve resistance to biotic stresses and tolerance to abiotic stresses in wheat harbors invaluable potential for wheat improvement.

9.8.3.2 Cytology, Cytogenetics and Evolution

Ae. geniculata is an allotetraploid (2n = 4x = 28) species. Its nuclear genome designation is M^oM^oUU and organellar genome is M^o (Kimber and Tsunewaki 1988; Dvorak 1998). Kihara (1937, 1954) designated the genome of Ae. geniculata C^uC^uM^oM^o (C^u is currently U) primarily due to the chromosomal pairing observed in hybrids with other Aegilops allotetraploids. Kihara (1929) and Kihara and Lilienfeld (1932) observed 5–11 bivalents in the F₁ hybrid Ae. triuncialis (genome C^uC^uCC; currently UUCC) and Ae. geniculata and in the reciprocal combination and concluded that the C^u (U) subgenome of Ae. triuncialis also exists in Ae. geniculata. Likewise, Kihara (1949) observed 6–9 bivalents and 0–3 trivalents in the F₁ Ae. geniculata x Ae. columnaris hybrid (genome C^uC^uM^cM^c; currently UUXⁿXⁿ), confirming the inclusion of the U genome in Ae. geniculata. On the other hand, Kihara (1929) observed 5–10 bivalents in the F₁ hybrid of Ae. geniculata x Ae. ventricosa (genome DDM^uM^u; currently DDNN) and configurations of 10 bivalents + 8 univalents were frequently observed in the F₁ of the reciprocal cross (Kihara and Lilienfeld 1932). Therefore, it was concluded that one of the two subgenomes in Ae. geniculata is close to the M genome of Ae. uniaristata or Ae. comosa. This conclusion was accepted because pairing in the F₁ hybrid Ae. geniculata x T. aestivum (genome BBAADD) had only 2–3 bivalents (Kihara and Lilienfeld 1932; Riley 1966a), indicating that Ae. geniculata does not carry the D subgenome. Genome analysis following direct crosses between Ae. geniculata and its putative diploid progenitors was performed by Kihara (1937), who observed 7 bivalents in the Ae. geniculata x Ae. umbellulata (genome UU) hybrid. Kimber and Abu-Baker (1981) reported the presence of 6 bivalents and one trivalent in the F₁ of this hybrid combination, thus, presenting direct evidence that one of the subgenomes of Ae. geniculata is U of Ae. umbellulata. Likewise, Kimber et al. (1988) reported the presence of 7 bivalents or 6 bivalents and one trivalent in the F₁ Ae. geniculata x Ae. comosa (genome MM) hybrid, verifying the presence of the M subgenome in Ae. geniculata. Yet, based on meiotic analysis of a number of hybrids, Kimber et al. (1988) concluded that, while the U subgenome of Ae. geniculata is much closer to the U genome of Ae. umbellulata than the M subgenome is to the M genome of Ae. comosa, and suggested that the M subgenome underwent substantial modifications at the tetraploid level. By this conclusion, they reinforced Kihara’s (1937, 1954) view that the M subgenome of Ae. geniculata is a modified genome. The presence of a modified M-subgenome in Ae. geniculata was further confirmed by cytogenetic and molecular data (Talbert et al. 1993; Friebe et al. 1999; Resta et al. 1996; Badaeva et al. 2002, 2004).

Resta et al. (1996) investigated the origin of the Ae. geniculata subgenomes by examining specific restriction fragments of repeated nucleotide sequences in the DNA of this allotetraploid species. The analysis showed that Ae. geniculata is closely related to Ae. biuncialis; in both species, one subgenome was closely related to the genome of Ae. umbellulata and the other was a modified genome of Ae. comosa. Consequently, they proposed the same genome formula, UUM°M°, for Ae. geniculata and Ae. biuncialis. Ae. neglecta and Ae. columnaris, which are also closely related to each other and have the same genomes, share the U genome with Ae. geniculata and Ae. biuncialis, but their second pair of subgenomes is unrelated to the M^o subgenome.

A similar conclusion was reached by Gong et al. (2006) who, using 31 ISSR primers, found that the genome constituents of Ae. geniculata had considerably changed as compared to the genomes of its ancestral diploid species. The genetic similarity index between Ae. geniculata and Ae. umbellulata was much higher than that with Ae. comosa, indicating that the U subgenome of Ae. geniculata had changed relatively minimally while its M^o subgenome was significantly modified.

Karyological studies also showed that one chromosomal set of Ae. geniculata corresponds to that of Ae. umbellulata (Senjaninova-Korczagina 1932; Pathak 1940; Chennaveeraiah 1960). The second chromosomal set is different, bearing median or sub-median centromeres and with no satellites or secondary constrictions (Senvaninova-Korchaginova 1932; Chennaveeraiah 1960). Since the karyotype of the second set differed from that of all the diploid species of Aegilops, Chennaveeraiah (1960) supported Kihara’s (1937, 1954) claim that the second chromosomal set of Ae. geniculata, the M^o genome, became modified at the tetraploid level.

Maan (1977b) reported that the cytoplasm of Ae. geniculata is similar to that of Amblyopyrum muticum. Similarly, Tsunewaki (1996) and Wang et al. (1997) suggested that a form of A. muticum might be the maternal and Ae. umbellulata the paternal parent of Ae. geniculata. Yet, Terachi et al. (1984), based on restriction fragment patterns of chloroplast DNA, concluded that the chloroplast genome of Ae. geniculata is unique, being distinctly different from that of all species of sections Aegilops and Comopyrum as well as from that of Amblyopyrum muticum. No diploid species with a chloroplast genome closely related to that of Ae. geniculata has been found in the Triticum, Aegilops or Amblyopyrum genera (Ogihara and Tsunewaki 1982; Terachi et al. 1984). Even Ae. umbellulata and A. muticum, whose chloroplast genomes are closest to that of Ae. geniculata, showed seven differences in ctDNA restriction fragments when compared to that of Ae. geniculata. It was therefore suggested that the cytoplasm of Ae. geniculata was derived from the M^o genome donor (Mukai and Tsunewaki 1975; Tsunewaki 1980). Consequently, the plasma type of Ae. geniculata was designated M^o (Tsunewaki 1980; Tsunewaki and Tsujimoto 1983; Kimber and Tsunewaki 1988; Dvorak 1998), and is different from that of the related allotetraploid species, Ae. biuncialis, Ae. neglecta and Ae. columnaris, that have the cytoplasm of Ae. umbellulata.

Tsunewaki (2009), reviewing the results of his and his coworkers on plasmon (plastom and chondrion) analysis of the Aegilops-Triticum group, maintained the conclusion that the origin of the M^o plasmon of Ae. geniculata is unclear. This allotetraploid species shares a genome constitution, UM, with Ae. biuncialis, and has a genome constitution related to that of Ae. columnaris and Ae. neglecta. Its plasmon, however, greatly differed from those of these three species in most respects, although similar to the Ae. columnaris plasmon in its phenotypic effects, and to the T, T2 and U plasmons of A. muticum and Ae. umbellulata, respectively, in the plastom.

Assuming the degree of plasmon dissimilarity between two species parallels differences in time of origin, Wang et al. (1997) and Tsunewaki (2009) speculated relative time of the origin of related polyploids by comparing their plasmon similarity with their closest diploid relatives. Thus, plasmon differences were compared between Ae. geniculata and its phylogenetically related species of section Aegilops (Ae. biuncialis, Ae. neglecta and Ae. columnaris) and the plasmons of the closest diploid relatives. The results suggest that the origin of Ae. geniculata is more ancient than that of the other related allotetraploid species. A similar conclusion on the early origin of Ae. geniculata was reached by Terachi et al. (1984) on the basis of plastom analysis.

Ae. geniculata contains 10.29 ± 0.008 pg 1C DNA (Eilam et al. 2008), which is 5.68% less DNA than that expected from the sum of the DNA of its two diploid parents, i.e., 10.91 pg (Ae. comosa contains 5.53 pg and Ae. umbellulata contains 5.38 pg; Eilam et al. 2007). The loss of DNA in Ae. geniculata was confirmed by Badaeva et al. (2002, 2004) who, based on differential C-banding and in situ hybridization, found that this allotetraploid exhibits substantial structural chromosome rearrangements, including deletion of chromosomal segments and reduction of heterochromatin content.

Early studies of the karyotype of Ae. geniculata revealed only one pair of SAT chromosomes on the somatic complement (Senyaninova-Korchagina (1932) or two such pairs on short arms (Matsumura 1940), and Pathak (1940) noted the presence of one pair with a satellite and another pair with a secondary constriction. Chennaveeraiah (1960) confirmed the presence of two SAT pairs on short arms, and also reported one pair with a knob due to extreme sub-terminal constriction, one pair with a median centromere, eight pairs with sub-median centromeres and four pairs with sub-median-sub-terminal centromeres.

Chennaveeraiah (1960) analyzed the karyotype of two taxa of Ae. geniculata, var. brachyathera (Pomel) Eig [=Ae. geniculata ssp. gibberosa (Zhuk.) Hammer] and subsp. globulosa Zhuk., and found no significant differences between them. In both, two satellite pairs were observed, confirming the observations of Matsumura (1940) and Pathak (1940). Both had a pair with a median centromere, another pair with an almost median centromere, and a pair with the extreme sub-terminal centromere. The rest of the pairs in both taxa had sub-median and sub-median-sub-terminal centromeres. The only difference between the two taxa was that ssp. globulosa had more pairs with sub-median-sub-terminal centromeres than var. brachyathera.

Chromosome morphology and variation in N-band distribution along the Ae. geniculata chromosomes were studied in 13 accessions by Landjeva and Ganeva (2000). The N-banding technique differentially stained all 14 chromosome pairs, enabling their identification. The most prominent bands were observed near the centromeres and in the intercalary regions of both arms, whereas telomeric bands were found in only seven chromosome pairs. Polymorphism for presence or absence of particular bands was observed among different accessions. The variable bands were predominantly located at the terminal and subterminal chromosome regions, and were also unevenly distributed over chromosomes. In their work, the researchers present a generalized idiogram of Aegilops geniculata (Landjeva and Ganeva 2000).

Badaeva et al. (2002, 2004), using C-banding technique, analyzed heterochromatin banding patterns in the somatic metaphase chromosomes of several accessions of Ae. geniculata. Chromosome morphology and C-banding patterns in most studied accessions were similar to those reported previously (Pathak 1940; Chennaveeraiah 1960; Friebe et al. 1999), allowing chromosome designations according to the standard nomenclature (Friebe et al. 1999). Chromosome modifications were mostly found in Turkish accessions involving a high frequency of chromosomal rearrangements, represented by paracentric inversions and intragenomic and intergenomic translocations. Some modifications were the result of either Robertsonian translocations or translocations with interstitial breakpoints (Badaeva et al. (2002, 2004). This is in accord with the finding of Furuta (1981a), who reported a high frequency of chromosomal rearrangements in Turkish Ae. geniculata accessions. On the other hand, Yen and Kimber (1990c) did not observe modifications of the U genome chromosomes in five Turkish Ae. geniculata accessions.

The findings reported by Badaeva et al. (2002, 2004) confirmed that Ae. geniculata evolved as a result of hybridization between Ae. umbellulata and Ae. comosa. Comparing Ae. geniculata with its diploid ancestors revealed differences in morphology and C-banding patterns of many chromosomes belonging both to the M^o and U genomes, indicating that both genomes had been modified, which agrees with previous data (Kimber and Abu-Baker 1981; Kimber et al. 1988; Kimber and Yen 1989; Yen and Kimber 1990d).

Talbert et al. (1993) cloned three repetitive DNA sequences found primarily in the U genome and two repetitive DNA sequences found primarily in the M genome and used these to monitor variation in species containing both genomes. The U genome of Ae. umbellulata and the U subgenome of the allotetraploid U-genome species were similar with regards to hybridization patterns observed with the U genome probes. Much more variation was found both among diploid Ae. comosa accessions and allopolyploids containing M subgenomes. The observed variation supports the cytogenetic evidence that the M subgenome in the allotetraploid species is more variable than the U subgenome. It also raises the possibility that part of the differential nature of the M subgenome of the allotetraploids may be due to variation within the diploid Ae. comosa, while other variations may have been generated at the tetraploid level.

Badaeva et al. (2004) also studied the distribution of hybridization sites in Ae. geniculata, by performing FISH with the repetitive DNA probes pSc119 and pAs1, as well as the distribution of NOR and 5S DNA loci, using the pTa71 (18S-26S rDNA), and pTa794 (5S rDNA) probes. FISH with pSc119 revealed signals on 10 chromosome pairs, while five chromosome pairs had pAs1 FISH sites. Large- or medium-sized pSc119 FISH sites were observed in telomeric regions of either one or both chromosome arms. Interstitial pSc119 FISH sites were located in the long arms of 6U and 7U, which were identified on the basis of their similarity with the corresponding chromosomes of Ae. umbellulata (Badaeva et al. 1996a; Castilho and Heslop-Harrison 1995). On the other hand, the pAs1 FISH sites were faint and located in interstitial chromosome regions and, in general, resembled the pAs1–labeling pattern of Ae. comosa chromosomes (Badaeva et al. 1996a, 1999).

Badaeva et al. (2004) observed two pairs of SAT chromosomes in Ae. geniculata, confirming previous reports (Pathak 1940; Chennaveeraiah 1960; Cermeño et al. 1984b; Yamamoto 1992b; Yamamoto and Mukai 1995). Multicolor FISH with the probes pTa71 and pTa794 exposed the inactivation of major NORs on the M-subgenome chromosomes, and redistribution of 5S rDNA sites and loss of some minor 18S-26S rDNA loci in both subgenomes (Badaeva et al. (2004). Their study detected two major and four minor 18S-26S rDNA and three 5S rDNA sites, whose locations and presence were different from those in the parental species; chromosome 1U had a major NOR but lacked the 5S rDNA site and 1M^o only had the 5S rDNA locus. Small 5S rDNA loci were detected in the short arms of all group-5 chromosomes, which also had either major (5U) or minor (5M^o) 18S-26S rDNA loci. Minor pTa71 FISH sites of various intensities were detected in telomeric regions of two large sub-metacentric chromosome pairs, presumably 2 and 3 Mg, and in a small metacentric chromosome pair, presumably a derivative of chromosome 6 M of Ae. comosa that lost the NOR during the formation of Ae. geniculata. A weak hybridization signal was occasionally observed in an interstitial region of the long arm of chromosome 6U, which is also present in chromosome 6U of Ae. umbellulata (Badaeva et al. 1996b). The data presented by Badaeva et al. (2004) substantiated that Ae. geniculata speciation was accompanied by modification of both parental genomes as a result of amplification, deletion, and re-distribution of various classes of repetitive DNA sequences and chromosomal rearrangements.

Chromosome 1U is one of the two SAT chromosomes of diploid Ae. umbellulata and is one of the most conserved chromosomes among the different allopolyploid species containing the U subgenome. Loss of the 5S rDNA locus on 1U of Ae. geniculata was probably caused by a species-specific translocation. Badaeva et al. (2004) confirmed that the second subgenome of Ae. geniculata, the M subgenome, is a modified subgenome that underwent many modifications because of amplification, elimination and redistribution of highly repetitive DNA sequences, as well as chromosomal rearrangements. Part of the significant intraspecific heterogeneity observed in Ae. geniculata was presumably derived from independent hybridization and introgression with other species.

Cermeño et al. (1984b) analyzed the activity of the NORs in somatic metaphase chromosomes of Ae. geniculata using a highly reproducible silver-staining procedure. Similar to the finding of Badaeva et al. (2004), Cermeño et al. (1984b) found that the U subgenome completely suppressed the NOR activity of the M^o subgenome. They identified two pairs of SAT chromosomes and two pairs of Ag-NORs, although four Ag-NORs pairs were expected, based on their count in their diploid progenitors. The number of nucleoli at interphase was as follows: 22 cells had 1 nucleolus, 175 cells had 2 nucleoli, 99 had 3, and 15 had 4.

The production and identification of a complete set of an intact Ae. geniculata chromosome additions to common wheat has been described (Friebe et al. 1999). C-banding and meiotic pairing analyses revealed that all added Ae. geniculata chromosomes were structurally identical to the Ae. geniculata parent accession. Consequently, all U and M^o subgenome chromosomes were tentatively assigned to their homoeologous groups based on C-banding, first meiotic metaphase pairing analyses and plant morphologies.

FISH signals were generally associated with constitutive heterochromatin regions corresponding to C-band-positive chromatin, including telomeric, pericentromeric, centromeric and interstitial regions of all the 14 Ae. geniculata chromosome pairs. The newly identified satellite DNA CL36, used by Koo et al. (2016), produced localized M^o-subgenome chromosome-specific FISH signals in Ae. geniculata and in the M genome of Ae. comosa ssp. heldreichii var. subventricosa but not in Ae. comosa ssp. comosa, suggesting that the M^o subgenome of Ae. geniculata derived from var. subventricosa. Friebe et al. (1999) suggested a different source of the Ae. geniculata M^o subgenome, based on comparison with the C-banding patterns that were established for Ae. umbellulata and Ae. comosa by Badaeva et al. (1996a, b). They confirmed that the U genome of Ae. geniculata was contributed by Ae. umbellulata, but claimed that the M^o genome derived from Ae. comosa ssp. comosa.

9.8.3.3 Crosses with Other Species of the Wheat Group

Chromosomal pairing at first meiotic metaphase of an Israeli accession of Ae. geniculata was regular: 0.06 univalents, 1.20 rod bivalents, 12.77 ring bivalents and 30.66 chiasmata per cell (Feldman 1963). The single pair with a sub-terminal centromere, belonging to the U subgenome, appeared as rod bivalents but, in many cells, it had two chiasmata. A higher number of rod bivalents (3.01 rod bivalents, 10.99 ring bivalents) and lower number of chiasmata per cell at first meiotic metaphase (24.99) was reported by Cuñado (1992) for Ae. geniculata from Turkey. Similar pattern of chromosomal pairing at first metaphase of meiosis of Ae. geniculata was reported by Cuñado et al. (1996a, b), who also analyzed the pattern of pachytene pairing by whole-mount surface-spreading of synaptonemal complexes under the electron microscope. Their data indicated that in more than 90% of the cells at the pachytene stage, all the chromosomes were associated as bivalents, while in the rest of the cells, they observed 12 bivalents and one quadrivalent, presumably the result of inter-subgenomic homoeologous pairing.

Furuta (1981a) studied meiosis in intraspecific hybrids of Ae. geniculata and found that many Turkish accessions differed by one or two reciprocal translocations. On the other hand, Yen and Kimber (1990c) did not observe any rearrangement in the U genome chromosomes in five Turkish accessions of Ae. geniculata.

Data on chromosomal pairing in F₁ hybrids between Ae. geniculata and its diploid parents, Ae. umbellulata and Ae. comosa, are presented in Table 9.5. Kihara (1937) reported that the Ae. geniculata x Ae. umbellulata hybrid had 7 bivalents, and similar observation was made by Kimber and Abu-Baker (1981) observed 6.42 univalents, 5.69 bivalents, 0.92 trivalents and 0.11 quadrivalents. The Ae. geniculata x Ae. comosa hybrid had 6.15 univalents, 2.65 rod bivalents, 3.05 ring bivalents and 1.15 trivalents (Kimber et al. 1988). The triploid hybrids pairing data substantiated the Kihara’s (1937, 1949, 1954) claim that Ae. geniculata originated from a hybridization between Ae. comosa and Ae. umbellulata, followed by chromosome doubling. Yet, the meiotic analysis of Kimber and Yen (1988) showed that the M genome of Ae. geniculata had undergone substantial modification; the U subgenome is much closer to the U genome of Ae. umbellulata than the M^o subgenome is to the M genome of Ae. comosa. However, the possibility exists that the U subgenome has also been somewhat modified (Kimber and Yen 1988).

Meiotic data from other F₁ triploid hybrid involving Ae. geniculata and Ae. uniaristata (Table 9.5) displayed low chromosomal pairing, indicating that there is low homology between the subgenomes of Ae. geniculata and the genomes of Ae. uniaristata (Table 9.5). The triploid hybrid Ae. geniculata x Ae. longissima had 2–6 bivalents, with mode of 4, and 0–1 trivalents (Kihara 1949). The Ae. geniculata x T. monococcum hybrid had 13.50 univalents, 3.42 rod bivalents, 0.06 ring bivalents and 0.18 trivalents (Bell and Sachs 1953). A similarly low level of pairing [0–5 (usual 1–3) bivalents] were reported earlier in this hybrid combination by Aase (1930), Bleier (1930 and Percival (1930).

Chromosome pairing at meiosis of F₁ hybrids between Ae. geniculata and allotetraploids sharing the U subgenome but differing in the second subgenome, showed that, in addition to pairing between the chromosomes of the shared subgenome, that pairing also occurred between several chromosomes of the non-shared subgenomes (Table 9.7). For instance, the F₁ tetraploid hybrid Ae. geniculata x Ae. biuncialis (hybrid genome M^oM^bUU) had 10.25 univalents, 4.20 rod bivalents, 2.30 ring bivalents, 1.25 trivalents and 0.25 quadrivalents (Kimber et al. 1988). The Ae. geniculata x Ae. columnaris (hybrid genome M^oXⁿUU) hybrid had 9.20 univalents, 4.45 rod bivalents, 2.70 ring bivalents, 1.10 trivalents and 0.30 quadrivalents (Kimber et al. 1988). The Ae. geniculata x Ae. kotschyi (hybrid genome M^oS^vUU) hybrid had 11.70 univalents, 1.60 rod bivalents, 4.42 ring bivalents and 1.42 trivalents (Feldman 1963), and Kimber et al. (1988) observed 10.65 univalents, 3.20 rod bivalents, 3.25 ring bivalents, 1.35 trivalents and 0.10 quadrivalents in this hybrid combination, and 11.95 univalents, 3.50 rod bivalents, 2.15 ring bivalents, 1.25 trivalents and 0.25 quadrivalents in the reciprocal hybrid. The Ae. geniculata x Ae. peregrina hybrid had 11.44 univalents, 2.02 rod bivalents, 5.00 ring bivalents and 0.84 trivalents (Feldman 1963). The Ae. geniculata x Ae. triuncialis hybrid had 5–11 (usually 7–8) bivalents (Kihara 1929).

Hybrids between tetraploid species with different genomic constitution had significantly less pairing than hybrids between tetraploid species that share one subgenome. For example, the Ae. cylindrica x Ae. geniculata (hybrid genome DCM^oU) hybrid had 3–8 bivalents (Aase 1930) and the Ae. geniculata x Ae. ventricosa (hybrid genome M^oUDN) hybrid had 3–10 bivalents (usually 7) (Kihara 1929) or 3–7 bivalents (all rod bivalents) (Percival 1930).

Chromosomal pairing was also studied in hybrids between Ae. geniculata and tetraploid and hexaploid wheat (Tables 9.9 and 9.10). The tetraploid hybrid Ae. geniculata x T. turgidum sp. turgidum (hybrid genome M^oUBA) had 0–3 rod bivalents (Aase 1930; Bleier 1928; Gaines and Aase 1926 Yen and Kimber (1990d) Genomic relationships of Triticum searsii to other S-genome diploid Triticum species. Genome 33: 369–374.

Kagawa 1929; Kihara 1929; Percival 1930; Sax and Sax 1924), and the Ae. geniculata x T. turgidum subsp. durum (genome M^oUBA) hybrid had 0–4 rod bivalents (Aase 1930). The T. aestivum x Ae. geniculata (genome BADM^oU) hybrid had 29.67 univalents, 2.62 bivalents and 0.03 trivalents (Riley 1996a), or 0–3 rod bivalents (Bleier 1928), indicating very low homology between the subgenomes of these two species. Similarly, low levels of pairing in the T. aestivum x Ae. geniculata hybrid was observed by Cifuentes and Benavente (2009), who examined chromosome pairing at meiotic metaphase in this F₁ pentaploid hybrid by FISH, which enabled simultaneous discrimination of the chromosomes of the A, B, D, U and M^o subgenomes. They observed 32.86 univalents, 1.04 bivalents, and 0.02 trivalents, of which more than 60% represented allosyndetic pairing between wheat and geniculata chromosomes. The average ratio of aestivum-geniculata associations was 5:1:12 for those involving the A, B and D subgenomes, respectively, indicating that somewhat higher homology exists between D subgenome chromosomes and those of Ae. geniculata than between A or B subgenomes chromosomes and Ae. geniculata chromosomes.

Fernandez-Calvin and Orellana (1992) analyzed meiotic associations at first metaphase in the pentaploid hybrid Ae. geniculata x T. aestivum, bearing the Ph1 gene (suppressor of homoeologous pairing) and the mutant phlb (a deficiency of Ph1 that enables high homoeologous pairing), using the C-banding technique. The observed associations revealed the same relative order: AD-M^oU > A-D > U–M^o > AD-B > UM^o-B in both low- and high-homoeologous-pairing hybrids.

Benavente et al. (2001) analyzed karyotypes of offspring of two different T. turgidum–Ae. geniculata amphiploids (2n = 8x = 56; genome BBAAM^oM^oUU) carrying Ph1 or lacking it (ph1c deletion), by GISH. The offspring, obtained after two generations of amphiploid selfing, had, on average, fewer chromosomes than expected. Most of the lost chromosomes belonged to Ae. geniculata. The two families differed greatly in the number of intergenomic translocations. The ph1c family showed nine translocations over 12 plants, while only one translocation was observed in the Ph1 family. All exchanges involved either the M^o and U chromosomes or the M^o and wheat chromosomes. The results suggest an epistatic effect of the ph1c deletion on the genetic diploidizing system that operates in Ae. geniculata, since translocated chromosomes are most likely derived from homoeologous recombination.

In summary, Ae. geniculata has very little homology with the subgenomes of either tetraploid or hexaploid wheat, as expressed by the minimal pairing in its hybrids with wheat species of these ploidy levels. Aase (1930) suggested that Ae. geniculata may be an autotetraploid. But, if so, more bivalents would be expected in its hybrids both with tetraploid and hexaploid wheat, and Kagawa’s studies (1929) on chromosome morphology in Ae. geniculata unambiguously rule out an autotetraploid interpretation of Ae. geniculata.

9.8.4 Aegilops biuncialis Vis.

9.8.4.1 Morphological and Geographical Notes

Ae. biuncialis Vis., common name Mediterranean Aegilops, [Syn.: Ae. lorentii Hochst.; Ae. machrochaeta Shuttl. & Huet. ex Duval-Jouve; T. macrochaetum (Shuttl. & Huet. ex Duval-Jouve) Richter; T. biuncialis Vill.; Ae. connata Ateud.] is a predominantly autogamous, annual, multi-tillered plant, 15–40(-50) cm tall (excluding spikes). The uppermost 1/3 or 1/4 of the culms is defoliated. Leaf blades are glabrous or ciliate, seldom hairy, short (2–5-cm-long) and narrow-linear. Its spike is narrow-lanceolate to narrow-elliptical, lax, 2.0–3.5-cm-long (excluding awns), awned, usually with two spikelets, rarely 3–4, the uppermost of which is not significantly smaller than the lower ones. The entire spike disarticulates at maturity and falls as a unit (umbrella-type dispersal unit). The lower rachis internode is shorter than the adjacent spikelet. There is one basal rudimentary spikelet, seldom two. The spikelets are narrow to broad elliptical, and their lower parts are sometimes slightly inflated. There are 4–5 florets, the two lower one being fertile. The glumes are 8–10-mm-long, awned, with unequally broad nerves that are unequally spaced. Glume awns are usually smooth underneath, all with the same breadth, but with unequal lengths in different spikelets. Terminal spikelet have three glume awns, which are 4–7-cm-long, and much longer and broader than those of the lateral spikelets. The central awn of the terminal spikelet is sometimes longer than its lateral awns. There are 2–3 awns on the lateral spikelets, and when there are three, the central awn is shorter than the lateral awns. The lemma is membranous, usually awned, but the awns are more poorly developed than the glume awns, plainly shorter and fewer, but always more than one per spikelet, usually 2 in lateral spikelets, and 3–4 in the terminal ones. All awns diverge at maturity. The caryopsis is free (Fig. 9.4c).

There is a difference of opinion among taxonomists concerning the valid name of this species. The name Ae. biuncialis was given by Visiani (1842), accompanied by an illustration but without a description. His description of the species appeared only ten years later (Visiani 1852). In the meantime, Hochstetter (1845) described the same species, collected by Lorent (1845), and named it Ae. lorentii. Zhukovsky (1928), Eig (1929a), Nevski (1934a, b), Tzvelev (1976) and Gandilian (1980) used the name Ae. biuncialis, whereas other taxonomists, e.g., Bor (1968, 1970), Tutin and Humphries (1980) and Hammer (1980), used the name Ae. lorentii. Yet, according to Article 44 of the International Code of the Botanical Nomenclature, the former name, which was published earlier and accompanied by an illustration with analysis, is the valid name; and so, it is, in practice (Mattatia and Feinbrun-Dothan 1986).

Ae. biuncialis exhibits a very wide morphological variation mainly involving the spike shape (elliptical to lanceolate), size (2 or 3, seldom 4 fertile spikelets), color, hairiness and awn width and length. This variation is reflected in the adaptation of Ae. biuncialis to different environmental conditions throughout its distribution area (van Slageren 1994). Because of this morphological variation, Zhukovsky (1928), Eig (1929a), and Hammer (1980) subdivided Ae. biuncialis into three varieties that mainly differed in the shape of the spikelets, hairiness and length of glumes and awns.

A high degree of intraspecific molecular variation was also revealed in Ae. biuncialis; analysis of several populations of this species from the Iberian-peninsula and the Balearic-islands, using AFLP DNA markers, showed high genetic variation within and between the studied populations (Monte et al. 1999, 2001). Similarly, Thomas and Bebell (2010), using RAPD and ISSR markers, revealed a significant amount of genetic variability in Ae. biuncialis from Greece. Rabokon et al. (2019), using intron-specific DNA polymorphism of β-tubulin gene family members as a molecular marker, uncovered DNA polymorphism sufficiently high to distinguish between different accessions of Ae. biuncialis.

Ae. biuncialis distributes in the Mediterranean and western Asiatic regions. It grows in Portugal (possibly), Spain, South France (including Corsica), Italy (including Sicily and Sardinia), Malta, Greece (including Crete and the Aegean Islands), Albania, Macedonia, Serbia, Bosnia-Herzegovina, Croatia, Romania, Bulgaria, South Ukraine, South Russia (Crimea and Cis-Caucasia), Trans-Caucasia (Armenia, Azerbaijan, Georgia), Turkey, northern Iraq, western Iran, Syria, Lebanon, Cyprus, Israel, Jordan, Libya, Tunisia, Algeria, and Morocco. Ae. biuncialis is common in southern Europe and the Aegean, Greece, Turkey, Bulgaria and Cyprus. It is well represented in the western arc of the Fertile Crescent but virtually rare in the central part and the eastern arc, in Cis- and Trans- Caucasia, southern Crimea and southern Ukraine. It is less common in North Africa. Ae. biuncialis is adventive in parts of central and northwestern Europe such as Germany, Switzerland and the Netherlands. Ae. biuncialis grows on a variety of soils, e.g., terra rossa, basalt, and rendzina soils. It is prevalent in the edges and openings of the sclerophyllous and deciduous oak forests and maquis, in degraded shrub formations, in semi-steppe herbaceous formations, in stony hillsides, abandoned fields, edges and within cultivation such as olive groves, vineyards, fruit tree plantations, and within or near barley and wheat fields, in disturbed and eroded areas and roadsides. It is common and locally abundant in generally dry, somewhat disturbed habitats, such as fallow wastelands, roadsides, and dry rocky slopes of hills and mountains. It grows at altitudes of 200–1750 m, rarely up to 2100 m in Lebanon. As a typical colonizer, the species can form massive stands, especially in regularly disturbed places. It grows in areas with annual rainfall varying from less than 100 mm up to 1100 mm, with most growing in areas with a range of 200–700 mm rain.

The distribution area of Ae. biuncialis is in the western and central parts of the distribution of the genus. It is a Mediterranean element extending into semi-steppical (West Irano-Turanian) region, occupying a wide variety of primary and secondary habitats. Ae. biuncialis usually grows in mixed stands with other Aegilops species, particularly those belonging to the U-genome cluster, i.e., the allopolyploids of section Aegilops, with which it may introgress. Interspecific hybrids and hybrid derivatives were frequently found in mixed populations of Ae. biuncialis, Ae. geniculata, and Ae. peregrina in Israel, indicating continuous genetic connections between these three species (Zohary and Feldman 1962; Feldman 1965a, b, c). Hybrids between Ae. biuncialis and common wheat, when the wild species grew in or close to the wheat fields, were reported by Loureiro et al. (2006), Loureiro and Escorial (2007), who estimated the field hybridization rate under central Spain conditions. Their study showed that the hybrid Ae. biuncialis x common wheat can be partially fertile by pollinating it by the wheat parent and thus, can introgress with the domesticated parent.

The distribution of the two diploid parents of Ae. biuncialis overlaps in western Turkey and Greece. It is assumed therefore, that this area is the center of origin of Ae. biuncialis. The distribution of Ae. biuncialis overlaps with and is much larger than that of its two parents. It has a sympatric distribution with the following species: A. muticum, Ae. speltoides, Ae. searsii, Ae, caudata, Ae. comosa, Ae. uniaristata, Ae. umbellulata, wild T. monococcum, wild T. timopheevii, wild T. turgidum, Ae. geniculata, Ae. neglecta, Ae. recta, Ae. columnaris and Ae. ventricosa, and an allopatric distribution with Ae. longissima, Ae. kotschyi, Ae. tauschii, Ae. crassa, Ae. vavilovii and Ae. juvenalis.

Ae. biuncialis contains genes that confer resistance to powdery mildew (Gill et al. 1985; Zhou et al. 2014), to leaf rust (Gill et al. 1985; Marais et al. 2003), yellow (stripe) rust (Damania and Pecetti 1990; Marais et al. 2003; Zhou et al. 2014), barley yellow dwarf virus (BYDV) (Makkouk et al. 1994), and Hessian Fly (Gill et al. 1985). It also contains genes that may improve tolerance to abiotic stresses such as drought and salt (Molnár et al. 2004; Colmer et al. 2006; Dulai et al. 2014). In this regard, Molnáret al. (2004) found that Ae. biuncialis genotypes originating from a dry habitat have better drought tolerance than wheat, rendering them good candidates for improving the drought tolerance of wheat through intergeneric crossing. In addition, it was found that one accession of Ae. biuncialis had a high gluten and grind quality because of a high percentage of γ-45.31 and γ-43.5 gliadins, while another accession had low gluten content, because of a low percentage of these gliadins (Ahmadpoor et al. 2014). A disomic addition line of a pair of chromosomes 1U of Ae. biuncialis in the background of common wheat, improved the end-product quality of wheat (Zhou et al. 2014). Also, chromosome 3 MB of Ae. biuncialis improved the grain micronutrient content, namely, higher K, Zn, Fe, and Mn contents in wheat (Farkas et al. 2014).

9.8.4.2 Cytology, Cytogenetics and Evolution

Ae. biuncialis is an allotetraploid (2n = 4x = 28) species. Its nuclear genome designation is UUM^bM^b and that of its organellar genome is U. Kihara (1937, 1954) designated the genome of Ae. biuncialis C^uC^uM^oM^o (current designation C^u = U and M^o = M^b), and Lilienfeld (1951) concluded that the M^o subgenome of Ae. biuncialis is a modified M genome, that was derived from either the M genomes of Ae. comosa or from the M^u (currently N) genome of Ae. uniaristata. The designation C^uC^uM^oM^o was based, in addition to genome analyses pairing data, on morphological characteristics and on karyotype (Senyaninova-Korchagina 1932). However, a later karyotypic study showed that one chromosome set corresponded very well with the chromosomes of Ae. umbellulata, whereas the second chromosome set did not correspond to any chromosomes of the diploid species (Chennaveeraiah 1960).

Sasanuma et al. (2006) presented evidence that the U subgenome of Ae. biuncialis had multiple origins deriving from different accessions of Ae. umbellulata. This conclusion was based on PCR–RFLP of the U genome-specific U31 fragment, developed by Kadosumi et al. (2005). Sasanuma et al. (2006) investigated the PCR–RFLP of this fragment in 48 accessions of Ae. biuncialis and found that most accessions possessed one allele of this fragment, whereas other accessions had a second allele. Since these two alleles exist in different accessions of Ae. umbellulata, Sasanuma et al. (2006) concluded that the U subgenome in Ae. biuncialis had multiple origins. The multiple origin of the M subgenome of Ae. biuncialis has been suggested by Chee et al. (1995),

The origins of the subgenomes of Ae. biuncialis and Ae. geniculata were investigated by examining the presence of specific restriction fragments of repeated nucleotide sequences in the DNA of the allopolyploid species (Resta et al. 1996). The analysis showed that Ae. biuncialis and Ae. geniculata are closely related, that the U subgenome of both species is closely related to the genome of Ae. umbellulata, whereas the second subgenome, M^b in Ae. biuncialis and M^o in Ae. geniculata, is a modified genome of Ae. comosa. Modification of the M^b and M^o subgenomes could be attributed to hybridization of allotetraploids sharing the U subgenome but differing in their second subgenome (Zohary and Feldman 1962; Feldman 1965a, b, c). C-banding and FISH studies also confirmed that the U subgenome of Ae. biuncialis derived from Ae. umbellulata and that the second subgenome, M^b, is a modified M genome of Ae. comosa (Badaeva et al. 2004). Like Resta et al (1996), Badaeva et al. (2004) also assumed that intraspecific divergence of Ae. biuncialis involved introgression of genetic material from other species. In addition to sharing the U and M subgenomes with the allotetraploid species Ae. geniculata, Ae. biuncialis shares the U subgenome with the allopolyploids Ae. columnaris (UUXⁿXⁿ), and Ae. neglecta (UUXⁿXⁿ), but differ from them in the second subgenome (Resta et al. 1996). In accord with this, Badaeva et al. (2004) found significant differences in the karyotype structure, in the total amount and distribution of C-heterochromatin and in the number and location of 5S and 18S-26S rDNA loci between Ae. geniculata and Ae. biuncialis on the one hand, and between Ae. columnaris and Ae. neglecta, on the one hand, evidence that these two groups of species contain the U genome of Ae. umbellulata but differ in the source of the second subgenome.

Several investigations were performed to identify the cytoplasm donor of Ae. biuncialis. Using restriction endonucleases, Ogihara and Tsunewaki (1982) found no differences in the chloroplast genome of two Ae. biuncialis accessions. Terachi et al. (1984) and Ogihara and Tsunewaki (1988) discovered that the chloroplast genome of Ae. biuncialis is almost identical with that of Ae. umbellulata and therefore concluded that Ae. umbellulata was the cytoplasm donor to Ae. biuncialis. Consequently, the designation U was given to the plasma type of Ae. biuncialis. These results were also confirmed by further studies on the chloroplast and mitochondrial DNA (Wang et al. 1997; Tsunewaki 2009). In contrast, based on studies of cytoplasm-substituted wheats, Maan (1975, 1978) and Panayotov and Gotsov (1975, 1976) proposed that Ae. uniaristata or Ae. comosa were the cytoplasm donors of Ae. biuncialis. However, the results of Terachi et al. (1984), Ogihara and Tsunewaki (1988), Wang et al. (1997), and Tsunewaki (2009) clearly confirmed the conclusion that Ae. umbellulata was the cytoplasm donor to Ae. biuncialis. Based on plasmon differences between the cytoplasms of the allotetraploids and those of Ae. umbellulata, Terachi et al. (1984), Wang et al. (1997), and Tsunewaki (2009) proposed that Ae. biuncialis is a young species whose origin was later than that of the origin of Ae. geniculata.

Ae. biuncialis contains 10.37 ± 0.039 pg 1C DNA (Eilam et al. 2008), which is 4.95% less DNA than that which is expected from the sum of the DNA of its two diploid parents, i.e., 10.91 pg (Ae. umbellulata contains 5.38 pg and Ae. comosa contains 5.53 pg; Eilam et al. 2007). The loss of DNA in Ae. biuncialis was also found by Badaeva et al. (2002, 2004) who, based on differential C-banding and FISH, found that Ae. biuncialis exhibits substantial structural chromosome rearrangements, including deletion of chromosomal segments and reduction of heterochromatin content.

Senyaninova-Korchagina (1932) was the first to describe the karyotype of Ae. biuncialis. She recognized seven types of chromosomes that have only primary constrictions and observed no SAT chromosomes. In contrast, Chennaveeraiah (1960) reported on three chromosome pairs with satellites on short arms, two pairs with markedly larger satellites, whereas those on the third pair were very small. All in all, he recognized 12 types of chromosomes, where one pair had an extreme subterminal centromere, three pairs had submedian-subterminal centromeres, and the rest of the pairs had submedian centromeres. No chromosome had a median centromere.

Teoh et al. (1983), using FISH with a repetitive clone that codes for rRNA, confirmed that there are three chromosome pairs in Ae. biuncialis exhibiting rRNA sites, whereas the expected number of pairs of rRNA is 4. Cermeño et al. (1984b) analyzed nucleolar activity in Ae. biuncialis using a highly reproducible silver-staining procedure and found that the U subgenome suppressed one pair of the NORs of the M^b subgenome.

Badaeva et al. (2002, 2004) studied the karyotype structure of Ae. biuncialis by analyzing heterochromatin banding patterns of their somatic metaphase chromosomes, as revealed by C-banding and FISH with the heterochromatin-limited repetitive DNA probes pSc119, pAs1, as well as the distribution of NOR and 5S DNA loci, revealed by pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) probes. The C-banding studies displayed a wide polymorphism resulting from chromosomal rearrangements represented by paracentric inversions and intra- and inter-subgenomic translocations. The results obtained confirmed that the allotetraploid species Ae. biuncialis was formed as a result of hybridization of the diploids Ae. umbellulata and Ae. comosa. The dissimilarity of the C-banding patterns and FISH sites in several chromosomes of this allotetraploid species and those of its ancestral diploid species indicated that chromosomal changes occurred at the tetraploid level. Badaeva et al. (2004) found that Ae. geniculata and Ae. biuncialis differed from each other and from the putative diploid progenitors in the inactivation of major NORs on the M-subgenome chromosomes, in the redistribution of 5S rDNA sites, and loss of some minor 18S-26S rDNA loci in Ae. geniculata and Ae. biuncialis. These differences indicate that various types of chromosomal alterations occurred during the formation and evolution of these allopolyploid species.

When analyzing the karyotype of three different accessions of Ae. biuncialis, Wang et al. (2013b) found that they all exhibited a similar karyotype. All Ae. biuncialis chromosomes had identifiable C-bands and FISH sites, which allowed for simultaneous discrimination of all U and M^b chromosomes. The U subgenome of Ae. biuncialis resembled the U genome of the diploid species Ae. umbellulata, whereas the M^b subgenome had some differences compared to the M genome of Ae. comosa. The C-banding pattern of the three studied accessions was similar to that reported by Badaeva et al. (2004).

In contrast to Wang et al. (2013a), Schneider et al. (2005), using FISH with the two repetitive DNA sequences pSc119.2 and pAs1 on root-tip metaphases, observed differences in the FISH patterns of all chromosomes among four Ae. umbellulata accessions, four Ae. comosa accessions, and three Ae. biuncialis accessions. The hybridization patterns of the analyzed Ae. biuncialis accessions were more variable, and differences were observed not only among the Ae. biuncialis lines but also between Ae. biuncialis and its diploid progenitors. The genetic variability of Ae. biuncialis was manifested by the different locations of the repetitive sequences, which led to differences in the FISH pattern. FISH polymorphism was detected in both the U and M subgenome chromosomes, but, as also reported by Chee et al (1995), the level of repetitive DNA variation in the M subgenome was much higher than in the U subgenome. Similar results were obtained by Gong et al. (2006) who, using ISSR markers, found that the U-subgenome of the allopolyploids of the U subgenome group, was very similar to that of Ae. umbellulata and was practically unchanged, while the other subgenomes were greatly altered in the allopolyploids, as was suggested by Zohary and Feldman (1962). Likewise, Kimber and Yen (1988), analyzing chromosome pairing in the hybrid between Ae. biuncialis and autotetraploid Ae. umbellulata, found that the U subgenome was relatively unmodified.

In addition, Schneider et al. (2005), produced and identified five different T. aestivum–Ae. biuncialis disomic addition lines. To differentiate between the added Ae. biuncialis chromosomes and those of common wheat, they used genomic in situ hybridization and detected no chromosome interchanges involving wheat and Ae. biuncialis chromosomes. Schneider et al. (2005) used three repetitive DNA clones (pSc119.2, pAs1, and pTa71) and identified the Ae. biuncialis disomic additions as 2M, 3M, 7M, and 3U.

9.8.4.3 Crosses with Other Species of the Wheat Group

Feldman (1963) studied chromosomal pairing at first meiotic metaphase of two Ae. biuncialis accessions and found regular behavior, i.e., an average of 0.97 rod bivalents and 13.03 ring bivalents per cell. Cuñado (1992) observed more rod bivalents These authors also analyzed the pattern of early meiotic zygotene pairing in Ae. biuncialis by whole-mount surface-spreading of synaptonemal complexes under the electron microscope. Their data indicate that at the zygotene stage, almost all cells (93%) had 14 bivalents, whereas only a few cells (7%) had 12 bivalents and 1 quadrivalent.

Chromosomal pairing at first meiotic metaphase of F₁ hybrids between Ae. biuncialis and its putative diploid parents, Ae. umbellulata and Ae. comosa, was studied by Kihara (1937, 1949). The Ae. biuncialis x Ae. umbellulata hybrid had an average of 7 bivalents (Kihara 1937) and the Ae. biuncialis x Ae. comosa hybrid had 5–8 bivalents with a mode of 7 and 0–2 trivalents (Kihara 1949). Kihara (1949) also studied hybrids between Ae. biuncialis and diploid species of section Sitopsis and noted much less pairing than in the hybrids between Ae. biuncialis and its putative diploid parents. Ae. biuncialis x Ae. speltoides had 3–7 bivalents and 0–3 trivalents, Ae. biuncialis x Ae. longissima had 0–5 bivalents and 0–2 trivalents, and the hybrid Ae. biuncialis x Ae. bicornis had 0–6 bivalents and 0–1 trivalents. On the other hand, the F₁ hybrid Ae. biuncialis x Ae. caudata had 5–7 bivalents with a mode of 7, and 0–3 trivalents (Kihara 1949), indicating a greater homology with Ae. caudata than with the Sitopsis species.

Hybrids between Ae. biuncialis and allopolyploid species sharing the U subgenome were studied by Kihara (1937), Feldman (1965c), and Kimber et al. (1988). Kihara observed 7 bivalents in the F₁ hybrid Ae. biuncialis x Ae. peregrina (hybrid genome UM^bS^vU) while Feldman reported 12.16 univalents, 1.16 rod bivalents, 5.38 ring bivalents, 0.76 trivalents, and 0.12 quadrivalents in this hybrid. The range of the bivalents in this hybrid was 5–9, indicating that in addition to the 7 bivalents between the chromosomes of the U subgenomes, some bivalents were also formed between the chromosomes of the differential genomes M^b and S^v. The F₁ Ae. geniculata x Ae. biuncialis hybrid (hybrid genome M^oUUM^b) had 10.24 univalents, 4.20 rod bivalents, 3.39 ring bivalents, 1.25 trivalents and 0.25 quadrivalents (Kimber et al. 1988). This hybrid also had pairing between chromosomes of the differential subgenomes M^o and M^b. Lindschau and Oehler (1936) observed very low chromosome pairing in the F₁ hybrid Ae. biuncialis x Ae. cylindrica (hybrid genome UM^bDC), indicating that Ae. biuncialis has no subgenome in common with Ae. cylindrica.

9.8.5 Aegilops neglecta Req. ex Bertol.

9.8.5.1 Morphological and Geographical Notes

Ae. neglecta Req. ex Bertol., common name tri-awn goatgrass, [Syn.: Ae. ovata L. emend. Roth; T. ovatum (L.) Raspail; Ae. triaristata Willd.; T. triaristatum Willd.) Godr. & Gren. in Grenier & Godron; T. neclectum (req. ex Bertol.) Greuter in Greuter & Rechinger] is a predominantly autogamous, annual, tufted, multi-tillered, 20–40(-50)-cm-tall (excluding spikes) plant. Its culms have few joints, are upright, or somewhat jointed near the ground, with the uppermost 1/4 or 1/3 being defoliated. Leaves are more or less hairy, often with ciliate margins. Its spike is 2.0–4.5-cm-long (excluding awns), lanceolate or narrowly ovoid and compact, with the lower parts inflated and ellipsoid, becoming suddenly narrow-cylindrical in the upper part, and awned. The entire spike disarticulates at maturity and falls as a single unit (umbrella-type dispersal unit). There are 3 basal rudimentary spikelets, seldom 2. There are 3–6 spikelets, usually 4, with the two lowest being narrow to broad-elliptical, proportionally large and lying against each other, and the upper two being narrow, sterile, and projecting conspicuously from the lower spikelets. The lower rachis internodes are shorter than the adjacent spikelet, whereas the upper rachis internodes are longer than the spikelets. There are 4 florets, the upper 2 being sterile. The glumes have curved, unequally broad, flattened nerves, with 2–3 awns that are equal in length to those of the lemma. There are usually three, 3.5–4.5-cm-long glume awns 2–3 on the lower spikelets and almost always 3 on the upper spikelets. There are 2–4 (usually 2) lemma awns on lower spikelets, usually none on the upper ones, and if they exist, they are weakly developed. Often the awns in the upper spikelets decrease in length so that all awns of the spike end at the same height. The awns of the mature spike are generally weak, seldom strong. The caryopsis is free (Fig. 9.4d).

Ae. ovata (senso lato) was presented in the species Plantarum (Linnaeus 1753) as containing two taxa, ovata (senso stricto) and triaristata. The taxon triaristata was separated from ovata and elevated to the specific rank by von Willdenow in (1806), but the name Ae. triaristata was illegitimate since it was already given to a part of the original Ae. ovata. Ae. neglecta Req. ex Bertol. was described by Bertoloni in 1834. Since it is the oldest available name to replace the triaristata-part of the old ovata of Linneaus, it is the legitimate name for this species (van Slageren 1994). Yet, a number of taxonomists and cytogeneticists [e.g., Zhukovsky (1928), Eig (1929a), Chennaveeraiah (1960), Kihara (1940a, b, 1954, 1963), Kihara and Tanaka (1970)] used the name triaristata, while others, e.g., Hammer (1980), used the name neglecta.

Ae. triaristata included tetraploid and hexaploid forms. The two forms were considered by Zhukovsky (1928), and Kihara 1954, 1963), as subspecies of Ae. triaristata, whereas Hammer (1980) considered them subspecies of Ae. neglecta. Hammer (1980) and van Slageren (1994) designated the tetraploid forms as ssp. neglecta and the hexaploid ones as ssp recta. Chennaveeraiah (1960), based on karyotype differences, elevated the hexaploid forms to the species rank and named it Ae. recta (Zhuk.) Chennav., but kept the tetraploid as Ae. triaristata.

Chennaveeraiah (1960) described few morphological differences between Ae. neglecta and Ae. recta, such as in the number of spikelets, with 2 to 3 in neglecta and 5 to 6 in recta. The awns in neglecta are spread and in recta, are slanted. Neglecta glumes are glabrous and in recta are hairy. In addition, Kihara (1963) noted that neglecta spikes suddenly taper, whereas those of recta, gradually taper. Yet, there are many morphological intermediates between the two species and sometimes it is difficult to differentiate between them (Kihara 1963).

Ae. neglecta has a wide morphological variation mainly in spikelet number, shape and size, hairiness, awn number and development. It is sometimes confused with its closest relatives Ae. recta and Ae. columnaris. Thomas and Bebell (2010), using RAPDs and ISSRs, reported on the occurrence of wide molecular diversity in Greek Ae. neglecta.

Ae. neglecta is an East Mediterranean and West-Asiatic element. It distributes in the northeastern Mediterranean region, that is in western Turkey, Greece (including the Aegean Islands and Crete), the Balkan (Croatia, Bosnia-Herzegovina, Serbia-Montenegro, North Macedonia, and Albania), Southern Crimea, and Syria, and in West Asia, i.e., northern Iraq, Cis-Caucasus, Trans-Caucasus, Northern Iran (rare) and Turkmenistan (Kihara 1963; van Slageren 1994; Ohta et al. 2016). Although Ae. neglecta was described from France and other West Mediterranean countries, there are no chromosome counts to assure that all these collections are tetraploids.

Ae. neglecta grows on a variety of soils, e.g., terra rossa, basalt, rendzina, and alluvial soils. It is common in the edges and openings of sclerophyllous and deciduous oak forests and maquis, in openings of shrub formations, degraded dwarf-shrub formations, semi-steppe herbaceous formations, pastures, stony hillsides, abandoned fields, edges of cultivation, disturbed and eroded areas and roadsides, within cultivations, such as olive groves, vineyards, fruit tree plantations, and within or near barley and wheat fields. It is also common, locally abundant in the Eastern Mediterranean but somewhat less common in its eastern distribution area. It grows at altitudes from almost sea level to 1600 m, rarely, up to 2000 m. As a typical colonizer, the species can form massive stands, especially in regularly disturbed places. It grows in areas with annual rainfall varying from less than 450 mm to 750 mm.

Ae. neglecta is an adventive in central and northwestern Europe, e.g., in Scotland, Belgium, Germany, Switzerland and in the Netherlands. It was introduced to the USA and reported as a “weed in fields of California and Virginia” by Hitchcock and Chase in their 1951 Manual of the Grasses of the United States. Ae. neglecta is still very restricted in North America, known in California, Oregon, New York and Virginia.

Ae. neglecta has a wide distribution in the central region of the genus distribution. It occupies a large variety of primary and secondary habitats. Ae. neglecta grows sympatrically with Ae. caudata, Ae. comosa, Ae. uniaristata, Ae. umbellulata, Ae. biuncialis, Ae. triuncialis, Ae. cylindrica and Ae. recta and allopatrically with A. muticum, Ae. speltoides, Ae. peregrina and Ae. columnaris. Hybrids and intermediate forms between Ae. neglecta and other species of the U-subgenome group, which usually grow in mixed stands, are occasionally found (Feldman 1965a).

Ae. neglecta contains genes that confer resistance to powdery mildew (Gill et al. 1985; Worthington et al. 2015), leaf rust (Gill et al. 1985; Marais et al. 2009), yellow (stripe) rust (Marais et al. 2009), barley yellow dwarf virus (BYDV) (Makkouk et al. 1994), and to Hessian Fly (Gill et al. 1985; El Bouhssini et al. 1998). Damania and Pecetti (1990) reported that Ae. neglecta is tolerant to drought and frost.

9.8.5.2 Cytology, Cytogenetics, and Evolution

Ae. neglecta is an allotetraploid (2n = 4x = 28), with a UUXⁿXⁿ genome designation (modified by Badaeva et al. 2008 from Dvorak 1998; Tsunewaki 2009). The designation of its plasmon is U, similar to that of Ae. umbellulata (Kimber and Tsunewaki 1988; Dvorak 1998; Tsunewaki 2009). Originally, Ae. neglecta was given the genomic formula C^uC^uM^tM^t (Kihara 1937, 1949; 1954; Morris and Sears 1967). The C^u subgenome (currently U) was derived from Ae. umbellulata and the M^t subgenome (Currenly Xⁿ) was thought to be a modified form of the Ae. comosa M genome (Kihara 1937, 1949). While Kihara did not analyze hybrids between Ae. neglecta and its assumed diploid parents, he determined the genomic formula of Ae. neglecta from chromosomal pairing in hybrids between Ae. neglecta and other allotetraploid species bearing U and M subgenomes. The Ae. neglecta (genome UUXⁿXⁿ) x Ae. biuncialis (genome UUM^bM^b) hybrid had 5–10 bivalents, with a mode of 8, the hybrid between neglecta and columnaris (genome UUXⁿXⁿ) had more than 12 bivalents (Kihara 1936, 1940a, b), and another work on neglecta x columnaris hybrid reported 1.98 univalents, 12.54 bivalents, 0.26 trivalents and 0.04 quadrivalents (Furuta and Tanaka 1970). Consequently, Kihara (1936, 1940a, b, 1954) assigned the genomic formulas C^uC^uM^tM^t to Ae. neglecta. Kimber and Yen (1989) and Yen and Kimber (1992b) reported that all the accessions of Ae. neglecta they used had an essentially unchanged U subgenome, which reinforced Kihara’s conclusion concerning the origin of the U subgenome. Tsuchiya (1956) produced hybrids between Ae. neglecta and its assumed diploid parent Ae. comosa, but most of the seeds did not germinate. In one hybrid that did germinate, the meiotic behavior of the chromosomes was not studied in detail, as the anthers degenerated. However, a few configurations observed at first metaphase of several meiotic cells, included 2 or 3 bivalents and many univalents. This may indicate that the second subgenome of Ae. neglecta is either different or greatly modified from the M genome of Ae. comosa. Additional hybrids between neglecta and comosa were not studied. Hybrids between Ae. neglecta and autotetraploid Ae. uniaristata showed that there is no N genome in neglecta (Kimber and Yen 1989).

Upon analysis of the karyotype of Ae. Neglecta, Chennaveeraiah (1960), like Senyaninova-Korchagina (1932), reported that there is only one pair of chromosomes with satellites on short arms, one pair with an extreme subterminal centromere, four pairs with submedian-subterminal centromeres, one pair with median centromeres and the rest with submedian centromeres. Senyaninova-Korchagina (1932) has called attention to the resemblance of one half of the chromosomes in Ae. neglecta to the chromosomes of Ae. umbellulata. The most distinguished character of the U genome is the single pair with the extremely subterminal centromere. Ae. umbellulata has two chromosome pairs with satellites, whereas Ae. neglecta has only one pair with satellites. It seems certain, however, that the U subgenome is present in Ae. neglecta either in a pure or in a slightly modified form. The second set of chromosomes includes five types, one pair with a median centromere and the rest with submedian centromeres. No secondary constrictions or satellites which are characteristic of the M genome are present here. The M^t genome of Ae. neglecta is either considerably modified from the basic type or it must be a genome foreign to Aegilops or at least to the known diploid analyzers of Kihara (1954).

Ae. neglecta and Ae. columnaris are closely related to each other and have the same genomes (Resta et al. 1996). Variations in restriction fragments of repeated nucleotide sequences showed that their second subgenome did not derive from the M genome of Ae. comosa. While they share the U subgenome with Ae. biuncialis and Ae. geniculata, their second subgenome is unrelated to the M^o or M^b subgenomes of these species. No relationship was found between this subgenome and the genome of any extant diploid species of Aegilops or any phylogenetic lineage leading to the extant diploid species (Resta et al. 1996; Dvorak 1998). This unknown genome was designated X^t by Resta et al. (1996) and Dvorak (1998), but was recently changed to Xⁿ, (X of neglecta) and consequently, the proposed genomic formula for Ae. neglecta and Ae. columnaris is UUXⁿXⁿ.

Analysis of the restriction fragment pattern of chloroplast DNA of Ae. neglecta and of Ae. umbellulata using restriction endonucleases (Ogihara and Tsunewaki 1982; Terachi et al. 1984), showed that the chloroplast genome (the plastom) of Ae. neglecta arose from Ae. umbellulata. Studies on the chondrion genome also showed that the cytoplasm (the plasmon) of Ae. neglecta derived from Ae. umbellulata (Terachi and Tsunewaki 1986). Comparison the plasmon of several U-subgenome allotetraploid species with that of Ae. umbellulata indicated that Ae. neglecta is of more recent origin than Ae. geniculata (Tsunewaki 2009). On the other hand, the four-tetraploid species, Ae. biuncialis, Ae. columnaris Ae. neglecta and Ae. triuncialis arose from Ae. umbellulata as female (Tsunewaki 1996, 2009). The distance between Ae. neglecta and Ae. umbellulata is 0.01 but is zero between the other three tetraploid and Ae. umbellulata. This result suggests that Ae. neglecta is the oldest of the four tetraploids.

Kadosumi et al. (2005), studying variation of the genome-specific PCR primer set U31 in Ae. umbellulata and Ae. neglecta, found three alleles of this DNA sequence in Ae. umbellulata, and two in Ae. neglecta. This result indicated that the U genome had at least two, probably more, independent origins in Ae. neglecta. Similarly, Meimberg et al. (2009) showed that Ae. neglecta originated from two independent hybridizations as indicated by presence of at least two different Ae. umbellulata chloroplast haplotypes. This multiple origin could have introduced genetic variability that increased the ecological amplitude and evolutionary success of Ae. neglecta.

Ae. neglecta contains 10.64 ± 0.404 pg 1C DNA (Eilam et al. 2008). The genome of Ae. neglecta, and that of its closest relative Ae. columnaris, are larger than the genomes of Ae. biuncialis and Ae. geniculata (Eilam et al. 2008). This may indicate that the genome of the, yet unknown, diploid donor of the subgenome Xⁿ is larger than the M genome of Ae. comosa, the putative donor of the M subgenome to Ae. biuncialis and Ae. geniculata.

Badaeva (2002) identified all the chromosomes of Ae. neglecta on the basis of morphology and C-banding patterns, while Badaeva et al. (2004) also used FISH. All accessions they analyzed possessed three pairs of NORs, two of which were attributed to the U and one to the Xⁿ subgenome. Yet, Senyaninova-Korchagina (1932) and Chennaveeraiah (1960) found only one pair of SAT chromosomes in Ae. neglecta. The values obtained by Badaeva (2002) and Badaeva et al. (2004) corresponded to the number of active NORs detected by Teoh et al. (1983) and Ag-NOR staining (Cermeño et al. 1984b). The absence of secondary constriction on two chromosome pairs carrying NORs indicated that the respective NORs were inactivated. Nevertheless, C-banding analysis showed the presence of active NORs on chromosomes 1U, 5U and a Xⁿ chromosome pair.

Comparison of 10 Ae. neglecta accessions from diverse geographical regions, revealed low variability in the C-banding patterns and translocation polymorphism (Badaeva 2002, and Badaeva et al. 2004). Badaeva et al. (2004) performed FISH with the heterochromatin-limited repetitive DNA probes pSc119, pAs1, and studied the distribution of NORs (18S-26S rDNA) and 5S rDNA loci using the pTa71 (18S-26S rDNA), and pTa794 (5S rDNA) probes. The data obtained confirmed significant differences in karyotype structure, in the total amount and distribution of heterochromatin, and in the number and location of 5S and 18S-26S rDNA loci between Ae. neglecta -Ae. columnaris, and Ae. geniculata—Ae. biuncialis. Ae. geniculata and Ae. biuncialis showed moderate heterochromatin content, with small- or medium-sized bands located in telomeric and interstitial chromosome regions. The C-banding patterns of these two-species corresponded with that of their diploid ancestors Ae. umbellulata and Ae. comosa. In contrast to the C-banding patterns of Ae. geniculata and Ae. biuncialis, Ae. neglecta and Ae. columnaris chromosomes were characterized by a high heterochromatin content, and large C-bands or C-band complexes located in pericentromeric, interstitial, and telomeric chromosome regions. Similarities in C-banding and FISH patterns of most Ae. columnaris and Ae. neglecta chromosomes suggest that they likely had a common ancestral diploid species. The differences in three chromosome pairs may indicate that the divergence of these two species was probably associated with chromosomal rearrangements and/or introgressive hybridization. Three intergenomic translocations between U and Xⁿ chromosomes exist in Ae. neglecta.

These data reinforced the conclusion of Resta et al. (1996) and Dvorak (1998) that Ae. neglecta and Ae. columnaris (genomes (UUXⁿXⁿ) had a different origin than Ae. geniculata and Ae. biuncialis (genomes M^oM^oUU and UUM^bM^b, respectively). The similarity of the C-banding patterns of Ae. neglecta and Ae. columnaris chromosomes suggest that they originated from a common ancestor, with Ae. umbellulata being the U-subgenome donor, while the donor of the second subgenome is yet to be determined. Moreover, Badaeva (2002) assumed that Ae. neglecta and Ae. columnaris exchanged genetic material through processes of introgressive hybridization with other tetraploid species bearing the U subgenome.

9.8.5.3 Crosses with Other Species of the Wheat Group

Using C-banding to study chromosome pairing at meiotic metaphase of the two subgenomes of Ae. neglecta, (Cuñado 1992) showed diploid-like meiotic behavior, namely, only bivalents were formed between fully homologous chromosomes. They analyzed the pattern of chromosome pairing at the meiotic zygotene and pachytene stages in Ae. neglecta by whole-mount surface-spreading of synaptonemal complexes under the electron microscope. The observations indicated that already at these early meiotic stages, the chromosomes were almost exclusively associated as bivalents.

There are no data on chromosomal pairing in hybrids between Ae. neglecta and its diploid parent Ae. umbellulata, the donor of the U subgenome. However, chromosomal pairing in the hybrid Ae. neglecta x autotetraploid Ae. umbellulata supported the assumption that Ae. neglecta contains the U subgenome and that this subgenome is fully homologous to the U genome of Ae. umbellulata (Kimber and Yen 1989; Yen and Kimber 1992b). On the other hand, there are no data on chromosomal pairing in the hybrid between Ae. neglecta and Ae. comosa, that was assumed to be the donor of the second subgenome of neglecta. Nevertheless, there are data on meiotic chromosomal pairing in hybrids between Ae. neglecta and other U-subgenome-bearing allotetraploids. The F₁ Ae. neglecta x Ae. columnaris hybrid had 11–14 bivalents with a mode of 12 bivalents (Kihara 1949), indicating that these two allotetraploids are very close to each other and share close genomes. The hybrid neglecta x geniculata had 5–10 bivalents, with a mode of 8 bivalents, 0–2 trivalents and 0–1 quadrivalents (Kihara and Nishiyama 1937), and the hybrid neglecta x biuncialis had similar pairing, i.e., 5–10 bivalents, with a mode of 8 bivalents, 0–1 trivalents and 0–1 quadrivalent (Kihara 1949). This degree of chromosomal pairing indicates that geniculata and biuncialis are close to neglecta but, since all three contain the U subgenome, the reduced pairing in the hybrids between them indicates that they differ in their second subgenome. The Ae. neglecta x Ae. triuncialis hybrid (genome UXⁿUC) had 0–9 bivalents, with a mode of 5 bivalents, and 0–1 trivalents (Lindschau and Oehler 1936; Kihara 1949), the hybrid Ae. neglecta x Ae. peregrina or Ae. neglecta x Ae. kotschyi (hybrids genome UXⁿS^vU) had 7–10 bivalents, with a mode of 9 bivalents, and 0–2 trivalents, indicating that Ae. neglecta differs in its second subgenome from Ae. triuncialis, Ae. peregrina and Ae. kotschyi. On the other hand, chromosome pairing in the pentaploid hybrid Ae. neglecta x Ae. recta (genome UXⁿUXⁿN) showed 14 bivalents and 7 univalents (Kihara 1937) indicating that the two species share two subgenomes and that the third subgenome of Ae. recta differ from the two subgenomes of Ae. neglecta.

Chromosomal pairing in hybrids between Ae. neglecta and diploid species having the S, C, and N genomes showed that the two subgenomes of Ae. neglecta are not homologous to those of the diploids. More specifically, the Ae. neglecta x Ae. speltoides hybrid (genome UXⁿS) had 2–8 bivalents, with a mode of 4–5 bivalents, the Ae. neglecta x Ae. caudata hybrid (genome UXⁿC) had 6.30 univalents, 4.48 rod bivalents, 0.17 ring bivalents, 1.73 trivalents and 0.05 quadrivalents (Kimber and Abu-Baker 1981), and the Ae. neglecta x Ae. uniaristata hybrid (genome UXⁿN) had 2–6 bivalents, with a mode of 4 bivalents (Kihara 1949).

The frequency of homoeologous pairing at first meiotic metaphase in F₁ hybrids between T. turgidum subsp. durum cv. Langdon and Ae. neglecta was determined by a genomic in situ hybridization (GISH) procedure that allowed simultaneous discrimination of durum and. neglecta chromosomes (Cifuentes et al. 2010). Chromosomal pairing was low in this hybrid; average pairing contained 22.71 univalents, 2.37 rod bivalents, 0.01 ring bivalents, 0.16 trivalents and 0.02 quadrivalents, with a total of 2.75 associations per cell. Some of the associations were auto syndetic, namely, 0.19 associations/cell were between A and B chromosomes of durum and 0.67 associations/cell were between U and Xⁿ chromosomes of neglecta. However, most of the associations were allosyndetic, with 1.39 associations/cell between A and UXⁿ chromosomes and 0.5 between B and UXⁿ chromosomes. Hence, interspecific durum-neglecta associations account for 69% of total first meiotic metaphase pairing. Chromosomes of the A subgenome were most frequently involved in pairing with their neglecta homoeologues than the B subgenome chromosomes (Cifuentes et al. 2010).

9.8.6 Aegilops recta (Zhuk.) Chennav.

9.8.6.1 Morphological and Geographical Notes

Ae. recta (Zhuk.) Chennav. [Syn.: Ae. triaristata Willd.; Ae. triaristata subsp. recta Zhuk.; Ae. neglecta Req. ex Bertol.; Ae. neglecta subsp. tecta (Zhuk.) Hammer; T. neglectum (Req. ex Bertol.) Greuter; T. rectum Bowden] is an annual, predominantly autogamous, 20–35-cm-tall (excluding spikes) plant. The uppermost 1/4 or 1/3 of the culms is defoliated. Leaf blades are more or less hairy, often with ciliate margins. Its spike is lanceolate, compact, 2.0–3.5-cm-long (excluding awns), awned, and generally becoming narrow in the upper parts. The entire spike disarticulates at maturity and falls as a unit (umbrella-type dispersal unit). The lower rachis internode is shorter than the adjacent spikelet. There are three basal rudimentary spikelets, seldom two. There are 3–6 spikelets, usually 4, with the two lowest being narrow to broad-elliptical, proportionally larger than the upper ones and lying against each other, while the upper two are usually fertile, seldom sterile, and project from the lower part of the spike. There are 4–5 florets, the two lower ones being fertile. The glumes are awned and broad, with curved, unequally broad, flattened nerves. There are usually 3 glume awns on the upper spikelets, and 2–3 on the lower spikelets. There are 2–4 lemma awns, usually 2, on lower spikelets and usually none on upper ones, and if they exist, they are poorly developed. Often the awns in the upper spikelets decrease in length so that all awns of the spike end at the same height. The awns of the mature spike are generally weak, seldom strong. The caryopsis is free (Fig. 9.4e).

Originally, Ae. recta has been included in Ae. neglecta (formerly Ae. triaristata) which contained tetraploid and hexaploid cytotypes. Zhukovsky (1928), and Kihara (1954, 1963) considered the two forms subspecies of Ae. triaristata, whereas Hammer (1980) and van Slageren (1994) considered them subspecies of Ae. neglecta, i.e., the tetraploid forms as ssp. neglecta and the hexaploid ones as ssp. recta (Zhuk,) Hammer.

The tetraploid and hexaploid subspecies of Ae. neglecta posed a taxonomic problem in the sense that they are almost undistinguishable, yet, the different ploidy level causes some reproductive isolation and subsequent speciation (van Slageren 1994). The association ‘recta’ with the hexaploid level was made for the first time by Senyaninova-Korchagina (1930) in her karyosystematical overview of Aegilops. Since then, the epithet ‘recta’ has been associated with the hexaploid level (e.g., Chennaveeraiah 1960; Bowden 1966; Löve 1984; Kimber and Feldman 1987). Yet, the F₁ hybrid between the tetraploid and the hexaploid subspecies had 14 bivalents and 7 univalents (Kihara 1937), indicating that the third subgenome in Ae. recta differ from the two subgenomes of Ae. neglecta. The F₁ hybrid is sterile (Chennaveeraiah 1960) or partially fertile (Kihara 1963). Therefore, according to Chennaveeraiah (1960), the two taxa should be treated as separate species. Consequently, Chennaveeraiah (1960) elevated the hexaploid forms to the species rank and named the new species Ae. recta (Zhuk.) Chennav.

Separation of Ae. neglecta and Ae. recta to two different species would be easy if morphological characters linked to the ploidy level can be identified. In this regard, Chennaveeraiah (1960) described few morphological differences between Ae. neglecta and Ae. recta, namely, there are 2 to 3 spikelets in Ae. neglecta, whereas Ae. recta has 5 to 6. Ae. neglecta awns are spread at maturity, while they are slanted in Ae. recta. Glumes are glabrous in Ae. neglecta and usually hairy in Ae. recta. In addition, Kihara (1963) noted that the spikes of Ae. neglecta suddenly taper, whereas those of Ae. recta taper gradually. Kimber and Feldman (1987) suggested that the hexaploid forms possess fertile terminal spikelet(s) while the tetraploid forms have sterile ones. Yet, there are many morphological intermediates between the two species, at times rendering it difficult to differentiate between them (Kihara 1963).

Aryavand et al. (2003) reported on a significant difference in stomatal counts and size in the leaves between the two species. Higher ploidy in Ae. recta is associated with fewer but larger stomata per unit leaf area, showing a highly negative correlation between stomatal count and size. Hence, this trait can assist in the identification of the two species. Additionally, Giraldo et al. (2016) developed two chloroplast DNA-based molecular markers that accurately discriminate Ae. recta from Ae. neglecta. The use of these markers, in addition to chromosome counting, facilitates further the ability to differentiate Ae. recta from neglecta.

Ae. recta exhibit relatively limited morphological variation involving spike size, color and hairiness. Its variation is relatively low compared with that of other polyploid species of Aegilops. Monte et al. (2001) used AFLP DNA markers to characterize the genetic diversity in Ae. recta populations distributed in the Iberian Peninsula and Balearic Islands. Ae. recta exhibited low variation in contrast with Ae. biuncialis, that presented a high degree of polymorphism.

Originally, there was confusion concerning the distribution of Ae. neglecta and Ae. recta (Ohta et al. 2016). The distribution of Ae. neglecta senso lato was usually described as Mediterranean and West Asiatic, namely, from Morocco and Portugal in the west to Transcaucasia, western Iran and Turkmenistan in the east. There was almost no attempt to determine the distribution of the two cytotypes of Ae. neglecta. Also, after the separation of the tetraploid forms to a different species, the geographical distributions of the two-species remained unclear. To determine more accurately the distribution of these two species, Ohta et al. (2016) analyzed the chromosome numbers of accessions of these two species from 137 populations, located in the western area of the species distribution from the Aegean Islands to Morocco. Taken together with data from previous studies, Ohta et al. (2016) revealed a difference in the geographical distribution of Ae. neglecta and Ae. recta: Ae. neglecta is distributed in the eastern part of the species area, whereas Ae. recta predominantly occurs in the western part, with the border between these two species on the western margin of the Aegean Sea. Near the border, Ae. neglecta and mixed populations of both species were sporadically found among populations of Ae. recta in the Balkan and Peloponnesus Peninsulas, while a few Ae. recta and mixed populations were found among populations of Ae. neglecta in the East Aegean Islands and West Anatolia (Ohta et al. 2016).

Support of the conclusions reached by Ohta et al. (2016) was provided by Baik et al. (2017), who carried out a karyological study of several populations of Ae. neglecta from different eco-geographical sites in North Algeria. Chromosome counting showed that all accessions were hexaploids, that is to say, Ae. recta. Similar results were obtained by Belkadi et al. (2003), who determined chromosome counts in two accessions of Ae. neglecta from Morocco and found them to be hexaploids.

Hence, Ae. recta is a West Mediterranean element. It grows in Algeria, Morocco, Portugal, Spain, southern France, Italy, Croatia, Bosnia-Herzegovina, Serbia, Bulgaria, Montenegro, North Macedonia, Albania, Greece, and western Turkey. It grows on terra rossa soil in edges and openings of sclerophyllus oak forests and maquis and dwarf-shrub formations, in abandoned fields, edges of cultivation, and in disturbed and eroded areas and roadsides. It is relatively common.

Ae. recta has a medium-sized distribution in the western part of the distribution of the genus. It grows sympatrically with its two parents, Ae. neglecta and Ae. uniaristata, in the Balkan, the Aegean islands and western Turkey. Since its two parents grow sympatrically in West Turkey and the Balkan, this is presumably the region where Ae. recta originated and from which it spread westward (Kihara 1963).

Ae. recta usually grows in mixed stands, with other species with which it introgresses. It grows sympatrically with Ae. umbellulata, Ae. comosa, Ae. uniaristata, Ae. caudata, Ae. neglecta, Ae. geniculata, Ae. biuncialis, Ae. triuncialis, and Ae. ventricosa, and allopatrically with Ae. columnaris.

In Europe, Ae. recta grows sympatrically with common wheat and spontaneous hybridization between these two species is known to occur (Zaharieva and Monneveux 2006). Arrigo et al. (2011) investigated introgression between common wheat and Ae. recta and compared wheat field borders to areas isolated from agriculture. All Ae. recta had 2n = 42. Individuals were characterized with AFLP fingerprinting, analyzed through two computational approaches (i.e., Bayesian estimations of admixture and fuzzy clustering), and sequences marking wheat-specific insertions of transposable elements. With this combined approach, Arrigo et al. (2011) detected substantial gene flow between wheat and Ae. recta, and noted significantly more admixed individuals close to wheat fields than in locations isolated from agriculture. Arrigo et al. (2011) concluded that reproductive barriers have been regularly bypassed during the long history of sympatry between wheat and Ae. recta.

Gill et al. (1985) reported that Ae. recta confers resistance to powdery mildew, leaf rust and Hessian Fly. El Bouhssini et al. (1998) also found that Ae. recta from Morocco showed resistance to Hessian Fly.

9.8.6.2 Cytology, Cytogenetics and Evolution

Ae. recta are an allohexaploid species (2n = 6x = 42). Its nuclear genome is designated as UUXⁿXⁿNN (modified from Dvorak 1998) and the plasmon genome is designated U, similar to that of Ae. umbellulata (Tsunewaki 2009). Kihara (1937), based on the observation of 14 ring bivalents in the pentaploid hybrid between tetraploid Ae. neglecta and hexaploid Ae. recta, concluded that Ae. recta contain the two subgenomes of Ae. neglecta, U and M^t (currently Xⁿ), and consequently, regarded Ae. recta as a cytotype of Ae. neglecta (Kihara 1963). Karyotype analyses confirmed that Ae. recta contain the two subgenomes of Ae. neglecta (Senyaninova-Korchagina 1930, 1932; Chennaveeraiah 1960). Kihara (1957), using indirect pairing data obtained from hybrids between Ae. recta and other allotetraploid species bearing the M subgenome, as well as from the morphological similarity between Ae. recta and Ae. neglecta, believed that the third subgenome of Ae. recta is a modified form of genome M of Ae. comosa. So, Kihara (1963) designated the genome of Ae. recta C^uC^uM^tM^tM^t2M^t2. This genomic formula indicated that Ae. recta are, in fact, an allo-auto-hexaploid containing two modified M subgenomes. But, Kihara’s assumption has never been confirmed by direct evidence from hybrids between Ae. recta and Ae. comosa.

As was pointed out by Senyaninova-Korchagina (1930, 1932) and Chennaveeraiah (1960), fourteen chromosome pairs of Ae. recta are identical to those of Ae. neglecta; thus, karyotypic results also revealed that two of the subgenomes of Ae. recta derived from Ae. neglecta. The third chromosomal set resembles the chromosomes of Ae. uniaristata, although Ae. uniaristata contains no chromosome with a median centromere. Bowden (1959) suggested that one subgenome of Ae. neglecta and Ae. recta derived from Amblyopyrum muticum, yet Chennaveeraiah (1960) rejected this suggestion because the karyotypes of Ae. neglecta and Ae. recta contained no A. muticum genome.

Evidence of the presence of the U subgenome in Ae. recta has been indirect, based largely on studies of hybrids between allopolyploid species, of which one species, Ae. geniculata, had previously been shown to have the Ae. umbellulata genome (von Berg 1937; Kihara 1937, 1940a, b, 1949; Kimber et al. 1988). Indirect evidence also came from the analysis of chromosome pairing in the F₁ hybrid between Ae. recta and a synthetic tetraploid Ae. umbellulata – Ae. uniaristata (hybrid genome UXⁿNUN), which showed good chromosome pairing; the average number of bivalents was 13.4 (Kihara 1963). On the other hand, the hybrid between Ae. neglecta (genome UUXⁿXⁿ) and this synthetic tetraploid (hybrid genome UXⁿUN) showed an average of only 8.3 bivalents. Hence, it was concluded that Ae. neglecta and Ae. recta share the U subgenome, while the third subgenome of Ae. recta (subgenome N) is entirely different from the second subgenome of Ae. neglecta (Xⁿ). Kihara (1963) assumed that the third subgenome of Ae. recta is closely related the genome of Ae. uniaristata, which was designated M^u, since Kihara (1954) believed it to be a modified M genome, and consequently, designated it M¹² (Kihara (1963). Accordingly, Kihara (1963) concluded that Ae. recta was formed via hybridization of Ae. neglecta x Ae. uniaristata. Since these two species grow in mixed stand in western Turkey and the southern Balkan, Ae. recta was formed there and then spread westward. But, data on chromosome pairing in interspecific hybrids do not agree with the assumption that the second and third subgenomes of Ae recta are modified M (Kimber et al. 1983; Resta et al. 1996).

The presence of the U subgenome in Ae. recta was also confirmed by crossing this species with an induced autotetraploid of Ae. umbellulata (2n = 4x = 28; genome UUUU) (Kimber and Yen 1989; Yen and Kimber 1992b). Chromosome pairing was analyzed at first meiotic metaphase in this pentaploid hybrid that had an average of 9.30 univalents, 4.76 rod bivalents, 3.16 ring bivalents, 2.83 trivalents, 0.33 quadrivalents and 0.10 pentavalents. The optimization analysis of the meiotic data fit the 3:2 model, indicating that there are three homologous genomes in the hybrid (Yen and Kimber 1992b). Since two U genomes were introduced from the autotetraploid Ae. umbellulata, it is quite clear that there is a U subgenome in the hexaploid Ae. recta which is homologous to genome U of Ae. umbellulata (Yen and Kimber 1992b).

Likewise, the origin of the third subgenome of Ae. recta was determined by studying chromosome pairing in the hybrid Ae. recta x an induced autotetraploid of Ae. uniaristata (2n = 4x = 28; genome NNNN), which showed an average of 9.85 univalents, 5.05 rod bivalents, 2.30 ring bivalents, 2.95 trivalents and 0.40 quadrivalents (Yen and Kimber 1992b). Also in this hybrid, the optimization analysis of the meiotic data fit the 3:2 model, indicating that there are three homologous genomes in the hybrid. Since two were introduced from the autotetraploid Ae. uniaristata, it was concluded that the third subgenome of the hexaploid Ae. recta was homologous to genome N of Ae. uniaristata (Yen and Kimber 1992b). However, since the pairing data of another hybrid involving a different line of Ae. recta and the autotetraploid of Ae. uniaristata indicated that some differences may exist between the N genomes, and since no data were available on the pairing of the Ae. recta chromosomes with Ae. comosa chromosomes, the existence of an N genome in Ae. recta was not unequivocally established. If Ae. uniaristata had contributed chromosomes to Ae. recta, they must have undergone substantial modification (Yen and Kimber 1992b).

The origin of the Ae. recta genome was also investigated by examining the presence of specific restriction fragments of repeated nucleotide sequences in its DNA (Resta et al. 1996). The data showed that all bands of Ae. neglecta and Ae. columnaris, both bearing genome UUXⁿXⁿ, were present in Ae. recta (Resta et al. 1996) indicating that one of these species is the tetraploid parent of Ae. recta. Ae. neglecta is morphologically very similar to Ae. recta and is therefore more likely a tetraploid ancestor of Ae. recta than Ae. columnaris (Resta et al. 1996).

Yen and Kimber (1992b) considered their study concerning the origin of the third subgenome of Ae. recta as inconclusive since they did not investigate the genetic connection between Ae. recta and Ae. comosa. This relationship was investigated by Resta et al. (1996), who showed that Ae. comosa did not contribute any of the three Ae. recta subgenomes. They determined the value of the repeated nucleotide sequence correspondence (RSC), that is, the fraction of marked bands shared between Ae. uniaristata and Ae. recta and found it to be 1.00, meaning that all marker bands of Ae. uniaristata were encountered in Ae. recta. This finding provided strong evidence that Ae. uniaristata donated the third subgenome to Ae. recta. Thus, the most probable origin of Ae. recta was the hybridization of Ae. neglecta with Ae. uniaristata. Resta et al. therefore proposed, to revise Kihara’s (1963) formula C^uC^uM^tM^tM^t2M^t2 for Ae. recta to UUX^tX^tNN (currently UUXⁿXⁿNN).

Tsunewaki (2009) reviewed his team investigations on the origins of the plasmons of the various diploid and polyploid species in the Aegilops-Triticum group. RFLP analyses of chloroplast (cp) DNA (Terachi et al. 1984; Ogihara and Tsunewaki 1988; Tsunewaki 1996) and mitochondrial (mt) DNA (Terachi and Tsunewaki 1986), as well as PCR–single-strand conformational polymorphism analyses of chloroplast and mitochondrial DNAs (Wang et al. 1997) showed that the plasmon of Ae. recta was identical to that of Ae. neglecta and that both were similar to that of Ae. umbellulata and therefore, were designated U. A similar conclusion from recent analyses of chloroplast DNA was reached by Bernhardt et al. (2017). The relative times of origin of the allopolyploid species of section Aegilops were inferred from the genetic distances of their plasmon from that of their putative maternal parent, Ae. umbellulata (Terachi et al. 1984; Wang et al. 1997; Tsunewaki 2009). Thus, it was suggested that the origin of Ae. geniculata was older than that of Ae. neglecta, which, in turn, was older than that of Ae. biuncialis and Ae. columnaris (Terachi et al. 1984; Wang et al. 1997; Tsunewaki 2009). Obviously, the origin of Ae. recta, which derived from hybridization of Ae. neglecta, as female parent, and Ae. uniaristata, was more recent than that of Ae. neglecta.

Ae. recta contain 16.22 pg 1C DNA (Eilam et al. 2008), which is 1.46% less DNA than expected from the sum of the DNA content of its two parents, i.e., 16.46 pg (Ae. neglecta contains 10.64 pg and Ae. uniaristata contains 5.82 pg; Eilam et al. 2007, 2008). The loss of some DNA in Ae. recta was also found by Badaeva et al. (2002, 2004) who, based on differential C-banding and FISH, found that Ae. recta exhibited several minor structural chromosome modifications, including deletion of chromosomal segments.

According to Senyaninova-Korchagina (1930, 1932), the hexaploid species, Ae. recta, had all the chromosomes present in the tetraploid species, Ae. neglecta, plus 7 chromosome pairs which had only submedian centromeres. Yet, the karyotype described by Chennaveeraiah (1960) showed that the description by Senyaninova-Korchagina (1930, 1932) of seven additional chromosome pairs in Ae. recta was incorrect. According to Chennaveeraiah (1960), the karyotype of Ae. recta consisted of two pairs with satellites of different sizes on short arms, one pair with an extreme subterminal centromere, seven pairs with submedian-subterminal centromeres, two pairs with median centromeres and the rest with submedian centromeres. The extra seven pairs in the hexaploid, therefore, consisted of one pair with satellites, one pair with median centromere, four pairs with submedian-subterminal centromeres and one pair with a submedian centromere. Therefore, this third set in Ae. recta is not in any way a duplicated complete set of either of the other two sets in the tetraploid parent, Ae. neglecta. In all, there are 16 types of chromosomes in Ae. recta (Chennaveeraiah 1960).

In accord with the findings of Senyaninova-Korchagina (1932) and Chennaveeraiah (1960), Teoh et al. (1983), using FISH with a repetitive DNA that codes for rRNA, found two pairs of SAT chromosomes and four pairs of rRNA sites in Ae. recta. Badaeva (2002) and Badaeva et al. (2004) observed that all analyzed accessions of Ae. recta possessed three SAT chromosomes and four major NORs on chromosomes 1U, 5U, 1Xⁿ, and 5N, only three of which were active. These values correspond to the number of SAT chromosomes and active NORs detected by Ag-NOR staining (Cermeño et al. 1984b). All in all, the results of C-banding analysis and FISH reported by Badaeva et al, (2004) in Ae. recta showed the presence of active NORs on chromosomes 1U, 5U and 1Xⁿ and inactivation of NOR on chromosome 5N.

Badaeva et al. (2004) analyzed the heterochromatin banding patterns of somatic metaphase chromosomes revealed by C-banding in several accessions of Ae. recta, and revealed low variation in C-banding patterns and translocation polymorphism. Four accessions from Spain, Portugal and Morocco had a Robertsonian translocation between chromosomes 6Xⁿ and 3N. Another Robertsonian translocation between 2Xⁿ and 3N was identified in an accession from Bulgaria. Badaeva et al. (2004, 2011) claimed that this relatively low variation of C-banding demonstrated that the formation of this allohexaploid involved only minor functional modifications of the parental genomes, while intraspecific divergence was accompanied by genome rearrangements, namely, translocations involving the total chromosome arms of all three subgenomes (Badaeva et al. 2011).

The distribution of C-bands on chromosomes of the U and Xⁿ subgenomes of Ae. recta was mostly identical and similar to that of chromosomes of Ae. neglecta. Likewise, the C-banding patterns of the N subgenome chromosomes of Ae. recta were similar to those of Ae. uniaristata chromosomes. It was possible to identify all Ae. recta chromosomes on the basis of morphology and C-banding patterns. Similar to its tetraploid parent, Ae. neglecta, the karyotype of Ae. recta was characterized by a high heterochromatin content (Badaeva 2002).

In addition, Badaeva et al. (2004) used FISH with the heterochromatin-limited repetitive DNA probes pSc119 and pAs1, and the pTa71 probe, to reveal the 18S-26S rDNA sites, and pTa794, to reveal the 5S rDNA sites. The pSC119 and pAs1 FISH patterns on U and Xⁿ subgenome chromosomes of Ae. recta were nearly identical to those of Ae. neglecta. Also, no differences in the labeling pattern of the N subgenome chromosomes were observed between Ae. recta and Ae. uniaristata (Badaeva et al. 2004). Therefore, in contrast to the conclusion of Yen and Kimber (1992b), the N subgenome of Ae. recta appeared to be only slightly structurally modified from that of Ae. uniaristata. Hence, the UXⁿ and the N subgenome chromosomes of Ae. recta are similar to those of Ae. neglecta and Ae. uniaristata, respectively, with regard to the distribution of C-bands, 45S and 5S rDNA loci and FISH sites of pSc119 and pAs1 (Badaeva 2002; Badaeva et al. 2004, 2011).

Four major (1U, 5U, 1Xⁿ, 5N) and two minor (7Xⁿ and 1N) NORs and seven 5S rDNA loci were detected on all group-1 and group-5 chromosomes in Ae. Recta, using pTa71 and pTa794. Thus, the number of NORs and 5S rDNA in Ae. recta was similar to the total number of these loci in the parental species. However, the number of minor 18S-26S rDNA sites was smaller than expected.

9.8.6.3 Crosses with Other Species of the Wheat Group

Chromosome associations at first meiotic metaphase were studied in the allohexaploid species, Ae. recta, using the C-banding technique (Cuñado 1992). Mean chromosomal pairing included 0.12 univalents, 5.31 rod bivalents, 15.57 ring bivalents, and 36.45 mean chromosome associations per metaphase cell. Use of C-banding enabled the analysis of associations between homologous chromosomes of each of the three subgenomes. Average homologous associations between chromosomes of the UXⁿ subgenomes showed 0.06 univalents, 3.16 rod bivalents, 10.78 ring bivalents and 24.09 mean chromosome associations per metaphase cell, and between chromosomes of the N subgenomes showed 0.05 univalents, 2.13 rod bivalents, 4.81 ring bivalents and 11.75 mean chromosome associations for a metaphase cell (Cuñado 1992). No difference between the UXⁿ and N subgenomes was noted in regards to the mean associations per bivalent.

The allohexaploid Ae. recta regularly form bivalents at first meiotic metaphase (Cuñado 1992). Cuñado et al. (2005) analyzed the pattern of synapsis at late zygotene and pachytene in Ae. recta using whole-mount surface-spreading of synaptonemal complexes under an electron microscope. It was revealed that the chromosomes were mostly associated as bivalents in these early meiotic stages, with a mean 0.17 multivalents per nucleus. It can be concluded that the mechanism controlling bivalent formation in this species acts mainly at zygotene, by restricting synapsis to homologous chromosomes, but also acts at pachytene, by preventing chiasma formation in the homoeologous associations (Cuñado et al. 2005).

Chromosomal pairing was studied in F₁ hybrids of Ae. recta x several allotetraploid species bearing the U subgenome. Only the Ae. neglecta x Ae. recta hybrid (genome UXⁿUXⁿN) had 14 ring bivalents and 7 univalents (Kihara 1937), indicating that only U and Xⁿ subgenomes are homologous to those of Ae. neglecta. On the other hand, the other hybrids exhibited a reduced level of pairing. More specifically, the hybrid Ae. geniculata x Ae. recta (genome M^oUUXⁿN) had 6–7 bivalents (Kihara 1937), Ae. recta x Ae. biuncialis (genome UXⁿNUM^b) had 6–11 bivalents and 0–1 trivalent (Kihara 1949), the hybrid Ae. recta x Ae. triuncialis (genome UXⁿNUC) had + 7(-9) bivalents (Kihara 1937) and the hybrid Ae. recta x Ae. peregrina (genome (UXⁿNS^vU) had 7–11 bivalents and 0–1 trivalent Kihara 1949). The amount of chromosomal pairing in these four hybrids showed that Ae. recta shared only one subgenome, i.e., U, with Ae. geniculata, Ae. biuncialis, Ae. triuncialis, and Ae. peregrina.

The hexaploid hybrid between Ae. recta and hexaploid Ae. crassa (genome UXⁿND^cX^cD) had 7–8 bivalents, with a mode of 8 bivalents, and 0–2 trivalents (Kihara 1949). Since most of the pairing in this hybrid was presumably autosyndetic between subgenomes D^c and D of Ae. crassa, the little remaining pairing was allosyndetic pairing between chromosomes of the homoeologous subgenomes.

9.8.7 Aegilops columnaris Zhuk.

9.8.7.1 Morphological and Geographical Notes

Ae. columnaris Zhuk. [Syn.: Triticum columnare (Zhuk.) Morris and Sears] is an annual, predominantly autogamous, multi-tillered plant. Its culms are 20–50-cm-tall (excluding spikes) and slightly geniculate at base. The leaves are narrow, linear-lanceolate, 2–7-cm-long and generally hairy. The spike is lanceolate or narrow, ovoid to oblong, becoming suddenly narrow in the upper half, generally awned, and 2.5–7.0 (usually 3.5–5.5)-cm-long (excluding awns). The entire spike disarticulates at maturity and falls as a unit (umbrella-type dispersal unit). There are 4–6 spikelets (usually 5) in each spike. The lowest two (seldom 3) are elliptical and longer than the adjacent rachis segments, the upper spikelets are equally long or shorter than the adjacent rachis segment. The upper spikelets bear small grains. There are 2–4 (usually 3) basal rudimentary spikelets. The glumes of the lower spikelets are elliptical, 7–11-mm-long, with two 3–5-cm-long awns, one being much wider than the other, with a deep cleft between. Glumes on upper spikelets often have 3 awns, which are shorter than the awns of the lower spikelets. Lemmas have 2–3 awns, are more slender and shorter than glume awns, and are generally present in all spikelets. The caryopsis is free Fig. 9.4f.

Ae. columnaris has relatively little morphological variation involving spike size, number of fertile and rudimentary spikelets and awn development. Sometimes it is difficult to distinguish it from Ae. neglecta and Ae. recta bearing only two awns per glume. According to van Slageren (1994), the main differences between Ae. columnaris and Ae. neglecta are: Ae. columnaris glumes of the lower 2–3 spikelets are elliptic-oblong, the apex usually has 2 awns–one large, 1.5–2.5-mm-wide at the base and often bifurcating above, and one very small and linear, 1 mm or less at the base–its spike is ovoid in the lower part, and more linear in the upper part, with 3–4(-6) spikelets, all fertile. Ae. neglecta glumes of the lower, fertile spikelets are obovate-elliptical, the apex usually has 3 awns of equal length and width at the base, its spike is ovoid-ellipsoid and inflated in the lower part, then abruptly constricted and almost linear, with 3–6 spikelets, of which the upper 1–3 are sterile.

Ae. columnaris is a Mediterranean-Western Asiatic element, occurring mainly in Turkey and the western arc of the Fertile Crescent, but scattered in the eastern part of the arc as well. The area of distribution extends westwards to Crete and eastwards to Transcaucasia and northwestern Iran (rare). It is uncommon throughout its range. Ae. columnaris grows in Greece (including Crete), Turkey, Syria, East Lebanon, North Iraq (rare), Iran (rare), Armenia, and Azerbaijan. It is found as an adventive plant in southern France. Ae. columnaris is mainly found on limestone, less frequently on basalt or grey-calcareous steppe soils. The soil textures are predominantly stony, with additional clay, (clay)-loam and occasionally sand. It grows in areas with a range of annual rainfall 450–1250 mm, indicating that it prefers a generally wetter environment than most Aegilops species. It occurs in degraded deciduous oak forests, deciduous steppe maquis, degraded dwarf-shrub formations, open dwarf-shrub steppe-like formations, pastures, abandoned fields, edges of cultivation, disturbed areas and roadsides. It grows at altitudes of 450–1990 m.

Ae. columnaris has a medium-sized distribution in the central part of the distribution of the genus. It occupies mainly open, secondary habitats and usually grows in mixed stands with Ae. biuncialis, Ae. neglecta, Ae. triuncialis, Ae. cylindrica and other species, with which it introgresses, particularly with other U-subgenome allopolyploids. These mixed populations contained different types of intermediate forms; some of which are rich in hybrids and hybrid derivatives, while others contain only a small number of introgressed lines. Series of intermediates between Ae. neglecta and Ae. columnaris were sampled at the edges of cultivation, near Malatya, Turkey (Zohary and Feldman 1962), and intermediates between Ae. triuncialis and Ae. columnaris were collected west of Malatya, Turkey, at roadsides, and north of Ankara, central Turkey, at roadsides and edges of wheat fields (Feldman 1965a). Karagoz (2006) presented evidence of spontaneous hybridization between Ae. columnaris and domesticated wheat.

Ae. columnaris grows sympatrically with A. muticum, Ae. speltoides, Ae. caudata, Ae. umbellulata, Ae. tauschii, wild T. monococcum, T. urartu, wild T. timopheevii, Ae. neglecta, Ae. triuncialis, Ae. cylindrica, Ae. crassa, and Ae. juvenalis, and allopatrically with Ae. searsii, Ae. geniculata, Ae. kotschyi and Ae. peregrina.

Accessions of Ae. columnaris confer resistance to powdery mildew, leaf rust and Hessian Fly (Gill et al. 1985). Chromosome 5Xⁿ of Ae. columnaris is resistant to leaf rust (Badaeva et al. 2018). Several accessions of Ae. columnaris are tolerant to drought and frost (Damania and Pecetti 1990).

9.8.7.2 Cytology, Cytogenetics and Evolution

Ae. columnaris is an allotetraploid species (2n = 4x = 28). Its nuclear genome is designated UUXⁿXⁿ (similar to the genome of Ae. neglecta; modified from Dvorak 1998) and the plasmon genome is designated U’, a sub-type of the U plasmon of Ae. umbellulata (Tsunewaki 2009). The origin of Ae. columnaris is not yet well understood (Resta et al. 1996; Yen et al. 1996; Dvorak 1998). Originally, the nuclear genome of Ae. columnaris was designated by Kihara (1937, 1949, 1954) as C^uC^uM^cM^c (C^u is currently U and M^c is modified M), closely related to the genome of Ae. Umbellulata, based on indirect evidence, namely, on analysis of chromosome pairing in hybrids between Ae. columnaris and other allotetraploid species that had previously been carry the Ae. umbellulata genome (von Berg 1937; Kihara 1937, 1940a, b, 1949). A direct cross between Ae. columnaris and Ae. umbellulata was analyzed by Kimber and Abu-Bakar (1981), who observed 6.17 univalents, 1.87 rod bivalents, 4.30 ring bivalents, 6.17 total number of bivalents and 0.83 trivalents at first meiotic metaphase of this triploid hybrid (hybrid genome UXⁿU). This pairing data indicated that Ae. columnaris indeed contains one subgenome that is homologous to genome U of Ae. umbellulata. Analysis of chromosome pairing in hybrids between Ae. columnaris and an induced autotetraploid line of Ae. umbellulata again showed that one subgenome of Ae. columnaris is homologous to that of Ae. umbellulata (Kimber and Yen 1988, 1989). The conclusion that one subgenome of Ae. columnaris originated from Ae. umbellulata was later confirmed by several authors (e.g., Resta et al. 1996; Badaeva 2002; Badaeva et al. 2004, 2018). Both genomes have similar chromosome morphologies, distribution of 5S and 45S rDNA loci, labeling patterns of pSc119.2 and pAs1 probes and weak hybridization with the (GTT)₁₀ probe (Badaeva et al. 2018).

Evidence for multiple origins of the U subgenome of Ae. columnaris was obtained by Kadosumi et al. (2005). Using the genome-specific PCR primer set U31, they analyzed variation of the U genome in 48 accessions of Ae. columnaris and 72 accessions of its diploid progenitor Ae. umbellulata. Three alleles were distinguishable by the length of the amplified sequence, namely, allele I (normal size with an MspI site), allele II (normal size without an MspI site), and allele III (shorter size caused by a 123 bp deletion). Sequence comparison indicated the inheritance of alleles I and III from the diploid to the tetraploid, suggesting multiple origins of the U subgenome of the tetraploid. Regarding allele II, however, the sequence comparison indicated that parallel mutations at the MspI site produced allele II several times. The phylogenetic tree based on the sequences of the U31 region, demonstrated the presence of a third lineage of the U genome from Ae. umbellulata to Ae. columnaris. Consequently, Kadosumi et al. (2005) concluded that the Ae. columnaris U subgenome had at least three independent origins.

The origin of the second subgenome of Ae. columnaris has not been satisfactorily determined. Initially, the second genome was designated by Kihara (1937, 1949, 1954) as M^c, i.e., a modified M genome, since, based on morphological comparisons and meiotic analysis, Kihara assumed that this subgenome was closely related to the M genome of Ae. comosa or to the M^u genome of Ae. uniaristata. Yet, the hybrids between the diploid species Ae. comosa or Ae. uniaristata and Ae. columnaris displayed relatively low pairing, indicating that there was no homology between any of the subgenomes of Ae. columnaris and the genomes of these diploid species. The Ae. comosa x Ae. columnaris hybrid showed three to seven bivalents per first meiotic metaphase cell, of which only one was a ring bivalent, and the hybrid Ae. uniaristata x Ae. columnaris showed four to six rod bivalents per cell (Kihara 1949). To resolve this incongruity, Kihara ((1963) suggested that the incomplete pairing between the M subgenomes of Ae. columnaris with the M or the M^u genomes of the diploid species, resulted from either divergence of the M at the diploid level, followed by extinction of some of the diploid taxa, or genomic modification at the tetraploid level. But, while resemblance between one chromosome set of Ae. columnaris to that of Ae. umbellulata was observed, karyotype analysis failed to find a genome resembling to that of Ae. comosa or Ae. uniaristata in Ae. columnaris (Chennaveeraiah 1960). Moreover, Chennaveeraiah noted that no genome in the diploid species of the Aegilops-Triticum group matched the second subgenome of Ae. columnaris.

Resta et al. (1996) employed variation in randomly selected families of repeated nucleotide sequences to study the origin of the subgenomes of Ae. columnaris. They determined the fraction of marker bands of these repeated sequences of a diploid shared with a polyploid which they called repeated nucleotide sequence correspondence (RSC). RSC varies from 0.00, if no marker band of a diploid is encountered in a polyploid, to 1.00, if all are encountered. They found that fifteen marker bands of Ae. umbellulata were shared with Ae. columnaris (RSC = 0.88), and that two marker bands of Ae. speltoides (RSC = 0.05) and one marker band of Ae. caudata (RSC = 0.03) were found in Ae. columnaris. Thus, only the RSC of Ae. umbellulata with Ae. columnaris differed significantly from 0.00, indicating that Ae. columnaris has a subgenome similar to that of Ae. umbellulata. The analysis also showed that Ae. columnaris is closely related to Ae. neglecta. Both species share the U subgenome with Ae. geniculata and Ae. biuncialis, but their second subgenome differs from those of Ae. geniculata and Ae. biuncialis, in other words, is not a modified M subgenome. As concluded by Chennaveeraiah (1960), they found no relationship between the second subgenome of Ae. columnaris and Ae. neglecta and a genome of any extant diploid species of the Aegilops-Triticum group. Consequently, the second subgenome of Ae. columnaris and Ae. neglecta was designated X^t by Resta et al. (1996) and the proposed genome formula for Ae. columnaris and Ae. neglecta was UUX^tX^t (Resta et al. 1996; Dvorak 1998). This genomic formula was later modified to UUXⁿXⁿ for Ae. columnaris and for Ae. neglecta (Badaeva 2002; Badaeva et al. 2004).

Differences in C-banding patterns, and the distribution of 5S and 45S rDNA loci also contradicted the conclusion that the second subgenome of Ae. columnaris is a modified M genome (Badaeva 2002; Badaeva et al. 2004). Considering data from C-banding and FISH analyses, Badaeva (2002) and Badaeva et al. (2004) suggested that the putative progenitor of the Xⁿ subgenome should have a relatively symmetric karyotype, with highly heterochromatic chromosomes visualized by C-banding. In hybridization patterns of pSc119.2 and pAs1 probes, the progenitor species should be similar to S, T, U, or C genomes of diploid Aegilops. An abundance of (GTT)_n microsatellite repeats is an important diagnostic feature of the Xⁿ subgenome. The Xⁿ subgenome is also unique with respect to the number and distribution of 5S rDNA loci, because other diploid or polyploid species of Aegilops and Triticum. except Ae. neglecta, carry two or, rarely, one, 5S rDNA locus per haploid genome (Dvorak et al. 1989; Badaeva et al. 1996b). In all cases, these loci are located on different chromosomes. Ae. columnaris has four 5S rDNA loci, two of which are located in one chromosome. None of the extant diploid species of Aegilops fit the criteria listed above. Consequently, none of them could be the direct progenitor of the Xⁿ genome, which agrees with previous results (Resta et al. 1996; Dvořák 1998). The Xⁿ subgenome was probably derived from an extinct yet an undiscovered extant diploid species of Aegilops.

While Resta et al. (1996) detected considerable differences between Ae. geniculata and Ae. biuncialis, on one side, and Ae. columnaris and Ae. neglecta, on the other, almost no divergence was found between Ae. columnaris and Ae. neglecta. This close relationship between the genomes of the latter two species is reflected in the high chromosomal pairing in the hybrid Ae. neglecta x Ae. columnaris, i.e.,12–14 bivalents per cell with a mode of 12 bivalents, and in the 20% pollen viability (Kihara 1949). In contrast, the hybrid Ae. biuncialis x Ae columnaris showed a mode of seven to eight bivalents per cell (Kihara 1937), Ae. biuncialis x Ae neglecta showed a mode of eight bivalents (Kihara 1937), and Ae. geniculata x Ae. neglecta showed a mode of eight bivalents, ranging from 6–12 bivalents per cell (Percival 1932; Kihara 1937). Hence, the data of chromosome pairing and the studies of Resta et al. (1996) showed that the species within each group (Ae. geniculata and Ae. biuncialis vs. Ae. columnaris and Ae. neglecta) have two subgenomes in common, whereas the species between the groups have only one subgenome in common, i.e., subgenome U. This contradicts the conclusion of Kihara (1937, 1949, 1954, 1963) who assigned the same basic genome formula to all these four species.

According to Mukai and Tsunewaki (1975), Tsunewaki (1993, 1996, 1980), Tsunewaki and Tsujimoto (1983), and Ikeda and Tsunewaki (1996), the cytoplasm of Ae. columnaris causes variegation (variegated yellowing of leaf color) in midwinter and growth depression in most common wheat lines bearing Ae. columnaris cytoplasm. In addition, it causes haploidy and haplo-diplo twinning in two particular common wheats and complete male sterility in about half of the common wheats tested. Since these genetic effects are similar to those caused by the cytoplasm of Ae. umbellulata, Mukai and Tsunewaki (1975), Tsunewaki (1980) and Tsunewaki and Tsujimoto (1983), classified Ae. columnaris cytoplasm as C^u (currently U) plasma type and concluded that Ae. umbellulata as the cytoplasm donor to Ae. columnaris. Ogihara and Tsunewaki (1988) used restriction enzymes to investigate variation due to fragment size mutations in the chloroplast DNAs of these species and classified the chloroplast DNAs of the Triticum and Aegilops species. This classification was in agreement with that of the plasma types assigned according to phenotypes arising from nucleus-cytoplasm interactions. On the basis of these studies, Ogihara and Tsunewaki (1988) confirmed the Ae. umbellulata origin of the Ae. columnaris cytoplasm. However, based on studies of the phenotypes in cytoplasm-substituted wheats, Maan (1975, 1978) and Panayotov and Gotsov (1975, 1976) proposed that Ae. uniaristata or Ae. comosa was the cytoplasm donor to Ae. columnaris. Yet, the results of Terachi et al. (1984), Ogihara and Tsunewaki (1988) and Wang et al. (1997) unequivocally showed that the chloroplast genome of Ae. columnaris is nearly identical to that of Ae. umbellulata.

Terachi and Tsunewaki (1986) reported that the mitochondrial genome diversity in the allopolyploid species of the wheat group is far more extensive than the chloroplast genome diversity. In this respect, they found that of the mitochondrial DNAs isolated from the allotetraploids with U-type cytoplasms, that of Ae. columnaris was the most differentiated. Whereas the mitochondrial DNA of the other allotetraploids was almost identical to that of Ae. umbellulata, that of Ae. columnaris differed to some extent from that of Ae. umbellulata (Terachi and Tsunewaki 1986; Wang et al. 1997). In agreement with the above findings, Ikeda et al. (1994) and Ikeda and Tsunewaki (1996) reported that alloplasmic wheats having Ae. columnaris cytoplasm, that are characterized by growth inhibition, show impaired mitochondrial cytochrome c oxidase. Hence, Ae. columnaris differs only by a small number of mutations from that of Ae. umbellulata, whereas almost no mutations exist in the chloroplast genome of Ae. columnaris, the allotetraploid undoubtedly originated from Ae. umbellulata as a female (Tsunewaki 2009). Because of the mitochondrial changes the cytoplasm of Ae. columnaris was designated U’ (Tsunewaki 2009).

The relative times of origin of the four allopolyploid species bearing U cytoplasm, i.e., Ae. neglecta, Ae. columnaris, Ae. biuncialis, and Ae. triuncialis, were inferred from genetic distances between their cytoplasms and that of Ae. umbellulata, their putative maternal parent. The number of differences between the plasmon of these allotetraploids and that of Ae. umbellulata indicates that Ae. neglecta is older than Ae. columnaris, Ae. biuncialis and Ae. triuncialis (Tsunewaki 2009).

Ae. columnaris contains 10.86 pg 1C nuclear DNA (Eilam et al. 2008). Its genome, and that of is closest relative Ae. neglecta, are larger than the genomes of Ae. biuncialis, Ae. geniculata, and Ae. triuncialis and smaller than those of Ae. peregrina and Ae. kotschyi (Eilam et al. 2008). This may indicate that the genome of the, yet unknown, diploid donor of the subgenome Xⁿ to Ae. columnaris is larger than both the M genome of Ae. comosa, the putative M subgenome donor to Ae. biuncialis and Ae. geniculata and the C genome of Ae. caudata, the putative C subgenome donor to Ae. triuncialis, but smaller than the S^l genome of Ae. longissima, the putative S^l subgenome donor to Ae. peregrina and Ae. kotschyi.

The karyotype of Ae. columnaris has three pairs of chromosomes with satellites on short arms (Chennaveeraiah 1960). The satellites on two chromosome pairs are fairly large and those on the third pair are comparatively smaller. One chromosome pair with large and another pair with smaller satellites resemble those of Ae. umbellulata (Chennaveeraiah 1960). There is a single pair with the characteristic extreme subterminal centromere similar to that of Ae. umbellulata; all the other pairs have submedian centromeres. (Chennaveeraiah (1960). Thus, the karyotype of Ae. columnaris differs considerably from those of Ae. geniculata and Ae. biuncialis (Chennaveeraiah 1960).

Three pairs of SAT chromosomes were also found in Ae. columnaris by Teoh et al. (1983). Using FISH with a repetitive DNA sequence coding for rRNA, they noted the presence of three pairs of rRNA sites, agreeing with the number of SAT chromosomes pairs. NORs activity studied in somatic metaphase chromosomes of Ae. columnaris by phase contrast, C-banding and Ag-staining (Cermeño et al. 1984b), showed that the U subgenome suppressed the activity of one pair of NORs of subgenomes Xⁿ. In accord with the finding of Chennaveeraiah (1960) and Teoh et al. (1983), Cermeño et al. (1984b) found three pairs of SAT chromosomes as well as three pairs of active NORs. Of the active three NOR pairs, two pairs were of the U subgenome and only one pair of the Xⁿ subgenome. The Xⁿ NOR showed reduced activity as was evident by the small size of the Ag-NORs of this chromosome pair (Cermeño et al. 1984b).

The heterochromatin banding patterns of somatic metaphase chromosomes of Ae. columnaris, as revealed by C-banding, contained broad C-banding polymorphism and chromosomal rearrangements, represented by paracentric inversion and intra- and inter-subgenomic translocations (Badaeva (2002) and Badaeva et al. (2004). Significant differences in karyotype structure and total amount and distribution of C-heterochromatin were observed between Ae. columnaris and Ae. neglecta, on the one hand, and Ae. geniculata and Ae. biuncialis, on the other, evidence supporting claims of different origins of these two groups of species. In turn, similarity of the C-banding patterns of Ae. columnaris and Ae. neglecta chromosomes suggested that they were derived from a common ancestor (Badaeva (2002). Ae. umbellulata was shown to be the U-subgenome donor of Ae. columnaris and Ae. neglecta, but the donor of the second subgenome of these two species was not determined by the C-banding pattern. Badaeva (2002) assumed that the second subgenome of these two tetraploid species resulted from introgressive hybridization.

In addition, Badaeva et al. (2004) used FISH with the heterochromatin-limited repetitive DNA probes pSc119 and pAs1 to study the distribution of these hybridization sites, as well as the distribution of NOR and 5S DNA loci revealed by pTa71 (18S-26S rDNA) and pTa794, (5S rDNA), respectively. Similar to the finding of Resta et al. (1996) that differentiated between Ae. columnaris and Ae. neglecta, on one hand, and Ae. geniculata and Ae. biuncialis, on the other hand, the studies of Badaeva et al. (2004) revealed significant differences between these two groups of species in the total amount and distribution of heterochromatin, and in the number and location of 5S and 18S-26S rDNA loci, indicating that these two groups of species have different origins. Similarities in C-banding and FISH patterns of most Ae. columnaris and Ae. neglecta chromosomes suggest that they were probably derived from a common ancestor, whereas distinct differences of three chromosome pairs may indicate that the divergence of these species was probably associated with chromosomal rearrangements and/or introgressive hybridization (Badaeva et al. 2004). The contribution of the U subgenome to Ae. columnaris and Ae. neglecta by Ae. umbellulata was again confirmed, however, the source of their second genomes remains unknown. FISH with the probes pSc119 and pAs1 detected polymorphic hybridization patterns in Ae. columnaris. Large to medium PSc119 sites were observed in telomeric regions of one or both arms of 11 of 14 chromosome pairs, and interstitial sites were observed in the long arms of chromosomes 6U and 7U. FISH with pAs1 revealed signals of different intensities on several chromosomes.

Ae. columnaris is characterized by a unique distribution of the hybridization sites of the two ribosomal probes, pTa71 and pTa794. (Badaeva et al. 2004). Two major NORs were detected on chromosomes 1U and 5U. These chromosomes also had 5S rDNA sites distal and proximal to the major NORs. The third major NOR was detected in the short arm of chromosome 1Xⁿ, which also had two 5S rDNA loci; one distal and the other proximal to the NOR. Thus, Ae. columnaris has three major MORs and four 5S rDNA loci, two of which are located in one chromosome arm. Such distribution of 18S-26S and 5S rDNA loci has no analogs in diploid or polyploid Aegilops/Triticum species, except for Ae. neglecta (Badaeva et al. 1996b). No minor NORs were observed in Ae. columnaris. The genetic nomenclature of Ae. columnaris chromosomes have not been established, and consequently, Badaeva et al. (2004) tentatively assigned chromosomes to homoeologous groups and genomes on the basis of their similarity with chromosomes of other Aegilops species.

According to Badaeva (2002), of all allopolyploid Aegilops species, Ae. columnaris showed the highest level of intraspecific chromosome variation. Two major chromosomal types, A and B, differing in karyotype structure, in the number of SAT chromosomes, and in the total amount and distribution of heterochromatin, could be distinguished in this species (Badaeva 2002). Broad C-banding polymorphism was revealed within each chromosomal type. Chromosome type A was found in 13 of 17 Ae. columnaris accessions (Badaeva 2002). These accessions had three pairs of SAT chromosomes, two of which were assigned to the U and one to the Xⁿ subgenomes. The length of secondary constrictions on the satellite chromosomes of the U subgenome (1U and 5U) significantly exceeded the length of the secondary constriction on the Xⁿ subgenome. The type A chromosomes of Ae. columnaris showed significant polymorphism due to both variability of the C-banding patterns and chromosomal rearrangements. A paracentric inversion of chromosome 7U was the most common type of chromosomal rearrangements found in six out of 13 lines with type A chromosomes. Two unrelated accessions possessed centromeric translocation between chromosomes 1U and 5U, accompanied by inactivation of one NOR locus (Badaeva 2002).

Type B chromosomes were found in four accessions (Badaeva 2002). These accessions contained only two pairs of SAT chromosomes, i.e., they lost the satellite on chromosome 1U. At the same time, the secondary constriction on chromosome 1Xⁿ, which is poorly expressed in accessions of group A, was clearly extended. In addition, the accessions with type B chromosomes possessed less heterochromatin compared to accessions of group A. They also showed different morphology and C-banding patterns than those of seven chromosome pairs belonging to both U and Xⁿ subgenomes. The observed differences were so significant that Badaeva (2002) could not explain them by simple chromosome translocation and/or inversions. She thus suggested that intraspecific chromosome diversity of Ae. columnaris is accompanied not only by chromosomal aberrations, but, to a great extent, by introgression of genetic material from other species. It may be assumed that type B chromosomes derived from type A, since the latter are similar to the karyotype of Ae. neglecta.

Data from C-banding, FISH, nuclear and chloroplast (cp) DNA analyses, and gliadin electrophoresis, enabled Badaeva et al. (2021a) to confirm the division of Ae. columnaris into two distinctive groups, C-I (equivalent to group A) and C-II (equivalent to group B). C-I group was more similar to Ae. neglecta than C-II, and less polymorphic than C-II. Most C-II accessions were collected from a very narrow geographic region, and they might have originated from a common ancestor. The authors suggest that C-II emerged from C-I relatively recently, probably due to introgression from another Aegilops species, and might be at the initial stage of a speciation process.

Badaeva et al. (2018) reported the development of 57 common wheat-Ae. columnaris introgression lines covering 8 of the 14 Aegilops chromosomes. Based on the compensating capability in substitution lines and the results of FISH analysis of the parental Ae. columnaris line with seven DNA probes, Badaeva et al. (2018) determined the genetic nomenclature of 11 of 14 chromosomes. Each of these 11 Ae. columnaris chromosome was characterized on the basis of C-banding pattern and the distribution of the following seven DNA sequences: two wheat rDNAs pTa71, and pTa794, two microsatellite sequences (GAA)₁₀ and (CTT)_9, (GAA/CTT)_n), (GTT)₁₀, and three tandemly repeated DNA families, pSc119.2 pAs1, and pAesp_SAT86 (Badaeva et al. 2015).

GISH on Ae. columnaris chromosomes failed to clearly discriminate between genomes. Therefore, Badaeva et al. (2018) tested different probes to find appropriate genome-specific markers. Distribution of the (GAA/CTT)_n sequence was similar to the C-banding pattern but, by contrast, the (GTT)₁₀ probe showed different labelling patterns on chromosomes of the two Ae. columnaris subgenomes. Poorly labelled chromosomes were assigned to the U subgenome, based on their similarity with Ae. umbellulata chromosomes, which display weak hybridization with this sequence. Heavily labelled chromosomes were assigned to the Xⁿ subgenome.

9.8.7.3 Crosses with Other Species of the Wheat Group

Chromosome pairing at first meiotic metaphase of Ae. columnaris was studied by Cuñado (1992) using C-banding. The lower C-heterochromatin content of the U subgenome differentiated them from the chromosomes of the Xⁿ subgenome. Mean chromosomal pairing per cell included 0.03 univalents, 2.23 rod bivalents, 11.73 ring bivalents and 25.70 chiasmata. Thus, Ae. columnaris exhibits diploid-like meiotic behavior. Pairing between homologous chromosomes of the U subgenome included 0.03 univalents, 1.58 rod bivalents, 5.39 ring bivalents and 12.36 chiasmata, while the Xⁿ genome included 0.00 univalents, 0.65 rod bivalents, 6.34 ring bivalents and 13.34 chiasmata. The somewhat higher pairing between chromosomes of the Xⁿ subgenome than between chromosomes of the U subgenome may be due to higher C-heterochromatin content in the latter subgenome (Cuñado 1992).

The pattern of zygotene and pachytene pairing in Ae. columnaris was analyzed by whole-mount surface-spreading of synaptonemal complexes, as viewed under an electron microscope (Cuñado et al. 1996b). The data indicated that at the early meiotic zygotene and pachytene stages the chromosomes were almost exclusively associated as 14 bivalents; only rarely as 12 bivalents and one trivalent.

Data of chromosomal pairing in the F₁ hybrid Ae. columnaris x Ae. umbellulata (genome UXⁿU) showed that Ae. columnaris contains one subgenome that is homologous to the genome of Ae. umbellulata. In contrast, the triploid hybrids between Ae. columnaris and the M-and N-genome diploid species, Ae. comosa and Ae. uniaristata, showed a much lower level of pairing, indicating that none of the subgenomes of Ae. columnaris are homologous to the M or N genomes. More specifically, Ae. columnaris x Ae. comosa (genome UXⁿM) had 3–7 bivalents and 0–3 trivalents and Ae. columnaris x Ae. uniaristata (genome UXⁿN) had 4–6 bivalents and 0–1 trivalents (Kihara 1949). The triploid hybrid Ae. columnaris x Ae caudata (genome UXⁿC) showed somewhat higher pairing than the hybrids of Ae. columnaris with the M and N genome species. Kihara (1949) observed 7–8 bivalents and 0–3 trivalents in Ae. columnaris x Ae. caudata, while Kimber and Abu-Bakar (1981) observed 7.47 univalents, 5.06 rod bivalents, 0.14 ring bivalents, 5.20 bivalents, 0.99 trivalents and 0.04 quadrivalents. This level of pairing indicates that one subgenome of Ae. columnaris, most probably subgenome U, is closer to the C genome than to the M and N genomes. The triploid hybrid Ae. columnaris x Ae. speltoides (genome UXⁿS) showed 5–7 bivalents with a mode of 7 bivalents and 0–4 trivalents (Kihara 1949). In contrast, the triploid hybrid Ae. columnaris x Ae. bicornis (genome UXⁿS^b) had 1–7 bivalents and 0–2 trivalents (Kihara 1949). No homology between the subgenomes of Ae. columnaris and Ae. bicornis was observed, and the high pairing of the hybrid with Ae. speltoides resulted presumably from the ability of the speltoides genome to promote homoeologous pairing.

The Ae. neglecta x Ae. columnaris hybrid (genome UXⁿUXⁿ) had 11–14 bivalents, with a mode of 12 bivalents (Kihara 1949). This level of pairing shows that the genomes of the two species are closely related; both contain the U genome and a slightly modified Xⁿ genome. Chromosomal pairing of other hybrids between Ae. columnaris and allotetraploid species having the U-subgenome and differing in the second subgenomes, usually had more than seven bivalents, indicating that, in addition to pairing between the chromosomes of the shared U subgenome, some pairing also occurs between chromosomes of the differential, unshared subgenomes. Thus, chromosome pairing data between Ae. triuncialis and Ae. columnaris hybrid (genome UCUXⁿ) (6–12 bivalents, with a mode of 10 bivalents, and 0–2 trivalents) shown by Lindschau and Oehler (1936), indicate that there is one subgenome common to both species, whereas the second subgenome is different. The Ae. geniculata x Ae. columnaris hybrid (genome M^oUUXⁿ) had 7.15 bivalents, 1.10 trivalents and 0.30 quadrivalents (Kimber et al. 1988) or 6–9 bivalents and 0–3 trivalents (Kihara 1949). The Ae. columnaris x Ae. biuncialis hybrid (hybrid genome UXⁿUM^b) had 7–9 bivalents, with a mode of 8 bivalents, and 0–2 trivalents (Kihara and Nishiyama 1937). The Ae. columnaris x Ae. kotschyi hybrid (genome UXⁿS^vU) had 7.05 bivalents and 0.90 trivalents (Kimber et al. 1983). The pentaploid hybrid between the hexaploid species Ae. juvenalis and Ae. columnaris (genome D^cX^cUUXⁿ) had 7.81 bivalents, 1.79 trivalents and 0.22 quadrivalents (McGinnis and Melnyk 1962), indicating that the two species share only one subgenome, i.e., U, and differ in the other subgenomes. The subgenome X^c of Ae. juvenalis, like X^c of Ae. crassa, is different from the Xⁿ of Ae. columnaris; the symbol ‘Xⁿ’ means that the origin of this subgenome is unknown (Dvorak 1998).

9.8.8 Aegilops triuncialis L.

9.8.8.1 Morphological and Geographical Notes

Ae. triuncialis L., common name Barbed Goatgrass [Syn.: Triticum triunciale (L.) Raspail; Aegilopoides triuncialis (L.) Á. Löve; Ae. aristata Req. ex Bertol.; Ae. echinata C. Presl: Ae. persica Boiss.] is an annual, predominantly autogamous, tufted, many–tillered plant. Its culms are without many joints, usually prostrate and then turning upright, (15-)20–45(-60)-cm-tall (excluding spikes), and defoliated in the upper quarter. The leaves are usually hairy, narrow-linear and 2.0–6.0-cm-long. The spike is narrow-lanceolate, 2.5–6.0-cm-long (excluding awns), usually awned, tapering to the tip, and well above the flag leaf on a long peduncle. The entire spike disarticulates at maturity and falls as a unit (umbrella-type dispersal unit), seldom as individual spikelets, each with its adjacent rachis segment (barrel-type dispersal unit). The rachis internode is generally approximately as long as the adjacent spikelet. There are three, and seldom 2, basal rudimentary spikelets. There are 3–8 (usually 4–5) spikelets, narrowly elliptical, usually all of which are potentially fertile. Each spikelet contains 4 florets, with the upper 2 being sterile. Glumes feature curved, unequally wide nerves, often covered with short, silvery hairs. There are 3 glume awns, unequal in length, which are usually smooth underneath, gradually tapering to the tip. The central glume awn of lower spikelets is shorter than its laterals. The central awn of glumes of the terminal spikelet is longer (4.5–7.0-cm-long) and wider than its laterals and is often the longest awn on the spike, and diverges almost at a right angle to the spike axis. Lemma awns are poor developed or absent. The caryopsis is free (Fig. 9.4g).

Ae. triuncialis is a polymorphic species exhibiting very wide morphological variation that mainly involves awn development (presence or absence), length and count, spike color and hairiness. This large morphological variation led taxonomists to subdivide the species into several intraspecific taxa. Zhukovsky (1928) classified Ae. triuncialis in a separate section, Surculosa Zhuk., and divided it into two subspecies, brachyathera Boiss. and kotschyi Boiss. Eig (1929a) also divided Ae. triuncialis into two subspecies: eu-triuncialis and orientalis Eig, with subsp. eu-triuncialis containing two varieties: typical and constantinopolitana Eig, and subsp. orientalis containing three varieties: assyriaca Eig, persica (Boiss.) Eig, and var. anathera Hausskn. et Bornm. Hammer (1980) also divided the species into two subspecies: triuncialis and persica (Boiss.) Zhuk, with subsp. triuncialis containing three varieties: triuncialis, flavescens Popova, and constantinopolitana Eig, and subsp. persica containing three varieties: persica (Boiss.) Eig, assyriaca Eig, and anathera Hausskn. et Bornm. In contrast to the above taxonomists who divided Ae. triuncialis into subspecies, van Slageren (1994) divided Ae. triuncialis into two varieties: triuncialis and persica (Boiss) Eig. Hammer’s subdivision (1980) into two subspecies, triuncialis and persica, will be used in this book.

Ssp. triuncialis has lateral spikelets glumes with 2–3 well-developed, 1.5-cm-long awns, while apical glumes have a well-developed, 5–8-cm central awn which is the longest awn of the spike, as well as 1–3-cm-long lateral awns. The spike always falls entire at maturity. Ssp. persica lateral spikelets have glumes with 1 or 2 teeth and a short, up to 1.5-cm-long awn. Its apical glumes have a well-developed, 2–5-cm-long central awn, and 2 short, 1–2-cm-long lateral awns, or lateral awns reduced to teeth. Lemmas of the terminal spikelet are awn-free, while the glumes bear 1 (1–3) awn. The spike sometimes disarticulates into individual spikelets, each with its adjacent rachis segment (barrel-type dispersal unit).

A new subspecies of Aegilops triuncialis L., ssp. bozdagensis Cabi & Doğan, confined to Denizli, Acıpayam, Bozdağ in southwestern Anatolia, was recently described (Cabi et al. 2018). It differs from the other two subspecies of Ae. triuncialis, triuncialis and persica, by the unawned glumes of the lateral spikelets, the three, up to 0.5-cm-long glume apex, with the middle one shorter than the others. The distribution map and notes on the biogeography and ecology of this subspecies, as well as an identification key of the three subspecies of Ae. triuncialis, are provided in Cabi et al. (2018).

Intraspecific variation was also detected on the molecular level. Nakai (1981), studying banding patterns of esterase isozymes in Ae. triuncialis and its putative parental diploid species, Ae. umbellulata and Ae. caudata, reported that zymogram phenotypes of both parents were quite uniform, whereas seven zymogram phenotypes were found among the 260 lines of Ae. triuncialis examined. Of these seven phenotypes, one (phenotype 3) had all bands of both parents, while the other six phenotypes differed greatly from phenotype 3. Thus, the zymogram phenotype in which the isozymes of both parental species are present, was considered by Nakai (1981) to be the most primitive of the seven types. Whether the phenotypes other than type 3 were due to introgressive hybridization or chromosomal rearrangement, is yet not determined.

Likewise, a RAPD analysis with 21 primers revealed more than 80% polymorphism between any two accessions of Ae. triuncialis collected in central Asia and North Caucasia (Okuno et al. 1998). Similarly, Monte et al. (1999, 2001), using RAPD and AFLP DNA markers, detected high polymorphism among Spanish populations of Ae. triuncialis and Thomas and Bebell (2010), using RAPD and ISSR analyses, observed significant genetic variability among 13 accessions of Ae. triuncialis from Greece. In contrast, Goryunova et al. (2010), using RAPD analysis, observed relatively low intraspecific polymorphism among 23 accessions of Ae. triuncialis that were collected from several areas of the species range.

Ae. triuncialis is a widespread Mediterranean/Western Asiatic/circumboreal element, occurring all over southern Europe and the Near East, extending eastwards into central Asia, and well-represented along most of the entire Fertile Crescent arc. It grows in Morocco, Algeria, Portugal, Spain, France (including Corsica), Italy (including Sardinia and Sicily), Slovenia, Croatia, Serbia, Bosnia-Herzegovina, Bulgaria, North Macedonia, Albania, Greece (including the Aegean Islands), Turkey, Cyprus, Syria, Lebanon, Israel (north), Iraq, Iran, Ukraine, Southern Crimea, Ciscaucasia, Transcaucasia (Georgia, Armenia and Azerbaijan), Kuwait, Saudi Arabia, Turkmenistan, Kazakhstan, Uzbekistan, Tajikistan, Kyrgyzstan, Afghanistan, and Pakistan. Ae. triuncialis grows in open areas and degraded forest and maquis, dwarf-shrub formations, steppe-like formations, pastures, roadsides and other disturbed habitats. It is also present in edges and within cultivation such as olive groves, vineyards, fruit tree plantations, and cereal crops, such as barley and wheat (with which it may form natural hybrids). Ae. triuncialis is also found in steppes but not in deserts, and, more rarely, in humid pastures, river terraces, and even at the seaside, apparently tolerating saline conditions. It grows on a variety of soils such as Mediterranean terra rossa, basalt and sandstone. It is also found on clay-and sandy loam, (sandy) clay, and gravel, and more rarely on loess, pure sands, and marly soils. It grows at altitudes from sea level up to 2700 m, usually in the 150–1800 m range. Ae. triuncialis grows under broad amplitudes of annual rainfall, varying from 125 mm up to 1400 mm per annum. Most data are, however, from the range 350–700 mm.

Ae. triuncialis has very large distribution (the largest in the genus), which overlaps with almost all of the distribution of the genus, except for the southeastern corner of the Mediterranean basin. It is common throughout its range and the most massive species in its distribution area. It has a very wide ecological amplitude, occupying a very large number of primary and secondary habitats. As a typical colonizer species, Ae. triuncialis can be found in massive stands and dominate a vegetation. It usually grows in mixed stands with other allotetraploid Aegilops species, with which it may introgress. According to Kihara (1954), Ae. triuncialis ssp. triuncialis with its var. typica covers the whole area of the species, while var. constantinopolitana is found in a very limited locality. The three varieties of ssp. persica are mainly in the Asiatic part of the Aegilops area. Among them one variety, persica, has the largest area extending to Iran, Transcaspian region and Afghanistan, while var. anathera has the same distribution except for not occurring in Afghanistan; var. assyrica has a very small habitat in Assyria.

The two diploid parents of Ae. triuncialis, Ae. umbellulata and Ae. caudata, have massive contact throughout Greece, Turkey, Syria, and Iran, where hybridization between them could have occurred, yielding Ae. triuncialis (Kihara 1954). However, the present distribution of Ae. triuncialis is very wide, much wider than that of its two diploid parents.

Ae. triuncialis grows sympatrically with almost all the species of the wheat group, except for Ae. bicornis, Ae. sharonensis, Ae. longissima, Ae. kotschyi and Ae. vavilovii. Intermediates between Ae. triuncialis and other allotetraploid species sharing the U genome are quite common in such mixed populations. Zohary and Feldman (1962) and Feldman (1965a) described intermediates between Ae. triuncialis and Ae. biuncialis and intermediates between Ae. triuncialis and Ae. neglecta in several locations in Turkey, and intermediates between Ae. peregrina and Ae. triuncialis in northern Israel. There is also evidence of hybridizations and introgression between Ae. triuncialis and common wheat (Hegde and Waines 2004). Arrigo et al. (2011) detected substantial gene flow between wheat and Ae. triuncialis growing close to wheat fields. Zaharieva and Monneveux (2006) assessed the probability of introgression between wheat and Ae. triuncialis in Europe, in areas where Ae. triuncialis occurred near or within wheat fields, and found that hybridization of Ae. triuncialis with wheat was quite common. Parisod et al. (2013), using 12 EST-SSR markers mapped on wheat chromosomes, obtained evidence of gene flow between common wheat and Ae. triuncialis growing in proximity to cultivated fields. Loci from the A subgenome of wheat were significantly less introgressed than sequences from the two other subgenomes, indicating differential introgression into Ae. triuncialis.

Ae. triuncialis is found as an adventive in central and northern France, Germany, Switzerland, Belgium, the Netherland, and the UK (van Slageren 1994). It was introduced with domesticated wheat into the USA (California, Oregon, Nevada, the New England area, and Pennsylvania) in the twentieth century and became a troublesome weed mainly on rangeland in California and Pennsylvania (Hitchcock 1951). Over the last several decades, Ae. triuncialis has rapidly spread into many annual grasslands and serpentine soil sites within California (Rice et al. 2013). It is unclear whether genetic differentiation, phenotypic plasticity, or both have allowed this species to invade competitive (i.e., high productivity, non-serpentine, annual grassland) and edaphically stressful (i.e., low productivity serpentine) environments (Rice et al. 2013). Despite its significant presence in northern California and in Pennsylvania, there have been no records of hybrids forming between common wheat and Ae. triuncialis in the USA (Watanabe and Kawahara 1999; Hedge and Waines 2004).

Accessions of Ae. triuncialis confer resistance to powdery mildew (Gill et al. 1985). leaf rust (Gill et al. 1985; Aghaee-Sarbarzeh et al. 2002), and to Hessian fly (Gill et al. 1985; Martín-Sánchez et al. 2003). Ae. triuncialis was highly resistant to Spikstersh, French and Swedish populations of cereal cyst nematode (Romero et al. 1998), and accessions of Ae. triuncialis were highly resistant to barley yellow dwarf luteovirus (BYDV) (Makkouk et al. 1994). Damania and Pecetti (1990) reported that several accessions of Ae. triuncialis were tolerant of drought and frost.

9.8.8.2 Cytology, Cytogenetics and Evolution

Ae. triuncialis is an allotetraploid species (2n = 4x = 28). Its nuclear genome is designated UUCC and CCUU (Kimber and Tsunewaki 1988; Dvorak 1998) and the plasmon genome of some forms is designated U, similar to that of Ae. umbellulata, while others contain plasmon C’, a subtype of the cytoplasm of Ae. caudata (Tsunewaki 2009). The origin of the subgenomes of Ae. triuncialis remains an enigma (Kihara 1954). Karyotype studies reported by Senyaninova-Korchagina (1932) and by Chennaveeraiah (1960) reached considerably different conclusions. While they agreed on the occurrence in this species of a subgenome similar to that of Ae. umbellulata, they differed in the observed nature of the second subgenome. Senyaninova-Korchagina (1932) described the karyotype of Ae triuncialis ssp. persica as composed of two sets of chromosomes, namely, one similar to that of Ae. umbellulata and the other to that of Ae. caudata, and the karyotype of ssp. triuncialis as composed of an Ae. umbellulata–chromosome set and another one different from that of Ae. caudata. Based on this, Senyaninova-Korchagina (1932) separated ssp. persica from Ae. triuncialis and elevated it to the specific rank, Ae. persica Boiss. In contrast, Chennaveeraiah (1960) reported that Ae. triuncialis ssp. persica had one set of chromosomes which corresponded to the genome of Ae. umbellulata, and a second set, though resembled, in most respects, the genome of Ae. caudata, differed from it in in having only one pair of SAT chromosomes. In Ae. triuncialis ssp. triuncialis he found, again, one chromosome set corresponding to that of Ae. umbellulata, whereas the second set differed not only from the typical genome of Ae. caudata but also from that of ssp. persica. Thus, Chennaveeraiah (1960) concluded that the Ae. triuncialis complex possesses chromosomal sets similar to those of Ae. umbellulata and Ae. caudata, but that either the chromosome set derived from Ae. caudata had undergone significant modifications or some other genome is involved.

Kihara (1940b) pointed out that karyotype analysis and genome analysis may lead to different results and, based on genome analysis, he included Ae. persica in Ae. triuncialis (Kihara 1954). Genome analysis (reviewed in Kihara 1954) showed that Ae. triuncialis originated from hybridization between the two diploids Ae. umbellulata and Ae. caudata and consequently, assigned to it the genome formula C^uC^uCC (currently C^u is U). Only one cross combination, Ae. caudata x Ae. triuncialis, was successful, displaying 7 bivalents and 7 univalents at first meiotic metaphase, thus, indicating unequivocally that Ae. triuncialis possesses the C subgenome that derived from Ae. caudata (Kihara 1949).

Since attempts to produce the triploid hybrid Ae. umbellulata x Ae. triuncialis were unsuccessful, the artificially produced amphidiploid of Ae. caudata x Ae. umbellulata (genome CCUU) was employed to determine whether the second subgenome of Ae. triuncialis is homologous to that of Ae. umbellulata (Kihara and Kondo 1943). The resulting amphidiploid was morphologically similar to ssp. triuncialis, whereas its karyotype was identical with that of ssp. persica (Kihara 1954), as described by Senyaninova-Korchagina (1932). Both the synthetic amphidiploid and ssp. persica had the umbrella-type disarticulation, but at the same time, the ear was easily broken in a barrel-type fashion in the upper part of the spike, a feature that is seldom found in ssp. triuncialis (Kihara 1954).

Kihara and Kondo (1943) used their own synthetic amphidiploid, and that of Sears (1939), to study chromosome pairing in hybrids between the synthetic amphidiploid and Ae. triuncialis. Chromosome pairing in the F₁ hybrid between the amphidiploid and the two subspecies of Ae. triuncialis was almost regular; the hybrid ssp. triuncialis x the amphidiploid had 11–12 bivalents (mode 12) and its reciprocal had 10–13 bivalents (mode 12); the hybrid ssp. persica x the amphidiploid had 11–13 bivalents (mode 12) and its reciprocal had 11–14 bivalents (mode 12) (Kihara and Kondo 1943). Pollen and seed fertility of these hybrids were low when the amphidiploid (C cytoplasm) was the female parent and higher in the reciprocal crosses when the natural subspecies bearing the U cytoplasm was the female parent (Kihara and Kondo 1943). The almost complete pairing in the hybrids between the synthetic amphidiploid and the two subspecies of Ae. triuncialis confirmed the karyotype studies which showed that the second subgenome of Ae. triuncialis is very close or even almost identical to the U genome of Ae. umbellulata. Thus, from the observation on chromosome pairing and fertility of the F₁ hybrid, Kihara and Kondo (1943) concluded that the genome-types of the two subspecies of Ae. triuncialis, triuncialis and persica, are identical or almost identical.

Additional evidence that Ae. triuncialis contains the U subgenome was presented by Kimber and Yen (1988, 1989), who observed 8.00 univalents, 2.08 rod bivalents, 2.37 ring bivalents, 2.87 trivalents, 0.52 quadrivalents, 0.07 pentavalents, and 0.02 hexavalents in the hybrid Ae. triuncialis x an induced autotetraploid Ae. umbellulata (hybrid genome UUUC). Using the numerical methods of chromosome pairing, Kimber and Yen (1988, 1989) determined that this type of chromosomal pairing best fits the 3:1 model, namely, three U and one C, indicating that Ae. triuncialis indeed contains a subgenome homologous to genome U of Ae. umbellulata. Studies on variation of repeated nucleotide sequences also confirmed the presence of U and C subgenomes in Ae. triuncialis (Dubcovsky and Dvorak 1994). They found that the number of bands of the repeated nucleotide sequences shared between each diploid species and Ae. triuncialis, was significantly higher than those shared with any other diploid species.

Early cytogeneticists were of the opinion that Ae. triuncialis is nearly an autotetraploid species (von Berg 1931; Karpechenko and Sorokina 1929; Kihara 1929, 1937). This opinion was based on the fact that the hybrid between Secale cereale and Ae. triuncialis had 3–7 bivalents (or sometimes even 6–7), which were attributed to autosyndesis of the triuncialis chromosomes. This level of pairing led Kihara and Lilienfeld (1932) to assume that Ae. triuncialis is not an autotetraploid, but rather, a segmental allopolyploid that was formed at a time when genomes U and C of the parental diploid species had not been widely differentiated.

Currently, the accepted view is that Ae. triuncialis is a typical allopolyploid (Kihara 1954, 1963). This view also gained support from the analysis of chromosomal pairing at first meiotic metaphase of a haploid plant of Ae. triuncialis (Chapman and Miller 1977), which showed 9.17 univalents, 2.10 bivalents, 0.17 trivalents, and 0.03 quadrivalents in the haploid plant. This low level of pairing indicated that the two subgenomes of Ae. triuncialis are less closely related than previously suggested (Chapman and Miller 1977).

Several accessions of Ae. triuncialis have the U plasmon type that was contributed by Ae. umbellulata (Tsunewaki 1980; Tsunewaki and Tsujimoto 1983). The chloroplast genome of these accessions showed identical restriction fragment patterns to those of Ae. umbellulata (Ogihara and Tsunewaki 1982, 1983; Terachi et al. 1984). Restriction fragment patterns of mitochondrial DNA isolated from one accession of Ae. triuncialis, have been analyzed using five restriction endonucleases and confirmed the presence of U cytoplasm (Terachi and Tsunewaki 1986).

Yet, by comparing morphological and physiological characters of alloplasmic wheat lines bearing Ae. triuncialis cytoplasm, Mukai et al. (1978) found that two accessions of Ae. triuncialis ssp. triuncialis have genetically different cytoplasms. One had a cytoplasm almost identical to that of Ae. umbellulata and the other had one similar to that of Ae. caudata. This finding was supported by restriction endonuclease analysis of the chloroplast DNA of these accessions (Ogihara and Tsunewaki 1982, 1983). Together, these observations indicated a possible diphyletic origin of Ae. triuncialis from the reciprocal crosses between Ae. caudata and Ae. umbellulata (Ogihara and Tsunewaki 1982). Murai and Tsunewaki (1986), aiming to clarify (a) the extent of chloroplast DNA variation within Ae. triuncialis, (b) the geographical distribution of accessions bearing different chloroplast genomes, and (c) the distribution of each chloroplast genome type among different taxa, further studied the diphyletic origin of Ae. triuncialis. These authors, using restriction endonuclease analysis of chloroplast DNA of 21 accessions of Ae. triuncialis, found that 13 accessions had the type 2a chloroplast genome derived from Ae. caudata, eight possessed the type 3 chloroplast genome of Ae. umbellulata, and the remaining five contained a new chloroplast genome (named type 2b), which differed from the 2a type, by a 0·3 kbp insertion. The accessions with type 2a (caudata) and type 3 (umbellulata) chloroplast genomes distribute in wide geographical areas, and occur in both of its subspecies, triuncialis and persica, whereas those with the type 2b chloroplast genome occurred only in Azerbaijan. Waines and Barnhart (1992) mistakenly suggested that Ae. umbellulata is the female parent of ssp. triuncialis (the typical subspecies), while Ae. caudata is the female parent ssp. persica. From these results, Murai and Tsunewaki (1986) drew the following two conclusions: (a) Ae. triuncialis has a diphyletic origin from the reciprocal crosses between Ae. caudata and Ae. umbellulata, and (b) the type 2b chloroplast genome arose from type 2a chloroplast genome by a 0·3 kbp insertion.

Likewise, Ogihara and Tsunewaki (1988), studying restriction fragment patterns of chloroplast DNAs from two alloplasmic lines of common wheat carrying cytoplasm of two different accessions of Ae triuncialis, discovered that one accession (code no. 26) showed identical restriction fragment patterns to those of Ae. umbellulata, whereas the second accession (code no. 38) had patterns identical to that of Ae. caudata. This finding corroborated the proposal that Ae. triuncialis was produced diphyletically by the reciprocal crosses between Ae. caudata and Ae. umbellulata.

Wang et al. (1997) performed PCR–single-strand conformational polymorphism (PCR-SSCP) analyses of 14.0-kb chloroplast and 13.7-kb mitochondrial DNA regions that were isolated from alloplasmic wheat lines carrying Ae. triuncialis cytoplasm. Their study, showing that Ae. triuncialis contains two plasmon types, U and C², supported the claimed dimaternal origin of Ae. triuncialis from reciprocal crosses between Ae. umbellulata and Ae. caudata. The plasmon of these species pointed to zero genetic distance between Ae. triuncialis (line 26) and Ae. umbellulata, and a large distance (average 0.074) between Ae. triuncialis and Ae. caudata. On the other hand, Ae. triuncialis (line 38) was a short distance (average 0.019) from Ae. caudata as compared to Ae. umbellulata (0.081). If intraspecific variation of Ae. caudata is subtracted from the distance between Ae. triuncialis (38) and Ae. caudata, the distance between them is small, suggesting that both types of Ae. triuncialis arose recently and at approximately the same time. Since the C cytoplasm of Ae. triuncialis differs slightly from that of Ae. caudata, Tsunewaki (2009) designated it as C’ (a subset of C).

In accord with the above, Chee et al. (1995), using genome-specific primer sets, reported that variation in the C subgenome of Ae. triuncialis resulted from variability in Ae. caudata rather than from frequent introgression events after formation of the allopolyploid, thus, suggesting a polyphyletic origin for Ae. triuncialis. Meimberg et al. (2009) concluded that Ae. triuncialis was formed in four independent origins, affecting differences in ecological tolerance among the independent origins of this species. Similarly, Vanichanon et al. (2003) claimed that the number of independent origins of an allopolyploid species was traditionally underestimated. They screened 84 primer sets to identify genome-specific primer sets for Aegilops triuncialis and its diploid progenitors; some of the primers were U genome-specific and others C genome-specific. A DNA sequence comparison revealed at least two or three independent formations of Ae. triuncialis.

Ae. triuncialis contains 9.93 ± 0.041 pg 1C DNA (Eilam et al. 2008), the smallest genome among the allotetraploid species of section Aegilops and, together with Ae. cylindrica, the smallest genome among all the allotetraploid species of the wheat group (Eilam et al. 2008). The 1C DNA content of Ae. triuncialis is 2.84% smaller than that which is expected from the sum of the DNA of its two diploid parents, namely, 10.22 pg (Ae. umbellulata, the donor of the U subgenome, contains 5.38 pg and Ae. caudata, the donor of the C subgenome, contains 4.84 pg; Eilam et al. 2007). The loss of DNA in Ae. triuncialis was also noted by Badaeva (2002) and Badaeva et al. (2004) who, based on differential C-banding and FISH, found that Ae. triuncialis exhibits substantial structural chromosome rearrangements, including deletion of chromosomal segments and reduction of heterochromatin content.

Early karyomorphological studies in Ae. triuncialis (Emme 1924; Sorokina 1928) did not show any presence of SAT chromosomes. According to Senyaninova-Korchagina (1930, 1932) the karyotype consists of 9 types of chromosomes, including only one pair with a satellite on a short arm, one pair with an extreme subterminal centromere, two pairs with submedian-subterminal centromeres and the rest with submedian centromeres. Chennaveeraiah (1960) analyzed the karyotype of the two subspecies of Ae. triuncialis and found marked differences between that described by Senyaninova-Korchagina (1930, 1932). The karyotype of ssp. persica studied by Chennaveeraiah (1960), was in agreement with that described by Senyaninova-Korchagina (1930, 1932) for this subspecies. However, Chennaveeraiah (1960) reported the existence of three chromosome pairs with satellites on short arms, with the satellites differ from each other in size. In addition, there are four pairs with extreme subterminal centromeres, two pairs have submedian-subterminal centromeres, and the rest with submedian centromeres. In all, there are 12 types of chromosomes (Chennaveeraiah 1960). Ssp. triuncialis (brachyathera Boiss. in Chennaveeraiah) has a different karyotype in that it has only two pairs with satellites on short arms, only three pairs with extreme subterminal centromeres (whereas there are four in ssp. persica), three pairs with submedian-subterminal centromeres, and the rest with submedian centromeres. In all, there are 12 types of chromosomes in this subspecies as well (Chennaveeraiah 1960). Because of the diversity of morphological traits within the species, it was suggested that different accessions of Ae. triuncialis show different types of karyotypes (Chennaveeraiah 1960).

According to Chennaveeraiah (1960), Ae. triuncialis ssp. persica has one set of chromosomes which corresponds to the U genome of Ae. umbellulata. The second set, which in most respects resembles the C genome of Ae. caudata, differs in that it has a only one pair with satellites. In Ae. triuncialis ssp. triuncialis, one set corresponds to the U genome, whereas the second set differs not only from the typical C genome set but also from that of ssp. persica. In this case, the second set does not even contain a single satellited pair. From this it appears that the C genome underwent tremendous modification.

In the light of the present knowledge of the karyotypes of these subspecies, the synthesized amphidiploid of caudata-umbellulata (Kihara and Kondo 1943) is expected to have 4 pairs with satellites, as both C and U genome sets have two SAT-pairs each. Yet, ssp. persica has three SAT chromosomes and ssp. triuncialis has two. In spite of this and other differences, Chennaveeraiah (1960) believed in the presence of both the C and the U subgenomes in the triuncialis complex, but in some, either the C genome had undergone significant modification or there was some involvement of a foreign genome instead of the C genome. The U genome has remained unchanged.

Teoh et al. (1983), using FISH with a cloned repetitive DNA sequence derived from common wheat coding for ribosomal RNA, observed three SAT pairs (like the report above), and three pairs of rRNA sites on the genome of Ae. triuncialis. On the other hand, two SAT chromosomes were observed in Ae. triuncialis by phase contrast of metaphase cells (Cermeño et al. 1984b), corresponding to the report by Chennaveeraiah (1960) for ssp. triuncialis. This number is in quite good agreement with the two Ag-NORs (2) and maximum number of four nucleoli observed by Cermeño et al. (1984b). Apparently, the U subgenome fully suppresses NOR activity of the C subgenome in Ae. triuncialis (Cermeño et al. 1984b).

In accord with the above, Al-Mashhadani et al. (1980) found two pairs of SAT chromosomes in Ae. triuncialis. They also reported the presence of four metacentric pairs, three submetacentric pairs and five subtelocentric pairs, whereas Tanaka and Matsumoto (1965) found four metacentric chromosome pairs, three submetacentric pairs and seven subtelocentric pairs. Ahmadabadi et al. (2002), analyzing 13 populations of Ae. triuncialis, collected from northwest regions of Iran, found that karyological characters such as total length of chromosomes, chromosome arm ratio, number and length of satellites, showed high variation. Sadeghian et al. (2015) also recorded intraspecific karyotype divergence in Ae. triuncialis from northwestern Iran. Studying samples of Ae. triuncialis from different eco-geographical sites in northern Algeria, Baik et al. (2017) revealed two cytotypes which differed in chromosome lengths, karyotype symmetry and in the presence or absence of the satellites. One cytotype had 7 metacentric, 4 submetacentric, and 3 subtelocentric pairs, whereas the second cytotype had 9 metacentric, 11 submetacentric, and 2 subtelocentric pairs. One cytotype, consisting larger chromosomes, was sampled from populations at low altitudes under a humid bioclimate, whereas the second cytotype, having smaller chromosomes, was found in populations of high steppe plains under a semi-arid bioclimate.

Badaeva et al. (2004) examined karyotypes of 21 accessions of Ae. triuncialis (13 of ssp. triuncialis and 8 of ssp. persica) by C-banding and of four accessions (three of ssp. triuncialis and one of ssp. persica) by FISH with the pSc119, pAs1, pTa71 (18S-26S rDNA), and pTa794 (5S rDNA) probes. The karyotype structure of the accessions studied was similar to that described by Chennaveeraiah (1960) for Ae. triuncialis var. persica. All Ae. triuncialis chromosomes had distinct C-banding patterns, similar to those of the parental species Ae. umbellulata and Ae. caudata that were described by Friebe et al. (1992a, b, 1995a, c) and Badaeva et al. (1996a). Only limited intraspecific polymorphism was observed, manifested by the presence or absence of certain bands and variation in C-band size. Chromosomal arrangements found in three of the 21 accessions were represented by single or multiple translocation breakpoints and by a paracentric inversion. The origin of additional modified chromosomes (chromosome 1U in line k-940; chromosome 5U in lines k-146 and k-1965) remains unknown. FISH with pSc119 revealed distinct hybridization sites in the telomeric regions of one or both arms of 12 chromosome pairs of Ae. triuncialis. Interstitial FISH sites were observed in the long arms of chromosomes 7U and one C genome chromosome. Comparison of the pSc119 labeling patterns with the parental species showed that both the number and intensity of hybridization signals on Ae. triuncialis chromosomes decreased.

An extended secondary constriction was always observed on chromosome 1C, whereas the satellites on the 1U-subgenome chromosomes were usually not visible. Two active NORs were observed using FISH, confirming previous observations. The signal size decreased in the order 1C > 5U > 1U > > 5C. A faint signal was occasionally observed in the middle of the long arm of chromosome 6U. Four approximately equal 5S rDNA sites were detected in all chromosomes of homoeologous groups 1 and 5. Thus, the distribution of ribosomal RNA gene families on Ae. triuncialis chromosomes is similar to those of the parental species (see Badaeva et al.1996b), suggesting that speciation in Ae. triuncialis was not associated with large genomic modifications. The low C-banding polymorphism and the low frequency of chromosomal rearrangements suggest a relatively recent origin of Ae. triuncialis. The study of Badaeva et al. (2004), while confirming the origin of the two subgenomes in Ae. triuncialis, also detected a certain level of modification of the C subgenome in this species.

Different repetitive sequences, namely, pSc119.2–1, pTa535-1, pAs1-1, (CTT)₁₀ and the 45S rDNA clone from wheat (pTa71), were hybridized to chromosomes of Ae. triuncialis and compared to the patterns of its diploid progenitors Ae. umbellulata and Ae. caudata (Mirzaghaderi et al. (2014). Like Badaeva et al. (2004), also Mirzaghaderi et al. (2014) found that the FISH patterns of the U and C subgenomes of Ae. triuncialis were, in general, similar to those of U and C genomes of Ae. umbellulata and Ae. caudata, respectively, although some differences were observed.

In contrast, Gong et al. (2006), using 31 ISSR primers, found that subgenome U showed few alterations, while subgenome C had undergone changes after allopolyploidization. Compared with their ancestral diploid genomes, the genetic similarity index between Ae. triuncialis and Ae. umbellulata was 0.6842, which was higher than that between Ae. triuncialis and Ae. caudata.

9.8.8.3 Crosses with Other Species of the Wheat Group

Kihara and Kondo (1943) reported that Ae. triuncialis ssp. triuncialis showed regular pairing (14 bivalents) at first meiotic metaphase. Fertility of this subspecies was high (94.8% pollen fertility and 88.4% seed fertility). Similar data were obtained in ssp. persica, namely, 14 bivalents, 96.7% pollen fertility and 92.1% seed fertility. Cuñado (1992), using C-banding, studied metaphase-I chromosome associations in Ae. triuncialis and observed 0.09 univalents, 5.45 rod bivalents and 8.46 ring bivalents, culminating to a mean 22.36 chromosome associations per cell. The pattern of zygotene and pachytene pairing in Ae. triuncialis, analyzed by whole-mount surface-spreading of synaptonemal complexes under the electron microscope (Cuñado et al. 1996b), indicated that the chromosomes were exclusively associated as bivalents at the zygotene stage, while at pachytene, one multivalent was observed in 7% of the cells.

Chromosomal pairing in the hybrid between Ae. triuncialis and one of its two diploid parents, Ae. caudata (genome UCC), included 6–7 bivalents, with a mode of 7, and 0–2 trivalents (Kihara 1949), indicating that Ae. triuncialis possesses one subgenome, C, that derived from Ae. caudata. On the other hand, the hybrid between Ae. triuncialis and another diploid species, Ae. speltoides (genome UCS) had only 4–7 bivalents (Kihara 1949). The lower pairing in the latter hybrid, in spite of the presence of Ae. speltoides genes promoting homoeologous pairing, showed that none of the subgenomes of Ae. triuncialis are close to the genome of Ae. speltoides.

Chromosome pairing in hybrids between Ae. triuncialis and other allotetraploid species sharing the U subgenome but differing in the second subgenome, showed that, in addition to pairing between chromosomes of the U subgenomes, some pairing also occurred between chromosomes of the differential subgenome. More specifically, the hybrid Ae. neglecta x Ae. triuncialis (genome UXⁿUC) had 0–9 bivalents, with a mode of 5, and 0–1 trivalents (Lindschau and Oehler 1936), or up to 9 bivalents (Kihara 1949). The hybrid Ae. geniculata x Ae. triuncialis (genome M^oUUC) had 5–11 bivalents (Kihara 1929), and the hybrid Ae. triuncialis x Ae columnaris (h genome (UCUXⁿ) had 6–12 bivalents, with a mode of 10, and 0–2 trivalents (Lindschau and Oehler 1936). Chromosome pairing between the hexaploid species bearing the U subgenome, Ae. juvenalis, and Ae. triuncialis (genome D^cX^cUUC) was 12.94 univalents, 5.37 rod bivalents, 2.27 ring bivalents (total 7.64 bivalents), 1.70 trivalents, 0.34 quadrivalents and 0.02 heptavalents (McGinnis and Melnyk 1962). This level of chromosome pairing showed the homology between the U subgenomes as well as some pairing between chromosomes of the differential subgenomes. Chromosome pairing with Ae. cylindrica, which shares the C subgenome with Ae. triuncialis (genome DCUC), included 3–12 bivalents (Percival 1930), indicating that up to three bivalents were formed between the chromosomes of the D and the U subgenomes.

Chromosome pairing in hybrids between Ae. triuncialis and several subspecies of tetraploid wheat, Triticum turgidum (genome UCBA), was very low, indicating no homology between the subgenomes of these species. More specifically, the hybrid Ae. triuncialis x ssp. dicoccoides had 0–7 bivalents (usually 2–4) (Kihara 1929) and 1–3 rod bivalents (Percival 1930). The hybrid Ae. triuncialis x ssp. dicoccon had 1–7 rod bivalents, usually 4–5 (Kihara 1929). The hybrid Ae. triuncialis x ssp. durum had 0–8 rod bivalents, usually 6 (Kihara 1929), and 1–6 bivalents (Percival 1930). The hybrid Ae. triuncialis x ssp. polonicum had 3–8 bivalents, usually 5–6 (Kihara 1929), and the hybrid Ae. triuncialis x subsp. turgidum had 1–3 rod bivalents (Percival 1930).

Cifuentes et al. (2010) studied chromosome pairing at meiotic first metaphase in hybrids between T. turgidum ssp. durum and Ae. triuncialis, using a genomic in situ hybridization (GISH) procedure that allows simultaneous discrimination between A, B and U, C subgenomes. Chromosome pairing in the hybrid presented 25.6 univalents, 1.16 rod bivalents, 0.002 ring bivalents and 0.024 trivalents, culminating to 1.21 associations/cell. The general picture that was drawn showed that A and B wheat subgenomes paired with each other less than U and C did in the examined hybrid. Interspecific wheat-triuncialis (AB-UC) pairing accounted for 57% of the total pairing in the hybrid; the A subgenome was always the wheat partner most frequently involved in pairing with the Ae. triuncialis homoeologues. Pairing between the U and C chromosomes in durum x Ae. triuncialis showed 0.55 bivalents/cell (45.8%), whereas pairing between chromosomes of subgenome A of durum and UC of triuncialis included 0.49 bivalents/cell (40.8) and that of subgenome B chromosomes and triuncialis 0.08 bivalents/cell (6.67%) (Cifuentes et al. 2010). Pairing between A and B chromosomes of durum was at an incidence of 0.08 bivalents/cell (6.67%). Evidently, there was more autosyndetic than allosyndetic pairing between chromosomes of Ae. triuncialis. These results support the suggestion that U and C subgenomes show a higher pairing affinity for each other, and for the wheat A genome, than any of the other pairs of the constituent subgenomes present in the hybrid. Thus, the U and C subgenomes of Ae. triuncialis are more closely related than any combination of the other subgenomes present in this hybrid.

Chromosome pairing in hybrids between hexaploid wheat, T. aestivum, and Ae. triuncialis (genome BADUC) presented 26.97 univalents, 3.77 bivalents, and 0.17 trivalents (Riley 1966a), indicating that no homology exists between the subgenomes of these species. A similarly low level of pairing was observed in this hybrid combination by Aase (1930) [0–3 bivalents] and by Kihara (1929) [0–5 bivalents, usually 1–3].

9.8.9 Aegilops kotschyi Boiss.

9.8.9.1 Morphological and Geographical Notes

Ae. kotschyi Boiss. [Syn.: Ae. triuncialis L. var. kotschyi (Boiss.) Boiss.; Ae. triuncialis ssp. kotschyi (Boiss.) Zhuk.; Triticum kotschyi (Boiss.) Bowden; Aegilemma kotschyi Á. Löve] is an annual, predominantly autogamous, tufted, 15–30(-40)-cm-tall (excluding spikes) plant. It is usually bushy, with many tillers, of which the upper parts are upright. The leaves are usually glabrous. The spike is narrow-lanceolate to ovate-lanceolate, 2.0–3.0(-4)-cm-long (excluding awns), which narrows toward the tip, and is awned. The entire spike disarticulates at maturity (umbrella-type dispersal unit). There are (3-)4–5(-6) spikelets, usually linear and narrow, appressed to the rachis or to each other, with the two lower spikelets usually longer than the adjacent rachis segment. There are 2–4 (usually 3) basal rudimentary spikelets. There are 3–4 florets in the two lower spikelets, 2–3 of which are fertile; the upper spikelets are only partly fertile with small seed. The glume is usually shorter than the lemma, with narrow, equally wide, parallel nerves. There are usually 3, glume awns, and when there are 2, the central awn is replaced with a tooth or a gap. Glume awns are flat, gradually tapering to the tip. There are 1–3 lemma awns, which are equal to or slightly shorter than glume awns. In total, there are 8–14 awns on the spike, which tend to spread at maturity. The caryopsis adheres to the lemma and palea (Fig. 9.4h).

The original specimen of this species was collected by the botanist Kotschy in Iran. Boissier first described it as a species in 1846, but later, in 1884, he changed his mind about this species and listed it as a variety of Ae. triuncialis L. (Eig 1929a). Indeed, Ae. kotschyi shows some morphological similarity with Ae. triuncialis, and consequently was also treated by Zhukovsky (1928) as a subspecies of the latter, namely, Ae. triuncialis L. ssp. kotschyi (Boiss.) Zhuk. However, according to Eig (1929a), Boissier erred by classifying Ae. kotschyi as a variety of Ae. triuncialis.

Genome analysis studies led Kihara (1937, 1949, 1954) designated the genome of Ae. triuncialis as C^uC^uCC and that of Ae. kotschyi as C^uC^uS^vS^v, indicating that these two allotetraploids are separate species. Moreover, Ae. triuncialis and Ae. kotschyi are distinguished by their acid phosphatase isoenzyme patterns (Jaaska 1978a). Ae. triuncialis has the acid phosphatase isoenzyme doublet characteristic of Ae. caudata (genome C), whereas this isoenzyme was not observed in Ae. kotschyi. These results, like those of Kihara (1949, 1954, 1937), show the absence of the C genome in the latter, implying that Ae. kotschyi differs from Ae. triuncialis in its genome composition and should be treated taxonomically as an independent species.

Based on almost complete chromosome pairing in interspecific F₁ hybrids between Ae. kotschyi and Ae. peregrina (Lindschau and Oehler 1936; Kihara 1937), Kihara (1954, 1957) considered these two species very closely related and therefore, included Ae. kotschyi in Ae. peregrina. While the inclusion of Ae. kotschyi in Ae. peregrina was accepted by some authors, e.g., Bowden (1959) and Morris and Sears (1967), that followed Bowden’ classification, included Ae. peregrina in Ae. kotschyi. However, other wheat scientists treated them as separate species (e.g., Chennaveeraiah 1960; Furuta 1981b; Kimber and Feldman 1987; Waines and Burnhart 1992). On taxonomical grounds, the taxonomists Eig (1929a), Hammer (1980) and van Slageren (1994) considered them separate species.

The two species are vicarious taxa that grow in two different phytogeographical regions; Ae. kotschyi is an Irano-Turanian element and Ae. peregrina is a Mediterranean element. The two species also differ by the following morphological characters: Ae. kotschyi has linear, appressed spikelets and both glumes of the lowest fertile spikelet always have 3 fine awns that are equally wide at the base. The awns of all glumes and lemmas being more or less of the same length in Ae. kotschyi. In Ae. peregrina, the spikelets are urn-shaped to elliptical and not appressed. The glumes are coarse, one glume of the lowest fertile spikelet has 3 awns that are equally wide at the base, the other glume has only 2 awns that are unequally wide at the base. The lemma awns are poorly developed and often absent. The spike is stout, with an irregular appearance, caused by variation in glume and lemma awn development. Glume awns are 2–4-cm-long, and lemmas have one or two 0.3–3-cm-long awns, which are always shorter than the glume awns and with 1 or 2 teeth.

Hammer (1980) followed Eig (1929a) in recognizing the following five varieties in Ae. kotschyi, var. kotschyi (=var. tipica Eig), var. leptostachys (Bornm.) Eig, var. palaestina Eig, var. caucasica Eig, and var. hirta Eig. Yet, in part four of Flora Palaestina, Feinbrun-Dothan (1986b) recognized only two varieties in Ae. kotschyi, var. kotschyi and var. brachyathera. The spike of var. kotschyi is oval-lanceolate to lanceolate in outline, and awned. It commonly grows in steppes. The spike of var. brachyathera is narrow-lanceolate; all spikelets are awnless, sometimes with the exception of the upper one, which is shortly and irregularly awned. The variety is rare and grows in Nubian sandstone hills together with var. kotschyi.

Ae. kotschyi has some morphological variation involving spike shape (ratio of length to width), compactness due to rachis segment length, spikelet size, awn length and development. Eig (1929a) did not encounter a form that can be described as a subspecies. Of note, Ae. kotschyi is fairly uniform in Israel and surrounding countries and seems to be so also in other parts of its area (Feinbrun –Dothan 1986a; b). The predominant taxon in all the distribution area is var. kotschyi.

Goryunova et al. (2010) used RAPD analysis to study genetic variation in 15 accessions of Ae. kotschyi and its phylogenetic relationships to other allopolyploid Aegilops species having the U subgenome. The majority of Ae. kotschyi accessions displayed a moderate variation; two accessions however, were distinct from the other accessions. The maximum genetic distance between Ae. kotschyi accessions was 0.20. The greatest separation within the U-subgenome cluster was observed for the US genome species Ae. kotschyi and Ae. peregrina. This result is in agreement with Eig (1929a), who noticed that these two species differ morphologically from the other U-subgenome species. Based on their morphological characters, Eig (1929a) isolated Ae. kotschyi and Ae. peregrina into a separate subsection, Adhaerens, in which the caryopsis is fused to floral bracts, while the other U-subgenome species were assigned to subsection Libera, in which the caryopsis is free.

The study reported by Zhang et al. (1992) indicated that repeated nucleotide sequence families were more variable in Ae. peregrina than in Ae. kotschyi . This agrees with the morphological studies of Eig (1929a), which also indicated greater variation in Ae. peregrina than in Ae. kotschyi .

Ae. kotschyi occurs mainly along the coast of eastern North Africa and the western arc of the Fertile Crescent but is rarer and displays a scattered presence in the central and eastern parts of the Fertile Crescent and central Asia. It grows in Uzbekistan, Turkmenistan, Afghanistan, Pakistan, southern Iran, Iraq (Mesopotamia), Kuwait, eastern Saudi Arabia, Azerbaijan, southeastern Turkey, Syria, Lebanon, Cyprus (rarely), Israel, Jordan, Egypt (lower Egypt and Sinai), Libya, and Tunisia. Ae. kotschyi grows on grey-calcareous steppe soil, white rendzina, loess, sandy-clay and sandy soils, in deciduous steppe maquis, in dwarf-shrub steppe-like formations, marginal dwarf-shrub formations, steppical plains, wadis, edges of cultivation, disturbed habitats and roadsides. It is very common and locally abundant, forming very dense populations in open disturbed habitats of the warm steppes. It is sporadic in primary, stable habitats. It grows at altitudes of 100–1100 m. Ae. kotschyi is one of the few Aegilops species that clearly extends into the Saharo-Arabian region.

Ae. kotschyi has a relatively large distribution in the south, central and eastern part of the distribution of the genus. It is a steppical (Irano-Turanian) element, penetrating into the desert (Saharo-Arabian) region. It is the southern-most species at many sites. In various parts of its distribution, it grows sympatrically with Ae. longissima, one of the putative donors of the S subgenome. In other parts of its distribution, Ae. kotschyi has mainly allopatric contact with Ae. umbellulata, the donor of its second subgenome. As its putative parents, Ae. longissima and Ae. umbellulata, have sporadic contact in semi-steppical, steppical or sub-Mediterranean regions of Syria, it is possible that this region was the center of origin of Ae. kotschyi, from which it spread both south- and eastward. Currently, Ae. kotschyi grows sympatrically in some sites with Ae. bicornis, Ae. longissima, tetraploid Ae. crassa, and Ae. vavilovii and allopatrically with Ae. searsii, Ae. caudata, Ae. umbellulata, Ae. geniculata, Ae. biuncialis, Ae. triuncialis, Ae. peregrina and Ae. tauschii.

The Mediterranean Ae. peregrina and the more arid species, Ae. kotschyi, come in contact in the southern coastal plain of Israel, in the border area between the steppe of the Negev and the Mediterranean territory. In this border belt, stands of both species are commonly in contact, and hybrids and hybrid derivatives were readily detected in several such mixed stands (Zohary and Feldman 1962; Feldman 1963). Triploid hybrids and hybrid derivatives between Ae. kotschyi and Ae. longissima were found in several site in the northern Israeli Negev (Feldman M, unpublished), indicating the possibility of gene flow from a diploid species to the allotetraploid. Similarly, spontaneous, introgression and stabilization of a DNA sequence from Ae. searsii into Ae. kotschyi was described by Weissman et al. (2005).

Ae. kotschyi contains gene(s) conferring resistance to leaf and stripe rust (Marais et al. 2005) and several accessions of this allotetraploid were resistant to widely virulent races of stem rust (Scott et al. 2014). Ae. kotschyi accumulates high concentrations of iron and zinc in its grains and it is a promising source for increasing the amount of these chemical elements in the grains of common wheat (Chhuneja et al. 2006). It is assumed that Ae. kotschyi contains genes for drought, heat, and salt tolerance (Kimber and Feldman 1987).

9.8.9.2 Cytology, Cytogenetics and Evolution

Ae. kotschyi is an allotetraploid species (2n = 4x = 28). Its nuclear genome was designated C^uC^uS^vS^v (Kihara 1963; Kihara and Tanaka 1970) and its plasmon genome as S^v (identical to the plasmon of Ae. searsii) (Tsunewaki 2009). Subgenome C^u is currently designated U and subgenome S^v is closely related to genome S^l of Ae. longissima and genome S^sh of Ae. sharonensis (Zhang et al. 1992; Dvorak 1998). Since the plasmon of Ae. kotschyi derived from the S^v donor, the genomic formula of this species is S^vS^vUU. Genome analyses reported the formation of about seven bivalents in the F₁ hybrid Ae . kotschyi x Ae. umbellulata (von Berg 1937; Kimber and Abu-Bakar 1981) and approximately seven bivalents in the hybrid between Ae. longissima and Ae. kotschyi (Kihara 1949). On the basis of these data, Kihara (1949) concluded that Ae. kotschyi contains the C^u (=U) subgenome from Ae. umbellulata and the S subgenome from a Sitopsis species and consequently, designated the genome of Ae. kotschyi as C^uC^uS^vS^v. While several authors (Talbert et al. 1991; Zhang et al. 1992; Friebe et al.1996) considered Ae. longissima, or the immediate precursor of Ae. longissima, and Ae. sharonensis (Zhang et al. 1992) as the source of the S subgenome of Ae. kotschyi, Kihara (1949) noted that the genome of Ae. longissima was not truly homologous to the S subgenome of this allotetraploid. Similarly, Feldman (1963) observed 5.48 univalents, 5.82 bivalents, 0.92 trivalents and 0.28 quadrivalents in Ae. kotschyi x Ae. longissima, showing that the S^v subgenome of Ae. kotschyi differs from the S^l genome of Ae. longissima by two reciprocal translocations. Therefore, the donor species of the S genome in Ae. kotschyi still remains uncertain (Kimber and Feldman 1987).

Sears (1941b) produced an amphidiploid (2n = 4x = 28; genome S^shS^shUU) from the cross of Ae. sharonensis and Ae. umbellulata. This amphidiploid was not very similar in its morphology neither to Ae. kotschyi nor to Ae. peregrina (Kihara 1954). Tanaka (1955b) produced another amphidiploid from the reciprocal cross, Ae. umbellulata x Ae. sharonensis. He then produced hybrids from reciprocal crosses between the resulting amphidiploid and Ae. peregrina. Chromosome pairing at first meiotic metaphase of the F₁ hybrids revealed almost complete pairing, i.e., 9–14 (mode of 13) bivalents and two multivalents that were formed in some of the PMCs. Pollen and seed fertility was relatively high. Tanaka (1955b) concluded that Ae. peregrina and Ae. kotschyi must have arisen as allotetraploids from a cross between Ae. umbellulata, with the U-genome, and Ae. sharonensis with S^sh-genome. The amphidiploid resembles Ae. kotschyi in several morphological characters, but differed from it by other characters, such as in the number of spikelets per spike, awn shape, and others.

Rubenstein and Sallee (1973) analyzed chromosome pairing in hybrids between Ae. kotschyi and different pairing genotypes of Ae. speltoides as well as with Ae. sharonensis and their result supported the view that the second subgenome of Ae. kotschyi showed a high degree of chromosome pairing with the S^sh genome of Ae. sharonensis, and less so with that of Ae. speltoides.

Jaaska (1978a, b), studying the acid phosphatase isoenzyme (isophosphatase) variation in Ae. kotschyi, found that the isophosphatases characteristic of Ae. umbellulata are expressed in Ae. kotschyi, thus confirming the presence of the U subgenome in this allotetraploid species. In addition, the observations did not contradict the proposition that Ae. kotschyi contains the S^v subgenome that either derived from Ae. bicornis, Ae. longissima, or Ae. sharonensis. However, the isophosphatase data did not provide any further information on the similarity of the S^v subgenome of Ae. kotschyi to one of the genomes of these diploids. On the other hand, the data clearly argue against the presumed involvement of the genome of Ae. speltoides in Ae. kotschyi, since the former species is distinct from Ae. bicornis, Ae. longissima, and Ae. sharonensis, by having isophosphatase of higher electrophoretic mobility not encountered in Ae. kotschyi.

Kimber and Yen (1988) and Yen and Kimber (1990d) used a new cytogenetic approach to examine the origin of the U and S^v subgenomes of Ae. kotschyi. Kimber and Yen (1988) studied chromosome pairing in F₁ hybrids between Ae. kotschyi and an induced autotetraploid line of Ae. umbellulata (hybrid genome S^vUUU), The pattern of pairing in this tetraploid hybrid best fit the 3:1 model, indicating that the U subgenome of Ae. kotschyi paired fairly well with the two U genomes of the Ae. umbellulata autotetraploid. Thus, they concluded that the U subgenome of Ae. kotschyi is homologous to the U genome of Ae. umbellulata, confirming that Ae. kotschyi contains a subgenome that derived from Ae. umbellulata. To determine the origin of the S^v subgenomes of Ae. kotschyi, Yen and Kimber (1990d) analyzed chromosome pairing in F₁ of the hybrid between Ae. kotschyi and an induced autotetraploid lines of three Aegilops species of section Sitopsis, namely, Ae. longissima, Ae. speltoides, and Ae. bicornis, as well as with the diploid Ae. speltoides. Their pairing data showed that the hybrid involving the autotetraploid Ae. longissima (hybrid genome S^vUS^lS^l) best fit the 3:1 model, i.e., subgenome S^v is homologous to S^l, while those involving the autotetraploid Ae. speltoides (hybrid genome S^vUSS) and autotetraploid Ae. bicornis (hybrid genome S^vUS^bS^b) best fit the 2:2 and 2:1:1 model, respectively. The triploid hybrid of Ae. kotschyi x diploid Ae. speltoides (hybrid genome S^vUS) best fit the 3:0 model. Thus, they concluded that although modifications occurred in the S^v subgenome of Ae. kotschyi, they were not extensive enough to clearly distinguish the S^v subgenome of Ae. kotschyi from the S¹ genome of Ae. longissima. Consequently, Yen and Kimber (1990d) decided that the genome of Ae. longissima is more similar to the S^v subgenome of Ae. kotschyi than those of Ae. speltoides and Ae. bicornis. However, the absence of the remaining two Sitopsis species, Ae. sharonensis and Ae. searsii, from their study precluded determination of the exact source of the S^v subgenomes. Additional hybrids between Ae. kotschyi and an autotetraploid line of Ae. searsii need to be made to study their genomic relationship (Yen and Kimber 1990d).

The genomes of Ae. longissima, Ae. sharonensis, Ae. bicornis, and Ae. searsii, are very close to one another, rendering it difficult to identify the diploid donor of the S^v subgenome. To resolve this problem, Talbert et al. (1991) used repeated nucleotide sequences to investigate the four species, but failed to distinguish the genomes of these four diploids from one another because only two repeated nucleotide sequences were used in their study.

To reinvestigate the origin of the two subgenomes of the closely-related allotetraploid species, Ae. kotschyi and Ae. peregrina, and the extent of their genome modification, Zhang et al. (1992) employed a developed phylogenetic technique (Dvorak and Zhang 1990; Zhang and Dvorak 1991) based on variation in repeated nucleotide sequences (Dvorak et al. 1988). They made use of twenty-seven randomly selected clones of repeated nucleotide sequences and one 5S rRNA gene clone to identify diagnostic bands and diagnostic hybridization intensities in the restriction profiles of the diploid Aegilops and Triticum species. The presence of each diagnostic band was then determined in Ae. kotschyi and in its closely related species Ae. peregrina. One subgenome in both allotetraploid species was found to be almost identical to the U genome of Ae. umbellulata and the other to the genome of Ae. longissima, Ae. sharonensis, or the internode in the phylogenetic tree of Aegilops/Triticum, immediately preceding the divergence of Ae. longissima and Ae. sharonensis. Twenty diagnostic bands were found to characterize the internode, 19 of which were encountered in Ae. kotschyi and Ae. peregrina, indicating that the S^v subgenome of the two allotetraploids was contributed by the Ae. longissima—Ae. sharonensis evolutionary lineage. Since the shared number of the diagnostic bands between the remaining Sitopsis species and the allotetraploids was close to zero, it is improbable that any of them contributed to the second subgenome of Ae. kotschyi and Ae. peregrina. Taken together, since Ae. kotschyi and Ae. peregrina originated from hybridization between Ae. umbellulata and a species in the lineage of Ae. longissima and Ae. sharonensis, the genome formulae of the two allotetraploids should be S^lS^lUUor S^shS^shUU (Zhang et al. 1992). However, these conclusions disagree with those reached following studies of the cpDNA and mtDNA of these species (Tsunewaki and Ogihara 1983; Ogihara and Tsunewaki 1988; Siregar et al. 1988; Wang et al. 1997; Tsunewaki 2009), which indicated that the cytoplasms of both allotetraploids were contributed by Ae. searsii. Whether this conflict indicates introgression of the cytoplasm from Ae. searsii into Ae. kotschyi and Ae. peregrina or some other causes is not known and requires further investigation. Until the unequivocal identification of the donor of the second subgenome of Ae. kotschyi and Ae. peregrina this subgenome will be designated S^v.

Both Ae. kotschyi and Ae. peregrina have the same S^v type cytoplasm and are thought to have received their cytoplasm from a genome donor with this type cytoplasm (Mukai and Tsunewaki 1975; Tsunewaki et al. 1978). The genetic characteristics of Ae. kotschyi and Ae. variabilis cytoplasms are identical, and differ greatly from those of Ae. umbellulata, and more or less resemble those of Ae. speltoides, Ae. longissima and Ae. bicornis (the cytoplasm of Ae. searsii was not studied (Mukai and Tsunewaki 1975; Tsunewaki et al. 1978; Tsunewaki 1980)). In an attempt to identify the donor of the cytoplasm to the Ae. kotschyi-Ae. peregrina complex, the restriction fragment patterns of chloroplast DNAs of the two allotetraploids were compared with those of several diploids of section Sitopsis (Tsunewaki and Ogihara 1983). The large subunit peptide composition of the chloroplast fraction I protein of Ae. kotschyi and Ae. peregrina is identical with that of Ae. bicornis, Ae. sharonensis and Ae. searsii (Hirai and Tsunewaki 1981), and the ctDNA restriction patterns of the former are identical to those of Ae. bicornis and Ae. searsii (Ogihara and Tsunewaki 1982; Tsunewaki and Ogihara 1983). Thus, Tsunewaki and Ogihara (1983) suggested that the cytoplasm donor of Ae. kotschyi and Ae. peregrina is Ae. bicornis or Ae. searsii. But, Tsunewaki and Ogihara (1983) also pointed out that the cytoplasm of Ae. bicornis differs from those of Ae. kotschyi and Ae. peregrina in the following two respects: first, Ae. kotschyi-Ae. peregrina cytoplasm induced complete male sterility in three of the 12 alloplasmic common wheats tested, whereas Ae. bicornis cytoplasm did not, and, secondly, the cytoplasm of Ae. kotschyi and Ae. peregrina induced haploids in the common wheat cultivar Salmon, but the Ae. bicornis cytoplasm did not. Obviously, Ae. bicornis has different cytoplasm DNA than Ae. peregrina and Ae. kotschyi, although Tsunewaki and Ogihara (1983) did not reveal any differences between the ctDNAs of these allotetraploids and that of Ae. bicornis. This might mean that their mitochondrial DNAs differ (Tsunewaki and Ogihara 1983).

To more accurately detect the diploid donor of the cytoplasm to Ae. kotschyi and Ae. peregrina, Terachi and Tsunewaki (1986), analyzing restriction fragment patterns of mitochondrial (mt) DNA isolated from Ae. kotschyi, Ae. peregrina, Ae. bicornis, and Ae. searsii, revealed that whereas the mitochondrial genomes of Ae. kotschyi and Ae. peregrina was identical to that of Ae. searsii, that of Ae. bicornis was somewhat different. Siregar et al. (1988) used alloplasmic lines of common wheat containing cytoplasm of several Sitopsis species, to compare the effect of the cytoplasm of Ae. bicornis, Ae. searsii and Ae. sharonensis to those of Ae. kotschyi and Ae. peregrina, on the fertility spectrum, haploid and twin induction and restriction fragment patterns of chloroplast and mitochondrial DNAs. In all these respects, the cytoplasm of Ae. searsii most closely resembled the cytoplasm of Ae. kotschyi and Ae. peregrina. Similar results were obtained by Terachi and Tsunewaki (1992), who used RFLPs among mtDNA digests of Triticum and Aegilops species that were analyzed by Southern blot hybridization with four cloned mitochondrial genes, as probes. With all of the probes used, mtDNA from Ae. searsii gave profiles identical to those of Ae. kotschyi. Thus, Ae. searsii was proposed as the cytoplasm donor to these two-allotetraploid species (Terachi and Tsunewaki 1986, 1992; Siregar et al. 1988; Tsunewaki 2009). Similarly, Wang et al. (1997), analyzing PCR–single-strand conformational polymorphism (PCR-SSCP) of 14.0-kb chloroplast (ct) and 13.7-kb mitochondrial mtDNA regions that were isolated from 46 alloplasmic wheat lines, concluded that Ae. searsii was the maternal ancestor of Ae. kotschyi and Ae. peregrina.

Due to the presence of three chloroplast haplotypes, two of which are shared with S‐genome diploids and one with Ae. umbellulata, Meimberg et al. (2009) suggested that Ae. kotschyi evolved through three independent hybridizations between the donor of the S^v subgenome and that of the U subgenome.

In order to more deeply understand the cytogenetic relationships between the two-closely related allotetraploid species, Ae. peregrina and Ae. kotschyi, Feldman (1963), Furuta (1981b) and Cuñado (1993b) studied chromosome pairing in F₁ hybrids between these two allotetraploid species. These studies reported the presence of almost complete pairing in these hybrids, i.e., 0.8–1.0 univalents, 11–12 bivalents and 1–2 multivalents. Evidently, the genome of Ae. peregrina differs, by at least two reciprocal translocations from that of Ae. kotschyi. It was pointed out by Furuta (1981a) that the genome of Ae. kotschyi significantly differs from that of Ae. peregrina. Pairing among chromosomes of the U subgenomes in Ae. peregrina x Ae. kotschyi hybrids was significantly higher than between chromosomes of the S^v subgenomes (Cuñado 1993b). Thus, the differences between peregrina and kotschyi could be exclusively attributed to the S^v subgenomes (Cuñado 1993b).

Based on genome analysis, Kihara (1937, 1949, 1954) concluded that the S^v subgenome of Ae. kotschyi is modified relative to those of the diploid species of Aegilops section Sitopsis, and, accordingly, designated it S^V. To explain the occurrence of modified subgenomes in most allopolyploid Aegilops species, Kihara (1954) suggested that either presently extinct species were the donors of the modified subgenomes or that the subgenomes were significantly rearranged during formation and evolution of the allopolyploids. The degree of S^v-subgenome modification in Ae. kotschyi was estimated differently in different studies; some authors reported that it is modified (Kihara 1940a, 1946, 1949, 1954; Chennaveeraiah 1960; Zohary and Feldman 1962; Kimber and Yen 1989), while others observed only minor differences compared to that of Ae. longissima (Zhang et al. 1992; Friebe et al. 1996).

In an attempt to explain the evolution of allopolyploid species of the wheat (Aegilops-Triticum) group, Zohary and. Feldman (1962) hypothesized that interspecific hybridization between allotetraploid species sharing one subgenome and differing in the second subgenome, also led to some pairing between the differential subgenomes, resulting in introgression and subsequently to modification of these differential genomes. The results of Zhang et al. (1992) showed no difference in divergence between the U subgenome of Ae. kotschyi and the genome of Ae. umbellulata, and only a minor difference was observed between the S^v subgenome of the allotetraploid and the genome of the Ae. longissima–Ae. sharonensis lineage. One of nine bands diagnostic for Ae. bicornis and 1 of 15 bands diagnostic for Ae. searsii, were detected in Ae. kotschyi. Whether these bands introgressed into the allotetraploid or their absence in Ae. longissima and Ae. sharonensis reflects evolution of the Ae. longissima—Ae. sharonensis lineage after the origin of the allotetraploids, remains to be determined (Zhang et al. 1992). The question of modification of the S^v subgenome by chromosome introgression (Zohary and Feldman 1962) may require further investigation, since the introgression of small chromosome segments might have escaped detection in the work of Zhang et al. (1992).

Following C-banding analysis, Badaeva et al. (2004) reported that the U subgenome in Ae. kotschyi is similar to the U subgenomes of all other polyploids bearing the U subgenome and to that of the parental diploid species Ae. umbellulata. On the other hand, the second subgenome of these allopolyploid species were modified compared to the donor chromosomes, as was suggested by Kihara (1954, 1963), Chennaveeraiah (1960), Kimber and Abu-Bakar (1981), Kimber and Zhao (1983), Kimber and Feldman (1987), and Kimber and Yen (1989), with the extent of subgenome modification varying between species. Similarly, Gong et al. (2006), using 31 inter-simple sequence repeat (ISSR) primers, compared the genome of Ae. kotschyi with those of its ancestral diploids. Based on the genetic similarity index between Ae. kotschyi and Ae. umbellulata, they reported that the allotetraploid contains one subgenome similar to the U genome of Ae. umbellulata, while the second subgenome was altered greatly in the allotetraploid.

Feldman (1963), Kawahara (1986, 1988), and Fernández-Calvín and Orellana (1991) found that most interchanges between Ae. kotschyi and Ae. peregrina involved chromosomes of the S^v subgenome. This suggests that genome rearrangement occurs more frequently in the S^v subgenome than in the U subgenome. Kawahara (1986, 1988) proposed that the modified genome of Ae. kotschyi probably evolved through its high structural variability rather than through introgression with other species. Yet, Furuta and Tanaka (1970) concluded from their experimental hybridization between Ae. peregrina and Ae. columnaris, that it may have been exchange of genetic material between the differential subgenomes of these allotetraploids, namely, S^v and Xⁿ, that modified them. Thus, introgression occurring between species sharing one subgenome and differing in the second subgenome can lead to formation of recombinant (modified) subgenomes. Since the S^v subgenomes of Ae. kotschyi and Ae. peregrina are close to each other, it is more difficult to obtain cytological and molecular evidence for modification of their S^v subgenomes through introgression, although morphological evidence indicated that such introgression indeed takes place (Feldman 1963). Assuming that the S^v subgenome of Ae. kotschyi derived from Ae. sharonensis and that of Ae. peregrina from Ae. longissima, as was suggested by Badaeva et al. (2004), the latter differs from the former diploid parent by a translocation (Ankori and Zohary 1962), or alternatively, one of the allopolyploids originated as a result of introgressive hybridization with Ae. searsii (Tsunewaki 2009 and references therein), which differs from Ae. longissima by a translocation (Feldman et al. 1979), such courses of events may explain the changes in the S^v subgenome of these allotetraploids.

Ae. kotschyi contains 12.64 ± 0.183 pg 1C DNA (Eilam et al. 2008), the largest genome among the allotetraploid species of Aegilops (Eilam et al. 2008). The 1C DNA content of Ae. kotschyi is 1.71% smaller than the DNA expected from the sum of the DNA of its two putative diploid parents, i.e., 12.86 pg (Ae. umbellulata, the donor of the U subgenome, contains 5.38 pg and Ae. longissima, the donor of the S^v subgenome, contains 7.48 pg; Eilam et al. 2007). If assuming that the S^v subgenome donor is Ae. sharonensis, then the 1C DNA content of Ae. kotschyi is 2.02% smaller than the DNA expected from the sum of the DNA of its two diploid parents, i.e., 12.90 pg (Ae. umbellulata, the donor of the U subgenome, contains 5.38 pg and Ae. sharonensis, the donor of the S subgenome, contains 7.52 pg; Eilam et al. 2007).

The karyotype figured by Senyaninova-Korchagina (1930, 1932) for Ae. triuncialis subsp. kotschyi Boiss. (Currently Ae. kotschyi) was similar to the one she figured for Ae. peregrina. In genome analytical studies, Kihara (1954, 1957) included Ae. kotschyi under Ae. peregrina, perhaps, partly due to the karyotypic results of Senyaninova-Korchagina (1930, 1932). However, the karyotypes studied by Chennaveeraiah (1960) revealed considerable differences between the karyotypes of Ae. kotschyi and Ae. peregrina. Ae. kotschyi has only two pairs with satellites on short arms whereas Ae. peregrine has three. The rest of the pairs in Ae. kotschyi consist of one with an extreme subterminal centromere, one with a median centromere, one with an almost median centromere, and the others with submedian centromeres. Thus, in all, there are 12 types of chromosomes in Ae. kotschyi (Chennaveeraiah 1960).

One half of the chromosomes of Ae. kotschyi corresponds to one half of the chromosomes of Ae. peregrina, which all correspond to the U genome of Ae. umbellulata (Chennaveeraiah 1960). Distinct differences in the second half of chromosomes in the two species were noted (Chennaveeraiah 1960). The second set in Ae. kotschyi consisted of chromosomes with only primary constrictions which were median or submedian. There were no pairs with satellite or secondary constriction in th S^v subgenome of Ae. kotschyi, whereas Ae. peregrina did have a pair with satellite in the second set of chromosomes. There was no chromosome with a median centromere in the entire set of Ae. peregrina, whereas Ae. kotschyi had one such pair. The chiasma frequency in meiotic first metaphase was also different in the two species; more rod bivalents formed in Ae. peregrina than in Ae. kotschyi (Chennaveeraiah 1960). Judging from karyotypic observations, Chennaveeraiah (1960) proposed that the second subgenome of Ae. kotschyi is similar to the M-genomes of species of section Comopyrum. It could even be foreign to Aegilops but never the same as the second subgenome in Ae. peregrina. Thus, also the study of Chennaveeraiah (1960) revealed that Ae. kotschyi and Ae. peregrina are separate species.

Like Chennaveeraiah (1960), Al-Mashhadani et al. (1980) also observed two satellite chromosome pairs in Ae. kotschyi from Iraq. The satellites in one pair were larger than in the other. In addition to the SAT chromosomes, five metacentric chromosome pairs, four submetacentric pairs and three subtelocentric pairs were observed. These observations are quite similar to those of Tanaka and Matsumoto (1965), and clearly differ from those of Chennaveeraiah (1960).

Badaeva et al. (2004) investigated the heterochromatin structure in the karyotype of Ae. kotschyi using C-banding and FISH with the heterochromatin-specific DNA probes pSc119 and pAs1 (non-coding, highly repeated DNA sequences), as well as of rDNA loci, using pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) probes. Ten accessions of Ae. kotschyi were examined by C-banding and one accession was analyzed by FISH. Although Ae. kotschyi has the same genome formula as Ae. peregrina, the C-banding patterns were different. Comparison of Ae. kotschyi with the diploid ancestors Ae. umbellulata and Ae. sharonensis revealed a higher degree of S^v subgenome modification in Ae. kotschyi than in Ae. peregrina. Chromosomes 4 Sv and 7 Sv of Ae. peregrina are nearly identical to 4S^l and 7S^l of Ae. longissima, whereas these chromosomes of Ae. kotschyi are similar to 4S^sh and 7S^sh of Ae. sharonensis. Previous studies performing isozyme (Hart and Tuleen1983) and C-banding analyses (Friebe et al. 1993, 1996; Friebe and Gill 1996) revealed that chromosomes 4S^l and 7S^l of Ae. longissima are involved in a species-specific reciprocal translocation that is absent in Ae. sharonensis. The presence of this translocation in Ae. peregrina and its absence in Ae. kotschyi suggests that Ae. peregrina originated from the hybridization of Ae. umbellulata and Ae. longissima and that Ae. kotschyi originated from the hybridization of Ae. umbellulata and Ae. sharonensis (Badaeva et al. 2004). Thus, the C-banding analysis showed that the S^v subgenome of Ae. kotschyi was derived from either the S^sh genome of Ae. sharonensis or its immediate precursor. The high frequency of chromosomal aberrations and reduction in the number and size of 18S-26S rDNA loci observed in the S^v subgenome compared to the S^sh and S^l of Ae. peregrina, suggested that Ae. kotschyi is an older species than Ae. peregrina.

FISH with clone pSC119 revealed signals of various sizes in telosomic regions of either the short or both arms of 12 chromosome pairs of Ae. kotschyi (Badaeva et al. 2004). Interstitial pSc119 FISH sites were detected in the long arm of chromosome 7U, while chromosome 6U had no such sites; distinct telomeric and interstitial FISH sites were present in Ae. umbellulata (Badaeva et al. 1996a). However, in the allotetraploid species, chromosome 6U has telomeric and interstitial pAs1 FISH sites that are absent in Ae. umbellulata.

Similar to Chennaveeraiah (1960) and Al-Mashhadani et al. (1980), Badaeva et al. (2004) found two pairs of satellite chromosomes in Ae. kotschyi, which coincides with the number of active NORs detected by Ag-NOR staining (Cermeno et al. 1984b) and in situ hybridization (Yamamoto 1992a). Like in Ae. peregrina, the NORs on Ae. kotschyi S^v-subgenome chromosomes were inactivated and accompanied with a decrease or loss of rDNA sequences. Consequently, the major NORs were observed on group 1U and 5U chromosomes. Ae. kotschyi also have several minor 18S-26S rDNA sites. Two consistent minor loci detected on group 5 Sv and 6 Sv chromosomes were associated with a significant reduction in copy number of 18S-26S rRNA genes. The hybridization patterns with pTa794 revealed four similar-sized 5S rDNA loci that were located on chromosomes of homoeologous groups 1 and 5.

9.8.9.3 Crosses with Other Species of the Wheat Group

Chennaveeraiah (1960) reported that meiosis is very regular in Ae. kotschyi exhibiting 14 bivalents at first meiotic metaphase of every PMC. Likewise, Feldman (1963) reported that all PMCs at first meiotic metaphase of Ae. kotschyi had 14 bivalents and 29.40 chiasmata/cell. Chromosome pairing in triploid hybrids between Ae. kotschyi and Ae. umbellulata indicated that the former contains one subgenome that is homologous to the genome of the latter (von Berg 1937; Kimber and Abu-Bakar 1981). Kimber and Abu-Bakar reported the presence of 7.04 univalents, 2.90 rod bivalents, 3.66 ring bivalents, 6.56 total number of bivalents and 0.29 trivalents (Table 9.5). Kihara (1949) reported 7 bivalents, five of which were rings, and up to one multivalent per cell in the triploid hybrid Ae. longissima x Ae. kotschyi. However, in the footnote to his Table 9.5, Kihara indicated that the genome of Ae. longissima was not truly homologous to the S^v subgenome of Ae. kotschyi. Likewise, Feldman (1963) studied chromosome pairing in the F₁ triploid hybrid Ae. kotschyi x Ae. longissima and observed 5.48 univalents, 5.82 bivalents, (of which 1.52 were heteromorphic), 0.92 (0–2) trivalents, 0.28 (0–1) quadrivalents and 11.92 chiasmata/cell. This pattern of chromosome pairing indicates the existence of homology between one subgenome of Ae. kotschyi and the S^l genome of Ae. longissima, although some structural differences (two reciprocal translocations) exist between the two species. On the other hand, chromosome pairing in the triploid hybrid Ae. kotschyi x Ae. caudata (genome S^vUC) had 10.70 univalents, 3.45 rod bivalents, 0.35 ring bivalents (3.80 total number of bivalents) and 0.90 trivalents (Kimber and Abu-Bakar 1981), indicating that the genome of Ae. caudata is homoeologous to the subgenomes of the allotetraploid.

Hybrids between Ae. kotschyi and both Ae. speltoides and Ae. sharonensis were produced and examined cytologically (Table 9.5). Chromosome pairing in the F₁ triploid hybrids between Ae. kotschyi and different pairing types of Ae. speltoides (genome S^vUS) reveals that the allotetraploid contains one genome that is related to the genome of the diploid. Interestingly, the high pairing and the intermediate pairing genotypes of Ae. speltoides did not promote pairing in the above hybrids (Rubenstein and Sallee 1973). Chromosome pairing in the F₁ hybrid Ae. kotschyi x Ae. sharonensis was somewhat higher than in the hybrid kotschyi x speltoides, confirming the presence of a subgenome in Ae. kotschyi that is homologous to the genome of Ae. sharonensis, but which differs from it by at least two reciprocal translocations.

Chromosome pairing in the F₁ tetraploid hybrid between several lines of the two subspecies of Ae. peregrina and Ae. kotschyi, was studied by Feldman (1963). The hybrids involving lines of Ae. peregrina ssp. peregrina and Ae. kotschyi had 0.84–0.94 univalents, 10.98–11.28 bivalents, 0.14–0.20 trivalents, 0.26–0.54 quadrivalents, 0.14–0.18 pentavalents, and 0.24–0.36 hexavalents. Hybrids involving lines of Ae. peregrina ssp. cylindrostachys and Ae. kotschyi had 0.84–1.02 univalents, 11.42–11.72 bivalents, 0.18–0.24 trivalents, 0.18–0.68 quadrivalents, 0.02–0.30 pentavalents, and 0.06–0.20 hexavalents. Evidently, the genomes of both subspecies of Ae. peregrina are homologous to that of Ae. kotschyi, but differ from it by, at least, two reciprocal translocations.

Similarly, Furuta (1981b) crossed 21 lines of Ae. kotschyi, collected in Egypt, Jordan, and Syria, with a line of Ae. peregrina that served as the common pollen parent. Chromosome pairing in the F₁ hybrids also displayed the presence of one or two reciprocal translocations between these two species. Furuta (1981b) studies revealed that variable and continuous chromosomal differentiation occurred between the chromosomes of Ae. peregrina and Ae. kotschyi. Based on these results, Furuta (1981b) concluded that the genome of Ae. kotschyi significantly differs from that of Ae. peregrina.

Comparison of chromosomal pairing of intraspecific hybrids in Ae. peregrina with hybrids between Ae. peregrina and Ae. kotschyi, enabled assessment of whether the U and S^v subgenomes had been altered during their evolution in one or both species (Cuñado 1993b). Chromosome pairing in the intraspecific hybrid peregrina x peregrina showed 0.18 univalents, 4.32 rod bivalents, 9.56 ring bivalents, and 23.44 associations/cell. Pairing in the U subgenome included 0.02 univalents, 2.14 rod bivalents, 4.84 ring bivalents, and 11.83 associations/cell, and pairing in the S^v subgenome showed 0.16 univalents, 2.18 rod bivalents and 4.72 ring bivalents (11.61 associations/cell (Cuñado 1993b). The number of associations/cell in the intraspecific hybrid of peregrina was lower than that of the parental lines Ae. peregrina (23.44 vs. 25.30; Cuñado 1993b). This decrease in association frequency cannot be attributed to structural changes since multivalents were not observed (Cuñado 1993b). Chromosome pairing in the hybrid peregrina x kotschyi included 0.99 univalents, 3.67 rod bivalents, 8.43 ring bivalents 0.47 trivalents, 0.35 quadrivalents, and 21.71 associations/cell. Pairing in the U subgenome showed 0.04 univalents, 2.05 rod bivalents, 4.93 ring bivalents, and 11.93 associations/cell, and pairing in the S^v subgenome had 0.95 univalents, 1.62 rod bivalents, 3.50 ring bivalents, 0.47 trivalents, 0.35 quadrivalents, and 9.78 associations/cell. Evidently, the S^v subgenome of the two species differs in a reciprocal translocation. These results are in agreement with those reported by Furuta (1981b) and Feldman (1963), although Furuta and Feldman found a higher number of interchanges in some hybrids involving other lines. The mean number of associations/cell between chromosomes of the U subgenomes were similar in the two hybrids. However, the frequencies of chromosome associations of the S^v subgenomes differed significantly in the peregrina x kotschyi as compared to the intraspecific hybrid. Thus, the differences between peregrina and kotschyi could be exclusively attributed to the S^v subgenomes (Cuñado 1993b).

Chromosome paring in tetraploid hybrids between Ae. kotschyi and several allotetraploid species bearing the U subgenome were studied by a number of groups. The hybrid Ae. geniculata x Ae. kotschyi (genome M^oUS^vU) had 11.70 univalents, 6.02 bivalents (of which 1.76 were heteromorphic) and 1.42 trivalents (Feldman 1963), while others reported 10.65 univalents, 3.20 rod bivalents, 3.25 ring bivalents, 6.45 total number of bivalents, 1.35 trivalents and 0.10 quadrivalents (Kimber et al. 1988). The hybrid Ae. columnaris x Ae. kotschyi (genome UXⁿS^vU) had 7–10 bivalents with mode of 9, 0–3 trivalents and 0–2 quadrivalents (Kihara 1949). The hybrid Ae. neglecta x Ae. kotschyi (genome UXⁿS^vU) had 7–10 bivalents with mode of 10, and 0–2 trivalents (Kihara 1949). The hybrid Ae. kotschyi x Ae. triuncialis (genome S^vUUC) had 6–9 bivalents (Kihara 1937). In all these hybrids, there were more than seven bivalents, indicating that, in addition to homologous pairing between the shared U subgenomes, homoeologous pairing also occurred between some chromosomes of the differential subgenomes.

Chromosome pairing in the hybrid common wheat x Ae. kotschyi (genome ABDS^vU) included 30.92 univalents, 1.98 rod bivalents, 0.02 ring bivalents, and 0.03 trivalents (Fernández-Calvín and Orellana 1991). Of the rod bivalents, 0.54 were between A-D chromosomes, 0.74 between AD-US^v chromosomes, 0.05 between AD-B chromosomes, 0.38 between US^v-B chromosomes and 0.25 between U-S^v chromosomes. The data indicated that more than 40% of the pairing in these three hybrids was autosyndetic (A-D + AD-B = 25%, and U-S^v = 16%) and that more than 50% was allosyndetic (AD-US^v + US^v-B = 51%) (Fernández-Calvín and Orellana 1991). The allosyndetic pairing between AD chromosomes and US^v chromosomes was twice as high as that between US^v and B chromosomes.

9.8.10 Aegilops peregrina (Hackel) Maire & Weiller

9.8.10.1 Morphological and Geographical Notes

Ae. peregrina (Hackel) Maire & Weiller [Syn.: Ae. variabilis Eig; Ae. peregrina (Hack. In Fraser) Eig; T. peregrinum Hack. In J. Fraser; Aegilemma peregrina (Hack.) Á. Löve] is a predominantly autogamous, annual, multi–tillered, 15–40-cm-tall (excluding spikes) plant. It branched and prostrate near the ground but upright at its upper parts. Its leaves are hairy or glabrous and spike is broad oval, linear to cylindrical, 1.2–7.5-cm-long (excluding awns), disarticulating entirely at maturity (umbrella-type dispersal unit), and usually awned. There are 2–4 (usually 3, exceptionally 1) basal rudimentary spikelets and 2–7, (usually 3–5), urn-shaped to elliptical spikelets, not appressed to the rachis or to each other. The spikelets become smaller to the tip of the spike, with the uppermost spikelet seldom becoming suddenly smaller. There are 3–6, usually 4–5 florets, with the upper 1–3 being sterile. The glume is usually tough and rough, 6–8-mm–long and 4–6-mm-wide, with weak, narrow, parallel, and equally long and wide nerves. There are 3 glume awns of the terminal spikelet, and 2–3 awns on lateral spikelets, with the central one replaced by a tooth or gap when there are only two. The number of awns never exceeds 3. Glume awns are narrow and flat at the base, strongly polymorphic and variable in number, width and length, and spread out at maturity. They are either equally broad, broader at the lower spikelets, or one is considerably broader (particularly in the lower spikelets). Their lengths are either equal, or differ (by 4–8 mm), or shorter in the middle as compared to its laterals. Lemma awns are weakly developed, and often missing, and when present, there is 1, very seldom 3, and always shorter than glume awns, often with more or less long teeth. The caryopsis adheres to the lemma and palea (Fig. 9.4i, j).

The species shows the most extraordinary variability (Fig. 9.5) and was therefore called Ae. variabilis by Eig (1929a), although he was perfectly aware that Hackel had already given the name Triticum peregrinum to a specimen of this species which had been casually introduced into Scotland. Eig preferred to give a new name to the species because Hackel’s specimen was atypical, calling for abandonment of the specific epithet. Maire and Weiller (1955) seem to be the first authors to have validly published the combination of Ae. peregrina, and transferred most of the intraspecific taxa of Eig from the species epithet variabilis Eig to the correct one, peregrina Hack.

Fig. 9.5

Spikes of Ae. peregrina (=Ae. variabilis Eig) representing portion of the morphological variation of this species (From Feldman 1963)

Eig (1929a) described two subspecies in this species, eu-variabilis (containing 7 varieties) and cylindrostachys (containing 3 varieties). Hammer (1980), accepting the transfer of Eig’s intraspecific taxa from variabilis to peregrina, designated Eig’s two subspecies as peregrina and cylindrostachys (Eig et Feinbrun) Hammer. In contrast to Eig and Hammer, van Slageren (1994) ranked the intraspecific taxa of Ae. peregrina as varieties: var. peregrina and var. brachyathera. The sub-classification of Hammer (1980) into subspecies will be used in this book.

The description of the two subspecies is as follows: Ssp. peregrina (Ae. variabilis ssp. eu-variabilis Eig & Feinbr.; Fig. 9.4i) has 1.5–4-cm–long (excl. awns), broadly ovate to lanceolate spikes, with 3–4 spikelets. The lower spikelets are longer than the adjacent rachis internodes. The glumes have 2–3 awns, with terminal spikelet glumes bearing three awns. The lemmas of all spikelets have 1–2 awns of notably uneven length (0.3–3-cm), flanked by 1–2 teeth. This subspecies is most variable in awn length. Eig (1929a) described seven varieties within this subspecies. Ssp. cylindrostachys (Eig & Feinbr.) Hammer (Ae. variabilis Eig ssp. cylindrostachys Eig & Feinbr.; Fig. 9.4j) has 3.5–7.5-cm–long (excl. awns), narrowly lanceolate to linear spikes, with (3-)5(-7) spikelets. The glumes of the lateral spikelets feature 2–3 sharp teeth, 1 or 2 of which may develop into a short awn (up to ± 7 mm-long, increasing to 1.5 mm subapically). The glumes of the apical spikelet bear 1–3, 1–3-cm-long awns. In the case of only one, the awn is flanked by acute teeth of up to 6-mm in length. All spikelet lemmas have 2–3 teeth. The rachis internodes are generally about as long as those of the adjacent spikelets. Awned variants are rare, and, when present, the awns on glumes are short whereas awns on lemmas are present only in the terminal spikelet. Eig (1929a) described three varieties within this subspecies. Subsp. cylindrostachys is less common than subsp. peregrina.

Ae. peregrina also exhibits wide variation at the biochemical and molecular levels. Nakai and Tsunewaki (1971) found variations in the zymograms of esterase isozymes in four Ae. peregrina accessions, analyzed using the gel electrofocusing method. Similarly, Nakai and Tsuji (1984), after examining four accessions of Ae. peregrina for acid phosphatase isozymes, using gel electrofocusing technique, reported on intraspecific variation of two variant phenotypes. Such intraspecific variation may be the result of adaptation to different environment conditions, as shown by Nevo et al. (1984), who electrophoretically analyzed allozymic diversity in two polymorphic esterase loci in 70 Ae. peregrina plants, collected from a microsite at Tabigha, north of the Sea of Galilee, Israel. The test involved a 100 m-transect, equally subdivided into basalt and terra-rossa soil types. Significant genetic differentiation across soil type was found over very short distances. The results suggested that allozyme polymorphisms in Ae. peregrina are adaptive and differentiate primarily by soil selection, probably through aridity stress.

RAPD analysis was used to study genetic variation and phylogenetic relationships among allopolyploid Aegilops species sharing the U-subgenome (Goryunova et al. 2010). In total, the group examined 115 DNA samples of eight allopolyploid species containing the U subgenome and of the diploid species Ae. umbellulata (genome U). Substantial interspecific polymorphism was observed in the majority of these allpolyploids. As with most other species, the 12 accessions of Ae. peregrina exhibited wide intraspecific variation. When establishing the phylogenetic relationships for the U-subgenome species, the authors noted the greatest separation within this group between the S^vU-subgenome species Ae. peregrina and Ae. kotschyi.

Ae. peregrina is a Mediterranean element, growing in Southern Italy (including Sicily), South Greece (including Crete and Rhodes), South Turkey, Iraq (lower Mesopotamia), Iran (northwest and south), Azerbaijan, Syria, Lebanon, Cyprus, Israel, Jordan, Egypt (lower), Libya, Tunisia, Algeria, and Morocco. In this region, Ae. peregrina thrives on a large variety of soils, in edges and openings of sclerophyllous oak forest, maquis, dwarf shrub formations, herbaceous formations, pastures abandoned fields, edges of cultivation, disturbed areas and roadsides. It grows at altitudes of 0–1600 m. The species is very common and locally abundant. It rapidly colonizes deserted fields as well as open, unstable, secondary habitats.

Ae. peregrina has a medium-sized distribution in the southwestern part of the distribution of the genus. Like Ae. triuncialis in the central and northern parts of the genus area, Ae. peregrina is the massive species in the southwestern part of the genus area. Its ecological amplitude is exceptionally large. It differs from its closely related species Ae. kotschyi, in that, in some areas, it grows sympatrically with both of its putative diploid parents, Ae. umbellulata and either Ae. longissima and, Ae. sharonensis or Ae. searsii. Ae. peregrina usually grows in mixed populations with other species, with which it introgresses. It may have contact with its two putative diploid parents in semi-steppical, steppical and sub-Mediterranean regions of Syria or Israel where Ae. peregrina might have originated and then spread southwards and westwards. Its distribution area is larger than those of its two putative parents.

Ae. peregrina grows sympatrically with Ae. speltoides, Ae. sharonensis, Ae. longissima, Ae. searsii, Ae. caudata, Ae. umbellulata, wild tetraploid wheat, T. turgidum ssp. dicoccoides, Ae. geniculata, Ae. biuncialis and Ae. triuncialis, and allopatrically with Ae. bicornis, Ae. comosa, the wild subspecies of T. monococcum, Ae. urartu, Ae. neglecta, Ae. columnaris, and Ae. kotschyi. Ae. peregrina forms mixed populations with Ae. geniculata and Ae. biuncialis in many parts of the Mediterranean phytogeographic region of Israel. In such populations, interspecific hybrids and hybrid derivatives between Ae. peregrina and the other two allotetraploids are quite common (Zohary and Feldman 1962; Feldman 1965a; Pazy and Zohary 1965). Ae. peregrina and its close relative Ae. kotschyi are vicarious species, the former growing in the Mediterranean phytogeographic region and the latter in semi-steppical and steppical (Irano-Turanian) regions. In the transition zone between southern Israel’s Mediterranean and steppical regions where Ae. peregrina and Ae. kotschyi have massive contact, there are many indications for gene flow between these two species (Feldman 1963). In northern Israel, there are mixed populations of four allotetraploids, namely, Ae. peregrina, Ae. geniculata, Ae. biuncialis and Ae. triuncialis, with intermediates between Ae. peregrina and Ae. triuncialis found in several such populations (Zohary and Feldman 1962; Feldman 1965a). In addition, hybrids between Ae. peregrina and wild and domesticated tetraploid wheat, as well as domesticated hexaploid wheat, were repeatedly found (Percival 1921; Feldman M, unpublished). In this regard, Weissman et al. (2005) described a spontaneous DNA introgression from domesticated hexaploid wheat into Ae. peregrina and the stabilization of this introgression in wild populations. Vardi and Zohary (1967) described triploid hybrids and hybrid derivatives in mixed populations of tetraploid Ae. peregrina and diploid Ae. sharonensis or Ae. longissima, indicating the possibility of gene flow between these species.

Accessions of Ae. peregrina carry genes that confer resistance to powdery mildew (Gill et al. 1985; Spetsov et al. 1997), leaf rust (Gill et al. 1985; Marais et al. 2008), stem rust (Anikster et al. 2005; Scott et al. 2014) and strip rust (Anikster et al. 2005; Liu et al. 2011a, b; Zhao et al. 2016). Accessions of this species were also found resistant to Hessian fly (Gill et al. 1985), and green bug (Gill et al. 1985), as well as to cereal cyst nematode (Coriton et al. 2009) and to root-Knot nematodes (Yu et al. 1990; Coriton et al. 2009). Several accessions of Ae. peregrina were salt tolerant (Farooq et al.1989).

9.8.10.2 Cytology, Cytogenetics, and Evolution

Ae. peregrina is an allotetraploid species (2n = 4x = 28). Its nuclear genome is designated S^vS^vUU (modified from Dvorak 1998) and its plasmon genome as S^v (identical to S^s of Ae. searsii) (Tsunewaki 2009). Early genome analysis studies showed that Ae. peregrina has the U subgenome from Ae. umbellulata (von Berg 1937; Kihara 1940a; Kimber and Yen 1989), findings that were later substantiated by a karyological study (Chennaveeraiah 1960), cytogenetic studies (Yu and Jahier 1992), biochemical studies (Jaaska (1978a, b) and subsequent molecular studies (Zhang et al. 1992; Badaeva et al. 2004). Yet, in contrast to the consensus concerning the nature and origin of the U subgenome of Ae. peregrina, that of the second subgenome is still enigmatic. Chennaveeraiah (1960), based on karyotypic considerations, suggested that the donor of the second subgenome is a species of the M-genome group. From his cytogenetic studies, Kihara (1946, 1949) proposed that the second subgenome of Ae. peregrina was contributed by a species of the Sitopsis group, possibly Ae. longissima. This proposal was supported by Talbert et al. (1991) and Friebe et al. (1996), whereas Zhang et al. (1992) assumed that either Ae. longissima, Ae. sharonensis or, more likely, the immediate precursor of these two-diploid species, was the donor of the S^v subgenome to Ae. peregrina. Badaeva et al. (2004), using C-banding method, revealed that Ae. peregrina contains a reciprocal translocation involving chromosomes 4 and 7 of the S subgenome, similar to the translocation that exists in Ae. longissima. Since this translocation does not present in Ae. sharonensis, they concluded that Ae. longissima was the source of the second subgenome of Ae. peregrina, and that Ae. peregrina arose after the separation of Ae. longissima from Ae. sharonensis. In contrast to the above, Ogihara and Tsunewaki (1988) and Siregar et al. (1988), studying restriction patterns of chloroplast DNA, proposed Ae. searsii as the cytoplasm donor to Ae. peregrina.

Sears (1941b) produced an Ae umbellulata—Ae sharonensis amphidiploid (2n = 4x = 28; genome UUS^shS^sh), which was not similar to Ae. peregrina, while the F₁ of the hybrid Ae. bicornis x Ae. umbellulata (Genome S^bU) had a similar ear-form to Ae. peregrina (Kihara 1954). Tanaka (1955b) also produced an amphidiploid from the Ae. sharonensis x Ae. umbellulata cross, that reportedly resembled Ae. kotschyi in many morphological characters, but differed from Ae. kotschyi and Ae. peregrina with respect to spikelet count and awn shape. Tanaka (1955b) reported that chromosome pairing in the F₁ hybrid of the amphidiploid x Ae. peregrina was almost regular, i.e., 0–4 univalents, with a mode of 2, 9–14 bivalents, with a mode of 13, and 0–2 trivalents or quadrivalents; pollen fertility was 71.5% and seed set 50.0%. These data implied that Ae. peregrina originated from the hybridization between Ae. sharonensis, or another closely related species, and Ae. umbellulata. Kihara (1954) regarded the S subgenome of Ae. peregrina as a modified subgenome and therefore, formulated it as S^v.

These findings were reinforced by Zhang et al. (1992), who identified diagnostic bands in Southern blots hybridized with repeated nucleotide sequences and one 5S rRNA gene. Their study confirmed that one subgenome in Ae. peregrina was identical to the U genome of Ae. umbellulata and that the other was identical to the S¹ genome of Ae. longissima or to the S^sh genome of Ae. sharonensis or, more likely, to the internode in the phylogenetic tree of Triticum immediately preceding the divergence of Ae. longissima and Ae. sharonensis. Their data indicated that the second subgenome of Ae. peregrina was contributed by the Ae. longissima-Ae. sharonensis evolutionary lineage, and not by any other Sitopsis species.

When the data of Zhang et al. (1992) are interpreted in the context of the phylogenetic tree of the wheat group, it appears that Ae. peregrina and Ae. kotschyi are of recent origin and evolved after the differentiation of Ae. umbellulata from Ae. caudata, and Ae. longissima and Ae. sharonensis from Ae. bicornis.

The results of the cytogenetic (Tanaka 1955b) and molecular (Zhang et al. 1992) studies on the source of the S^v subgenome in Ae. peregrina and Ae. kotschyi disagree with the inference based on chloroplast (cp) DNA and mitochondrial (mt) DNA (Ogihara and Tsunewaki 1988; Siregar et al. 1988; Wang et al. 1997; Tsunewaki 2009), which indicated that the cytoplasm of both allotetraploids was contributed by Ae. searsii and not by Ae. longissima.

Both Ae. peregrina and Ae. kotschyi are allotetraploid with the same S^v-type cytoplasm and are thought to have received their cytoplasm from a genome donor with the S^v-type cytoplasm (Mukai and Tsunewaki 1975; Tsunewaki et al. 1978). Studies on the large subunits of the chloroplast Fraction I protein (Hirai and Tsunewaki 1981) and chloroplast DNA restriction pattern (Ogihara and Tsunewaki 1982; Tsunewaki and Ogihara 1983), suggested that the S^v cytoplasm derived either from Ae. searsii or Ae. bicornis. While it is obvious that Ae. bicornis has different phenotypic effects in alloplasmic lines of bread wheat than the cytoplasm of Ae. searsii (Siregar et al. 1988), studies of the cytoplasm of Ae. peregrina and Ae. kotschyi, did not reveal any differences between their ctDNAs and that of Ae. bicornis, suggesting that these species differ in their mitochondrial genomes (Tsunewaki and Ogihara 1983).

Indeed, while aiming to more accurately determine the diploid donor of the cytoplasm to Ae. peregrina and Ae. kotschyi, Terachi and Tsunewaki (1986), analyzing restriction fragment patterns of mtDNA isolated from various Aegilops species, revealed that whereas the mitochondrial genomes of Ae. peregrina and Ae. kotschyi are identical to that of Ae. searsii, that of Ae. bicornis is somewhat different. Thus, Ae. searsii was proposed as the cytoplasm donor to these two allotetraploid species (Tsunewaki 2009 and references therein). Whether this conclusion indicates introgression of the cytoplasm from Ae. searsii into Ae. peregrina (and Ae. kotschyi) or some other cause, is currently not known and requires further investigation.

Wang et al. (1997) analyzed PCR–single-strand conformational polymorphism (PCR-SSCP) of 14.0-kb ct and 13.7-kb mt DNA regions that were isolated from 46 alloplasmic wheat lines. In accord with the above, they found that the genetic distances between Ae. searsii and both Ae. peregrina and Ae. kotschyi were moderate (0.008 and 0.011, respectively) and between the two allotetraploids was small (0.005). Therefore, Wang et al. (1997) designated the cytoplasm of Ae. searsii S^v, like the designation of the cytoplasm of Ae. peregrina and Ae. kotschyi. Apparently, Ae. peregrina and Ae. kotschyi had a monophyletic origin (Wang et al. 1997). In this respect, it is interesting to note that Meimberg et al. (2009) assumed a single origin for Ae. peregrina.

In his attempt to explain the origin of the modified genomes in the Aegilops allotetraploid species, Kihara (1963) assumed subgenome donors to be either extinct or yet unknown diploid species, or that independent chromosome differentiation occurred within the subgenome. Kawahara (1986, 1988) proposed an alternative and more simple explanation for the existence of modified subgenomes in the Aegilops-Triticum group. He assumed that this group contains several diploid species with stable genomes, while other diploids have less stable and more variable genomes. In an allotetraploid species with the genomic combination of one stable and one variable subgenomes, structural differentiation or segmental rearrangements would accumulate in the variable genome. Since one genome is stable, serving as a genetic buffer, the chromosome structure of the second genome would change far more rapidly than that of the corresponding genome of the diploid species. To verify this hypothesis, Kawahara (1986, 1988) identified the breakpoints of spontaneous reciprocal translocations in each subgenome. In Ae. peregrina and Ae. kotschyi he identified seven translocations, three being between the S^v subgenome chromosomes, two between the U and the S^v subgenomes and one between the U subgenome chromosomes. The breakpoint of the remaining translocation was assumed to be on a S^v subgenome chromosome but was not identified (Kawahara T, unpublished). Evidently, the number of breakpoints on the modified S^v is clearly about twice that on the pivotal U subgenome. Therefore, Kawahara (1986, 1988) concluded that genome rearrangement occurs more frequently in the modified genomes than in the pivotal ones, and that the modified genomes likely evolved through their high structural variability.

The coexistence of a modified and unchanged subgenome within the same nucleus of an allotetraploid Aegilops species, led to the hypothesis that interspecific hybridization between allotetraploid species sharing one subgenome, but differing in the other subgenome, results in the modification of the differential subgenomes via hybridization and gene exchange (Zohary and Feldman 1962; Feldman 1965a). These authors studied this evolutionary process using Ae. peregrina as a model, and argued that the extensive morphological variation in this species was indicative of genome modification. They presented evidence for the existence of hybrids and hybrid derivatives between Ae. peregrina and several other allotetraploid species that share the U subgenome and grow with Ae. peregrina in mixed populations. Moreover, F₁ hybrids between Ae. peregrina and other U-subgenome allopolyploids exhibit, in addition to pairing of the homologous U-chromosomes, also pairing between several chromosomes of the differential genomes (Feldman 1965c), even though such pairing is usually precluded (Riley 1966b). Upon backcrossing to either parent, this pairing may lead to the formation of introgressed (modified) subgenomes.

Furuta and Tanaka (1970) examined whether introgression occurs between tetraploid species belonging to the U-subgenome group of Aegilops, by carrying out cytological and morphological analyses on hybrid progenies of three cross-combinations: Ae. neglecta (UUXⁿXⁿ) x Ae. columnaris (UUXⁿXⁿ), Ae. peregrina (UUS^vS^v) x Ae. columnaris, and Ae. biuncialis (UUM^bM^b) x Ae. columnaris. In all cases, the U subgenome was the common buffer subgenome and subgenomes Xⁿ, S^vand M^b form 5, 3.5 and 2 bivalents, respectively, with the Xⁿ subgenome. In the most closely related combination, Ae. neglecta x Ae. columnaris, introgression was cytologically and morphologically confirmed. In Ae. peregrina x Ae. columnaris, only introgression between two homoeologous chromosomes was observed, whereas in the third distinctly related combination, Ae. biuncialis x Ae. columnaris, no introgression took place. These observations suggest that introgression is a function of the relationship between species and impacts the resulting modified genomes in the Aegilops allotetraploid species (Furuta and Tanaka 1970).

Support for the above hypothesis came from Nakai and Tsuji (1984) who, using the gel electrofocusing technique, examined acid phosphatase isozymes in four accessions of Ae. peregrina and reported that the S^v subgenome of Ae. peregrina and Ae. kotschyi, had been modified by introgressive hybridization from Ae. geniculata and Ae. columnaris. In contrast, Jaaska (1978a, b) noted from his acid phosphatase data, no indication of the occurrence of subgenome recombination in Aegilops species sharing a common subgenome. However, his results did not necessarily overthrow Zohary and Feldman’s (1962) hypothesis, as the banding profiles of the allotetraploids presented may reflect complex evolutionary processes. Likewise, C-banding and FISH analyses performed by Badaeva et al. (2004), strongly suggested that the S^v subgenome of Ae. peregrina was derived from Ae. longissima, and that it is not structurally altered relative to that of the parental species. This agrees with molecular data of Zhang et al. (1992), who obtained no evidence for an extensive modification of the S^v subgenome relative to that of its diploid donor, Ae. longissima.

In contrast to the above, Gong et al. (2006) used 31 ISSR (inter-simple sequence repeat) primers to study genomic evolutions among 23 species of Aegilops. The results indicated that the genome constituents of the allopolyploid species had considerably changed through evolution compared with their ancestral diploid species. Genome U showed little alterations in U-containing allopolyploids, while others had undergone changes after allopolyploidization. Gong et al. (2006), found that Ae. peregrina and Ae. kotschyi were more similar to Ae. umbellulata than to Ae. searsii, despite the fact that Ae. searsii was their cytoplasmic donor (Terachi et al. 1990). They concluded that their findings supported the pivotal-differential hypothesis proposed by Zohary and Feldman (1962).

Ae. peregrina contains 12.52 ± 0.181 pg 1C DNA (Eilam et al. 2008), one of the largest genomes among the allotetraploid species of Aegilops (Eilam et al. 2008). The 1C DNA content is 2.64% less DNA than that expected from the sum of the DNA of its two diploid parents, namely, 12.86 pg (Ae. umbellulata, the donor of the U subgenome, contains 5.38 pg and Ae. longissima, the assumed donor of the S^v subgenome, contains 7.48 pg; Eilam et al. 2007).

The karyotype of Ae. peregrina described by Sorokina (1928) did not show the presence of satellites. However, according to Senyaninova-Korchagina (1930, 1932), there are 8 types of chromosomes, including one pair with satellites. Chennaveeraiah (1960) revealed three pairs with satellites on short arms, two pairs with large satellites and the third pair with smaller satellites. One pair has an extreme subterminal centromere, two pairs have submedian-subterminal centromeres, and the rest have submedian centromeres. No pairs have median centromere. In all, there are 12 types of chromosomes. One set of chromosomes, consisting of a chromosome pair with small satellites, a pair with large satellites, a pair with extreme subterminal centromere, two pairs with submedian centromeres and two pairs with submedian-subterminal centromeres, corresponds to the karyotype of the U genome of Ae. umbellulata. The second set of chromosomes contains one pair with large satellites, and the rest with submedian centromeres but with different arm ratio. Based on these karyotypic features, Chennaveeraiah (1960) concluded that this set falls into the M-group rather than the S-group.

A repetitive DNA sequence, derived from T. aestivum, coding for ribosomal RNA, was used as a probe in FISH analysis of Ae. peregrina and Ae. kotschyi genomes (Teoh et al. 1983). Similar to the finding of Chennaveeraiah (1960), Teoh et al. (1983) also found three pairs of SAT chromosomes and three pairs of rRNA sites. In contrast, Cermeño et al. (1984b) found only two SAT chromosomes and two pairs of Ag-NORs in Ae. peregrina. In parallel, they found up to four nucleoli in interphase cells and reported that the U subgenome completely suppressed the NOR activity of the Ae. peregrina S^v subgenome.

Badaeva et al. (2004) studied structure of the Ae. peregrina genome by analyzing heterochromatin banding patterns of its somatic metaphase chromosomes, as revealed by study of 20 accessions by C-banding and three accessions by FISH with the heterochromatin-limited repetitive DNA probes pSc119 and pAs1, as well as the distribution of NOR and 5S DNA loci revealed by pTa71 (18S-26S rDNA) and pTa794 (5S rDNA) probes. All Ae. peregrina chromosomes were highly heterochromatic and with distinct C-banding patterns, allowing their easy identification. Similar to Badaeva et al. (2004), Zhao et al. (2016) identified each of the 14 pairs of Ae. peregrina chromosomes using a FISH probe combination of pSc119.2, pTa71 and pTa-713. Using N Banding, Jewell and Driscoll (1983) succeeded to identify nine of the 14 possible chromosomes of Ae. peregrina that were added to common wheat as a monosomic addition.

Significant C-banding polymorphism was detected in Ae. peregrina (Badaeva et al. 2004). However, the frequency of chromosomal aberrations was comparatively low, which agrees with meiotic pairing data in intraspecific hybrids of this species (Feldman 1963; Furuta 1981b; Kawahara 1986, 1988; Yu and Jahier 1992). Rearranged chromosomes were only found in four accessions. These modified chromosomes likely arose as a result of introgression of genetic material from a related species, followed by meiotic recombination. (Badaeva et al. 2004).

Studies based on isozyme (Hart and Tuleen1983) and C-banding analyses (Friebe et al. 1993, 1996; Friebe and Gill 1996) revealed that chromosomes 4S^l and 7S^l of Ae. longissima are involved in a species-specific reciprocal translocation that is absent in Ae. sharonensis. The presence of this translocation in Ae. peregrina and its absence in Ae. kotschyi suggests that Ae. peregrina originated from the hybridization of Ae. umbellulata with Ae. longissima and that Ae. kotschyi originated from the hybridization of Ae. umbellulata with Ae. sharonensis; both hybridizations occurred after the differentiation of Ae. longissima from Ae. sharonensis.

While the U subgenome of Ae. peregrina is similar to that of Ae. umbellulata (Friebe et al. 1995b, 1996; Badaeva et al. 1996a), Badaeva et al. (2004) found some differences in the size and position of C-bands of the corresponding chromosomes. On the other hand, the S^v subgenome was nearly identical to the S^l genome of Ae. longissima, as also reported by Friebe et al. (1993), Friebe and Gill (1996), and Badaeva et al. (1996a), indicating that Ae. peregrina derived from hybridization of Ae. umbellulata with Ae. longissima. Little modification occurred in the S^v subgenome at the tetraploid level, contradicting earlier findings (Chennaveeraiah 1960; Zohary and Feldman 1962; Feldman 1965a).

Although Ae. peregrina has a genome constitution similar to that of Ae. kotschyi, the C-banding patterns of these two species were different (Badaeva et al. 2004). Comparison of Ae. kotschyi with the diploid ancestors Ae. umbellulata and Ae. sharonensis, revealed a higher degree of genome modification in Ae. kotschyi as compared to Ae. peregrina. Chromosomes 4 Sv and 7S^vof Ae. peregrina are nearly identical to 4S^l and 7S^l of Ae. longissima, whereas chromosomes 4S^vand 7S^vof Ae. kotschyi are similar to 4S^sh and 7S^sh of Ae. sharonensis. The high frequency of chromosomal aberrations and reduction in the number and size of 18S-26S rDNA loci observed in the S^v subgenome of Ae. kotschyi, compared to the S^sh genome of Ae. sharonensis and S^v of Ae. peregrina, suggest that Ae. kotschyi is an older species than Ae. peregrina.

FISH with clone pSC119 revealed signals of various sizes in telosomic regions of either the short or both arms of 13 Ae. peregrina chromosome pairs. An interstitial pSc119 FISH site was detected in the long arm of Ae. peregrina chromosome 7U, whereas chromosome 6U had no pSc119 FISH site, despite the fact that distinct telomeric and interstitial FISH sites are present in Ae. umbellulata (Badaeva et al. 1996a). Chromosome 6U of Ae. peregrina has a telomeric and an interstitial pAs1 FISH site, which are absent in Ae. umbellulata. Another a pAs1 FISH site was detected in the middle of the satellite of chromosome 1U of Ae. peregrina.

In contrast to Chennaveeraiah (1960), Badaeva et al. (2004) found only two pairs of satellite chromosomes in Ae. peregrina and Ae. kotschyi, which coincides with the number of active NORs detected by Ag-NOR staining (Cermeno et al. 1984b) and in situ hybridization (Yamamoto 1992a). In both species, major NORs were observed on group 1 and 5 from the U-subgenome chromosomes. Ae. peregrina and Ae. kotschyi also have several minor 18S-26S rDNA sites. Two consistent minor loci detected on group 5 and 6 of the S^v-subgenome chromosomes were associated with a significant reduction in copy number of 18S-26S rRNA genes. The hybridization patterns with pTa794 were similar in both species; four similar-sized rDNA loci were located on chromosomes of homoeologous groups 1 and 5. As was found by Cermeño et al. (1984b), the NORs on S^v subgenome chromosomes were inactivated and were accompanied with a decrease or loss of rDNA sequences.

9.8.10.3 Crosses with Other Species of the Wheat Group

Chennaveeraiah (1960) reported that all PMCs of Ae. peregrina had 14 bivalents at first meiotic metaphase, two of which were usually rod bivalents. No multivalents were observed. Likewise, Feldman (1963), who studied chromosome pairing in different lines of both Ae. peregrina subspecies, noted that all lines had 14 bivalents and 27.86 to 32.34 chiasmata/cell. Feldman (1963) also studied chromosome pairing at first meiotic metaphase in F₁ hybrids between different lines of Ae. peregrina. Average chromosome pairing in hybrids between lines of ssp. peregrina included 0.0–0.14 univalents, 10.0–14.0 bivalents, 0.0–0.04 trivalents, 0.0–1.96 quadrivalents and 26.10–26.74 chiasmata. Average chromosome pairing in the hybrid between two lines of ssp. cylindrostachys contained 0.12 univalents, 11.92 bivalents, of which one was heteromorphic, 0.04 trivalents, 0.98 quadrivalents and 26.80 chiasmata. Average chromosome pairing in F₁ hybrids between lines of ssp. peregrina x lines of ssp. cylindrostachys, presented 0.06–0.56 univalents, 12.02–12.304 bivalents, of which 0.02–0.46 were heteromorphic, 0.02–0.08 trivalents, 0.78–0.96 quadrivalents, and 26.04–26.46 chiasmata. Average chromosome pairing in the intraspecific hybrids was somewhat less regular than pairing in the parental lines, namely, though they had a low frequency of univalents, they had fewer bivalents, some multivalents and a lower number of chiasmata per cell.

Cuñado (1992) used C-banding to analyze chromosome pairing at first meiotic metaphase of Ae. peregrina. This technique facilitated the pairing of the whole complement as well as of specific-subgenome chromosomes. Ae. peregrina had an average of 0.04 univalents, 2.61 rod bivalents, 11.34 ring bivalents and 25.30 chromosome associations per cell (Cuñado 1992). U subgenome chromosome pairings showed 0.04 univalents, 1.41 rod bivalents, 5.54 ring bivalents, and 12.50 chromosome associations per cell, while S^v subgenome chromosome pairings included 1.20 rod bivalents, 5.80 ring bivalents and 12.80 chromosome associations per cell. Slight differences were observed between the pairing data of the two subgenomes, presumably because of the presence of one chromosome pair with a subterminal centromere in the U subgenome and higher C-heterochromatin content in the S^v subgenome. Cuñado et al. (1996b) also studied chromosome pairing in early stages of meiotic prophase of Ae. peregrina and found, in zygotene, 14 bivalents in 9 cells and 12 bivalents and one multivalent in one cell, whereas, in pachytene, all cells had 14 bivalents. These data indicate that bivalent pairing in Ae. peregrina occurs already at early stages of meiosis.

Chromosome pairing in F₁ triploid hybrids of Ae. peregrina with its putative diploid progenitors, Ae. umbellulata and Ae. longissima, are presented in Table 9.5. Pairing in peregrina x umbellulata included 6.95 univalents, 1.59 rod bivalents, 5.41 ring bivalents, 0.02 trivalents and 12.44 associations/cell. Pairing in peregrina x longissima showed 6.77 univalents, 2.11 rod bivalents, 2.98 ring bivalents, 0.30 trivalents, 0.73 quadrivalents, 0.04 other multivalents and 11.42 associations/cell. Similar results were obtained by Feldman (1963), who studied chromosome pairing in F₁ hybrids between several lines of Ae. peregrina and Ae. longissima. Average chromosome pairing included 6.58–6.72 univalents, 6.52–6.72 bivalents, 0.28–0.34 trivalents, and 14.36–15.12 chiasmata/cell. One hybrid combination had 6.82 univalents, 4.90 bivalents, 0.16 trivalents, 0.90 quadrivalents, 0.06 pentavalents and 13.08 chiasmata/cell. The above data substantiated that Ae. umbellulata and Ae. longissima are the parental species of Ae. peregrina. The S^v subgenome in two lines of Ae. peregrina (one studied by Yu and Jahier 1992 and the second by Feldman 1963) differed from the S^l genome of Ae. longissima by a reciprocal translocation. Thus, the U subgenome of Ae. peregrina remained nearly unchanged from that of Ae. umbellulata, whereas the S^v subgenome of Ae. peregrina underwent some changes compared to the S^l genome of Ae. longissima and is therefore structurally differentiated by at least one interchange (Yu and Jahier 1992).

Yu and Jahier (1992) also studied chromosome pairing in F₁ triploid hybrids between Ae. peregrina and three other diploid species of section Sitopsis, Ae. bicornis, Ae. searsii and a low-pairing line of Ae. speltoides (Table 9.5). Pairing in peregrina x bicornis exhibited 7.57 associations/cell, pairing in peregrina x searsii had 5.82 associations/cell, and pairing in the peregrina x speltoides hybrid had 4.31 associations/cell. The relatively low level of pairing in these three hybrids showed that the genomes of these three Sitopsis species are only homoeologous to the S^v subgenome of Ae. peregrina.

Feldman (1965c) studied chromosome pairing in hybrids between Ae. peregrina and two other allopolyploid species of Aegilops sharing the U-subgenome, Ae. geniculata and Ae. biuncialis (Table 9.7). The F₁ of the Ae. biuncialis x Ae. peregrina ssp. peregrina hybrid (genome UM^bUS^v) showed 12.16 univalents, 6.54 bivalents, 0.76 trivalents and 0.12 quadrivalents, and the F₁ Ae. biuncialis x Ae. peregrina ssp. cylindrostachys hybrid showed 11.82 univalents, 6.60 bivalents, 0.70 trivalents, and 0.22 quadrivalents. The F₁ Ae. geniculata x Ae. peregrina ssp. peregrina hybrid (genome M^oUUS^v) had 11.44 univalents, 7.02 bivalents and 0.84 trivalents, and the F₁ Ae. geniculata x Ae. peregrina ssp. cylindrostachys hybrid had 11.42 univalents, 6.63 bivalents, 1.00 trivalent and 0.08 quadrivalent. Kihara (1937) observed 6–8 bivalents in the F₁ hybrid Ae. geniculata x Ae. peregrina and 7 bivalents in the F₁ hybrid Ae. biuncialis x Ae. peregrina, and Kihara (1949) observed 7–11 bivalents in the Ae. neglecta x Ae. peregrina hybrids, Lindschau and Oehler (1936) observed 2–12 bivalents in Ae. peregrina x Ae. triuncialis (genome S^vUUC) and Kihara (1937) observed 7–9 bivalents in the reciprocal combination. All these hybrids had more than 7 bivalents, indicating that pairing also took place between chromosomes of the differential genomes of these species.

Cuñado (1993a) also analyzed chromosome pairing in the F₁ Ae. triuncialis x Ae. peregrina hybrid and observed 12.74 univalents, 3.82 rod bivalents, 2.66 ring bivalents, 0.66 trivalents, 0.08 quadrivalents and 10.74 association/cell. The S^v—subgenome chromosomes were distinguishable from the U and C chromosomes by their larger size and higher C-heterochromatin content, which facilitated determination of the homomorphic and heteromorphic pairing in the hybrid. Average homomorphic pairing between the homologous chromosomes of the U subgenome, showed 2.04 rod bivalents, 2.50 ring bivalents and 6.66 associations/cell, and heteromorphic pairing between the homoeologous chromosomes of the C and S^v differential subgenomes included 1.78 rod bivalents, 0.16 ring bivalents, 0.66 trivalents, 0.08 quadrivalents and 3.58 associations/cell.

Meiotic pairing was analyzed by Fernández-Calvín and Orellana (1991) at first meiotic metaphase of the F₁ pentaploid hybrids common wheat x Ae. peregrina ssp. peregrina, common wheat x ssp. cylindrostachys, and common wheat x Ae. kotschyi. The hybrids had either the Ph1 gene of common wheat that induces low pairing in hybrids or the ph1b mutant that allows homoeologous pairing in hybrids. The use of C-banding technique enabled identification of the various chromosomes in the different pairing configurations. Chromosome pairing in the hybrid common wheat x Ae. peregrina ssp. peregrina var. typica (hybrid genome BADUS^v) included 30.99 univalents, 1.69 rod bivalents, 0.10 ring bivalents, 0.03 trivalents and 0.01 quadrivalents. Among the rod bivalents, 0.33 were between A and D chromosomes, 0.67 were between AD-US^v chromosomes, 0.01 were between AD-B chromosomes, 0.37 were between US^v-B chromosomes and 0.31 were between U-S^v chromosomes. Chromosome pairing in the hybrid common wheat x Ae. peregrina ssp. cylindrostachys presented 28.99 univalents, 2.81 rod bivalents, 0.06 ring bivalents and 0.10 trivalents. Among the rod bivalents, 0.60 were between A-D chromosomes, 1.20 between AD-US^v chromosomes, 0.14 between AD-B chromosomes, 0.42 between US^vB chromosomes, and 0.45 between U-S^v chromosomes. Chromosome pairing in the hybrid common wheat x Ae. kotschyi (hybrid genome BADUS^v) had 30.92 univalents, 1.98 rod bivalents, 0.02 ring bivalents and 0.03 trivalents. Among the rod bivalents, 0.54 were between A-D chromosomes, 0.74 between AD-US^v chromosomes, 0.05 between AD-B chromosomes, 0.38 between US^v-B chromosomes and 0.25 between U-S^v chromosomes. These data indicated that more than 40% of the pairing in these three hybrids was autosyndetic (A-D + AD-B = 25%, and U-S^v = 16%) and more than 50% was allosyndetic (AD-US^v + US^v-B = 51%) (Fernández-Calvín and Orellana 1991). The allosyndetic pairing between AD chromosomes and US^v chromosomes was twice as frequent as that between US^lvand B chromosomes. The genotype of Ae. peregrina ssp. cylindrostachys seemed to promote homoeologous pairing in the hybrid with common wheat (an increase of approximately a bivalent/cell). Its effect was detectable in the low pairing, but not in the high pairing hybrid (Fernández-Calvín and Orellana 1991).

Evidence for the existence of a gene(s) promoting pairing in a line of Ae. peregrina was obtained by Farooq et al. (1990) who analyzed chromosome pairing in hybrids of common wheat x three different accessions (A, B, and E) of Ae. peregrina. These authors found significant differences in the frequencies of homoeologous chromosome pairing at first meiotic metaphase. Hybrids between common wheat and Ae. peregrina accessions A and B showed very little pairing, as indicated by a chiasma frequency of 1.0 and 1.5 per cell, respectively. On the other hand, the hybrid between common wheat and Ae. peregrina accession E, showed significantly more homoeologous pairing (mean chiasma frequency was 12.6/cell). The level of such pairing was essentially the same as that between the hybrids of common wheat ph1b x Ae. peregrina accessions A and B. However, when the ph1b mutant was hybridized with accession E, the level of chromosome pairing significantly increased further (mean chiasma frequency was 17.52/cell), indicating the presence of pairing promoter gene(s) in Ae. peregrina accession E, which are epistatic to the wheat Ph1 allele, and which positively interact with its mutant form to further increase the ph1b ceiling to homoeologous pairing in wheat.