Plant Systematics and Evolution

, Volume 301, Issue 2, pp 749–759 | Cite as

Genetic insights into the hybrid origin of Abies × borisii-regis Mattf.

  • Eleni Bella
  • Sascha Liepelt
  • Laura Parducci
  • Andreas D. Drouzas
Original Article


Abies × borisii-regis Mattf. (King Boris fir) is a taxon endemic to the southern Balkan Peninsula, described as a hybrid between the widespread A. alba Mill. (Silver fir) and the Greek endemic A. cephalonica Loud (Greek fir). Even though A. × borisii-regis has attracted much research attention in the past, its origin, geographical distribution and taxonomic status are not fully elucidated and molecular evidence for hybridization is missing. To shed more light on this issue, we analyzed representative populations from these three Abies taxa using paternally inherited (chloroplast) and maternally inherited (mitochondrial) DNA markers. Both Silver and Greek fir could be clearly distinguished using mitochondrial markers, while we observed a mixture of maternal lineages in the A. × borisii-regis populations. In contrast, using chloroplast markers, we could not identify species-specific haplotypes, but a neighbor-joining analysis of population genetic distances revealed two separate clusters for the Silver fir and the Greek fir, while the A. × borisii-regis populations were placed in intermediate positions. Our results are in agreement with the hypothesis that the A. × borisii-regis populations investigated are a result of hybridization between A. cephalonica and A. alba.


Abies cephalonica A. alba A. × borisii-regis Hybrid cpDNA mtDNA 



The present work was part of the Postgraduate Diploma Thesis of EB, in the Postgraduate Studies Programme “Conservation of Biodiversity and Sustainable Exploitation of Native Plants” (, School of Biology, Aristotle University of Thessaloniki, Greece), which partially funded this work. The SSR work was carried out in the facilities of the Department of Genetics, Development and Molecular Biology of the School of Biology and the authors are expressing their gratitude to Prof. Theodore Abatzopoulos for the use of the LI-COR® 4200L DNA Analyzer, to Prof. Anastasia Kouvatsi for the use of gel documentation equipment and to Ass. Prof. Alexandros Triantafyllidis and Dr. Nikoleta Karaiskou for their contribution in the analyses. ADD sincerely thanks the Forest Services in Amfissa, Vytina, Sparti, Konitsa, Metsovo, Cephalonia and Kalavryta, as well as the Management Authority of the Aenos National Park in Cephalonia island (Greece) for the information and the support provided during the sampling. The authors also thank two anonymous reviewers for their constructive comments and suggestions to the manuscript.

Supplementary material

606_2014_1113_MOESM1_ESM.pdf (58 kb)
Supplementary material 1 (PDF 57 kb)
606_2014_1113_MOESM2_ESM.pdf (248 kb)
Supplementary material 2 (PDF 248 kb)


  1. Arnold ML (1997) Natural hybridization and evolution. Oxford University Press, OxfordGoogle Scholar
  2. Athanassiadis NH (1986) Forest botany (part II): trees and shrubs of Greece. Giahoudis-Giapoulis Publications, Thessaloniki (in Greek)Google Scholar
  3. Chater AO (1964) Abies Miller. In: Tutin TG, Heywood VH, Burges NA, Valentine DH, Walters SM, Webb DA (eds) Flora Europaea, vol 1. Cambridge University Press, Cambridge, pp 37–38Google Scholar
  4. Christensen KI (1986) Abies Miller. In: Strid A (ed) Mountain flora of Greece, vol 1. Cambridge University Press, Cambridge, pp 38–41Google Scholar
  5. Christensen KI (1997) Abies Miller. In: Strid A, Tan K (eds) Flora Hellenica, vol 1. Koeltz Scientific Books, Königstein, pp 1–3Google Scholar
  6. Clement M, Posada D, Crandall KA (2000) TCS: a computer program to estimate gene genealogies. Mol Ecol 9:1657–1660PubMedCrossRefGoogle Scholar
  7. Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15Google Scholar
  8. Drouzas AD (2000) Determination and analysis of variation of Greek fir species (Abies sp.), by using biochemical and molecular markers. Doctoral thesis, Aristotle University of Thessaloniki (in Greek, English summary)Google Scholar
  9. Du Rietz GE (1930) The fundamental units of biological taxonomy. Svensk Botanisk Tidskrift 24:333–428Google Scholar
  10. Excoffier L, Lischer HEL (2010) Arlequin suite ver 3.5: a new series of programs to perform population genetics analyses under Linux and Windows. Mol Ecol Resour 10:564–567. doi: 10.1111/j.1755-0998.2010.02847.x PubMedCrossRefGoogle Scholar
  11. Fady B, Conkle MT (1993) Allozyme variation and possible phylogenetic implications in Abies cephalonica Loudon and some related Eastern Mediterranean firs. Silvae Genet 42:351–359Google Scholar
  12. Fady B, Arbez M, Ferrandès P (1991) Variability of juvenile Greek firs and stability of characteristics with age. Silvae Genet 40:91–100Google Scholar
  13. Farjon A, Rushforth KD (1989) A classification of Abies Miller (Pinaceae). Notes R Bot Gard Edinb 46:59–79Google Scholar
  14. Felsenstein J (2005) PHYLIP (phylogeny inference package) version 3.6. Distributed by the author. Department of Genome Sciences, University of Washington, SeattleGoogle Scholar
  15. Gao J, Wang B, Mao JF, Ingvarsson P, Zeng QY, Wang XR (2012) Demography and speciation history of the homoploid hybrid pine Pinus densata on the Tibetan plateau. Mol Ecol 21:4811–4827. doi: 10.1111/j.1365-294X.2012.05712.x PubMedCrossRefGoogle Scholar
  16. Georgolopoulos G, Drouzas AD (2011) Species identification in the Pinaceae family by using molecular markers. In: Peraki E, Kopaka K, Pyrintsos S (eds) Abstracts book of the 12th Congress of the Hellenic Botanical Society, Rethymno, pp 50–51Google Scholar
  17. Gömöry D, Longauer R, Liepelt S, Ballian D, Brus R, Kraigher H, Parpan VI, Parpan TV, Paule L, Stupar VI, Ziegenhagen B (2004) Variation patterns of mitochondrial DNA of Abies alba Mill. in suture zones of postglacial migration in Europe. Acta Soc Bot Pol 73:203–206. doi: 10.5586/asbp.2004.027 CrossRefGoogle Scholar
  18. Gömöry D, Paule L, Krajmerova D, Romsakova I, Longauer R (2012) Admixture of genetic lineages of different glacial origin: a case study of Abies alba Mill. in the Carpathians. Plant Syst Evol 298:703–712. doi: 10.1007/s00606-011-0580-6 CrossRefGoogle Scholar
  19. Hatziskakis S, Papageorgiou AC, Gailing O, Finkeldey R (2009) High chloroplast haplotype diversity in Greek populations of beech (Fagus sylvatica L.). Plant Biol 11:425–433. doi: 10.1111/j.1438-8677.2008.00111.x PubMedCrossRefGoogle Scholar
  20. Konnert M, Bergmann F (1995) The geographical distribution of genetic variation of silver fir (Abies alba, Pinaceae) in relation to its migration history. Plant Syst Evol 196:19–30. doi: 10.1007/BF00985333 CrossRefGoogle Scholar
  21. Kormutak A (1985) Study on species hybridization within the genus Abies. Acta dendrobiologica. VEDA, BratislavaGoogle Scholar
  22. Kormutak A (2004) Crossability relationships between some representatives of the Mediterranean, Northamerican and Asian firs (Abies sp.).VEDA, BratislavaGoogle Scholar
  23. Liepelt S, Kuhlenkamp V, Anzidei M, Vendramin GG, Ziegenhagen B (2001) Pitfalls in determining size homoplasy of microsatellite loci. Mol Ecol Notes 1:332–335. doi: 10.1046/j.1471-8278.2001.00085.x CrossRefGoogle Scholar
  24. Liepelt S, Bialozyt R, Ziegenhagen B (2002) Wind-dispersed pollen mediates post-glacial gene flow among refugia. Proc Natl Acad Sci USA 99:14590–14594. doi: 10.1073/pnas.212285399 PubMedCentralPubMedCrossRefGoogle Scholar
  25. Liepelt S, Cheddadi R, De Beaulieau JL, Fady B, Gömöry D, Hussendörfer E, Konnert M, Litt M, Longauer R, Terhürne-Berson R, Ziegenhagen B (2009) Postglacial range expansion and its genetic imprints in Abies alba (Mill.)—a synthesis from palaeobotanic and genetic data. Rev Palaeobot Palynol 153:139–149. doi: 10.1016/j.revpalbo.2008.07.007 CrossRefGoogle Scholar
  26. Liepelt S, Mayland-Quellhorst E, Lahme M, Ziegenhagen B (2010) Contrasting geographical patterns of ancient and modern genetic lineages in Mediterranean Abies species. Plant Syst Evol 284:141–151. doi: 10.1007/s00606-009-0247-8 CrossRefGoogle Scholar
  27. Liu TS (1971) A monograph of the genus Abies. National Taiwan University, TaipeiGoogle Scholar
  28. Magri D, Vendramin GG, Comps B, Dupanloup I, Geburek T, Gomory D, Latalowa M, Litt T, Paule L, Roure JM, Tantau I, van der Knaap WO, Petit RJ, de Beaulieu JL (2006) A new scenario for the Quaternary history of European beech populations: palaeobotanical evidence and genetic consequences. New Phytol 71:199–221. doi: 10.1111/j.1469-8137.2006.01740.x CrossRefGoogle Scholar
  29. Manni F, Guérard E, Heyer E (2004) Geographic patterns of (genetic, morphologic, linguistic) variation: how barriers can be detected by “Monmonier’s algorithm”. Hum Biol 76:173–190. doi: 10.1353/hub.2004.0034 PubMedCrossRefGoogle Scholar
  30. Mattfeld J (1926) Die Europäischen und Mediterranen Abies Arten. Die Pflanzenareale 1:22–29 (in German)Google Scholar
  31. Mattfeld J (1930) Uber hybridogene Sippen der Tanne. Bibliotheca Botanika 100:1–84 (in German)Google Scholar
  32. Mergen F, Burley J (1964) Abies karyotype analysis. Silvae Genet 13:63–68Google Scholar
  33. Mitsopoulos DJ, Panetsos CP (1987) Origin of variation in fir forests of Greece. Silvae Genet 36:1–15Google Scholar
  34. Moulalis D (1986) Diagnosis and characterization of fir hybrids. Scientific Annals of the Department of Forestry and Natural Environment, Aristotle University of Thessaloniki, vol 29(ΚΘ)-7, pp 371–404Google Scholar
  35. Page RDM (1996) TreeView: an application to display phylogenetic trees on personal computers. Comput Appl Biosci 12:357–358. doi: 10.1093/bioinformatics/12.4.357 PubMedGoogle Scholar
  36. Panetsos CP (1975) Monograph of Abies cephalonica Loudon. Ann For 7:1–22Google Scholar
  37. Panetsos KP (1990) Species-provenance test of Mediterranean firs. In: Ducrey M, Oswald H (eds) Proceedings of the international workshop on: Mediterranean firs—adaptation, selection and silviculture”, Avignon. EUR 13491, pp 29–42Google Scholar
  38. Panetsos KP (1992) Variation in the position of resin canals in the needles of Abies species and provenances. Ann For Sci 49:253–260. doi: 10.1051/forest:19920304 CrossRefGoogle Scholar
  39. Parducci L, Szmidt AE (1999) PCR-RFLP analysis of cpDNA in the genus Abies. Theor Appl Genet 98(5):802–808. doi: 10.1007/s001220051137
  40. Parducci L, Szmidt AE, Madaghiele A, Anzidei M, Vendramin GG (2001) Genetic variation at chloroplast microsatellites (cpSSRs) in Abies nebrodensis (Lojac.) Mattei and three neighboring Abies species. Theor Appl Genet 102:733–740. doi: 10.1007/s001220051704 CrossRefGoogle Scholar
  41. Peakall R, Smouse PE (2006) GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Notes 6:288–295. doi: 10.1111/j.1471-8286.2005.01155.x CrossRefGoogle Scholar
  42. Petit RJ, Brewer S, Bordács S, Burg K, Cheddadi R, Coart E, Cottrell J, Csaikl UM, van Dam B, Deans JD, Fineschi S, Finkeldey R, Glaz I, Goicoechea PG, Jensen JS, König AO, Lowe AJ, Madsen SF, Mátyás G, Munro RC, Popescu F, Slade D, Tabbener H, de Vries SMG, Ziegenhagen B, de Beaulieu JL, Kremer A (2002) Identification of refugia and post-glacial colonisation routes of European white oaks based on chloroplast DNA and fossil pollen evidence. For Ecol Manag 156:49–74. doi: 10.1016/S0378-1127(01)00634-X CrossRefGoogle Scholar
  43. Pons O, Petit RJ (1996) Measuring and testing genetic differentiation with ordered versus unordered alleles. Genetics 144:1237–1245PubMedCentralPubMedGoogle Scholar
  44. Riesenberg LH, Wendel J (1993) Introgression and its consequences in plants. In: Harrison R (ed) Hybrid zones and the evolutionary process. Oxford University Press, New York, pp 70–109Google Scholar
  45. Saitou N, Nei M (1987) The neighbour-joining method: a new model for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425PubMedGoogle Scholar
  46. Scaltsoyiannes A, Tsaktsira M, Drouzas AD (1999) Allozyme differentiation in the Mediterranean firs (Abies, Pinaceae). A first comparative study with phylogenetic implications. Plant Syst Evol 216:289–307. doi: 10.1007/BF01084404 CrossRefGoogle Scholar
  47. Semerikova SA, Semerikov VL, Lascoux M (2011) Post-glacial history and introgression in Abies (Pinaceae) species of the Russian Far East inferred from both nuclear and cytoplasmic markers. J Biogeogr 38:326–340. doi: 10.1111/j.1365-2699.2010.02394.x CrossRefGoogle Scholar
  48. Stebbins GL (1950) Variation and evolution in plants. Columbia University Press, New YorkGoogle Scholar
  49. Stebbins GL (1969) The significance of hybridization for plant taxonomy and evolution. Taxon 18:26–35CrossRefGoogle Scholar
  50. Strid A, Tan K (1997) Flora Hellenica, vol I. Koeltz Scientific Books, KönigsteinGoogle Scholar
  51. Taberlet P, Fumagalli L, Wust-Saucy AG, Cossons J-F (1998) Comparative phylogeography and post-glacial colonization routes in Europe. Mol Ecol 7:453–464. doi: 10.1046/j.1365-294x.1998.00289.x PubMedCrossRefGoogle Scholar
  52. Turrill WB (1937) XI-On the flora of the near east: XVIII. New species, new records and notes. Kew Bull 2:79–86Google Scholar
  53. Tzedakis PC (1993) Long-term tree populations in northwestern Greece trough multiple Quaternary climatic cycles. Nature 364:437–440. doi: 10.1038/364437a0 CrossRefGoogle Scholar
  54. Tzedakis PC, Lawson IT, Frogley MR, Hewitt GM, Preece RC (2002) Buffered tree population changes in a quaternary refugium: evolutionary implications. Science 297:2044–2047. doi: 10.1126/science.1073083 PubMedCrossRefGoogle Scholar
  55. Vendramin GG, Lelli L, Rossi P, Morgante M (1996) A set of primers for the amplification of 20 chloroplast microsatellites in Pinaceae. Mol Ecol 5:595–598. doi: 10.1111/j.1365-294X.1996.tb00353.x PubMedCrossRefGoogle Scholar
  56. Wagner DB (1992) Nuclear, chloroplast, and mitochondrial DNA polymorphisms as biochemical markers in population genetic analyses of forest trees. New For 6:373–390. doi: 10.1007/BF00120653 CrossRefGoogle Scholar
  57. Wang XR, Tsumura Y, Yoshimaru H, Nagasaka K, Szmidt AE (1999) Phylogenetic relationships of Eurasian pines (Pinus, Pinaceae) based on chloroplast rbcL, matK, rpl20-rps18 spacer and trnV intron sequences. Am J Bot 86:1742–1753PubMedCrossRefGoogle Scholar
  58. Wang XR, Szmidt AE, Savolainen O (2001) Genetic composition and diploid hybrid speciation of a high mountain pine, Pinus densata, native to the Tibetan plateau. Genetics 159:337–346PubMedCentralPubMedGoogle Scholar
  59. Wang B, Mao JF, Gao J, Zhao W, Wang XR (2011) Colonization of the Tibetan Plateau by the homoploid hybrid pine Pinus densata. Mol Ecol 20:3796–3811. doi: 10.1111/j.1365-294X.2011.05157.x PubMedCrossRefGoogle Scholar
  60. Watano Y, Imazu M, Shimizu T (1996) Spatial distribution of cpDNA and mtDNA haplotypes in a hybrid zone between Pinus pumila and P. parviflora var. pentaphylla (Pinaceae). J Plant Res 109:403–408. doi: 10.1007/BF02344555 CrossRefGoogle Scholar
  61. Ziegenhagen B, Fady B, Kuhlenkamp V, Liepelt S (2005) Differentiating groups of Abies species with a simple molecular marker. Silvae Genet 54:123–126Google Scholar

Copyright information

© Springer-Verlag Wien 2014

Authors and Affiliations

  • Eleni Bella
    • 1
    • 2
  • Sascha Liepelt
    • 3
  • Laura Parducci
    • 4
  • Andreas D. Drouzas
    • 2
  1. 1.Postgraduate Studies Program “Conservation of Biodiversity and Sustainable Exploitation of Native Plants (BNP)”, School of BiologyAristotle University of ThessalonikiThessaloníkiGreece
  2. 2.Laboratory of Systematic Botany and Phytogeography, School of BiologyAristotle University of ThessalonikiThessaloníkiGreece
  3. 3.Conservation BiologyUniversity of MarburgMarburgGermany
  4. 4.Department of Ecology and Genetics, Plant Ecology and Evolution, Evolutionary Biology CentreUppsala UniversityUppsalaSweden

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