Introduction

Shearer (1993a) reviewed the freshwater ascomycota and provided a definition and methods to study their taxonomy, systematics, geographical distribution, ecology, and evolution. This definition and the study techniques have since commonly been used (e.g., Zhang et al. 2017a; Dong et al. 2018; Lu et al. 2018b; Luo et al. 2019; Yang et al. 2019b). Unlike other fungi, which can be often traced to a specific host, freshwater fungi have mostly been collected from “submerged wood” (Wei et al. 2018; Yu et al. 2018; Wang et al. 2019). Thus, there has been little research on the hosts of freshwater fungi. The geographical distribution of freshwater fungi is also unclear and little can be concluded as distribution patterns are still largely based on the locations of mycologists (Shearer 1993a); thus further work are needed in these areas.

Dothideomycetes and Sordariomycetes, the largest classes of Ascomycota, have been comprehensively reviewed (e.g., Hyde et al. 2013, 2020c; Maharachchikumbura et al. 2016; Hongsanan et al. 2020a, b). Some websites, e.g. (http://fungi.life.illinois.edu/) (Shearer and Raja 2010), (https://www.marinefungi.org/) (Jones et al. 2019), (https://www.dothideomycetes.org/) (Pem et al. 2019a) and (https://www.freshwaterfungi.org/) (Calabon et al. 2020), are devoted to taxonomy and classification of Dothideomycetes, provide a database of freshwater ascomycetes and online resources for marine fungi. There has been an increased interest in the taxonomy and phylogeny of freshwater fungi (Lu et al. 2018b; Yang et al. 2018; Bao et al. 2019b, c; Boonmee et al. 2019), but further research will be carried out. Luo et al. (2019) gave accounts of freshwater Sordariomycetes and provided a modified backbone tree and detailed information on distribution, holotypes, sequence data and sexual-asexual morph connections of taxa. They also introduced 47 new taxa based on fresh collections from China and Thailand, which indicated there is a high diversity of freshwater Sordariomycetes.

In this article, a comprehensive study on freshwater Dothideomycetes is carried out based on new collections from China, Egypt and Thailand, as well as studies of specimens in fungal herbaria and literature reviews. Multigene phylogenetic analyses and morphological studies have resulted in nine new genera, 33 new species, 16 new combinations and 17 new geographical and habitat records being reported. Considering that many freshwater fungi are scattered in Dothideomycetes and the study of freshwater fungi have greatly advanced, there is a need to revise all freshwater Dothideomycetes with updated knowledge, and to provide a better understanding of their morphology and phylogeny. This monograph on freshwater Dothideomycetes provides the latest generic concepts, descriptions, illustrations and keys to genera and species. Using data from this study, we present several common morphological traits that appear to be adaptations of microfungi that live in flowing freshwater. Additionally, the global distribution of freshwater Dothideomycetes is reviewed in this study to show the poorly studied areas.

Materials and methods


Sample collection and specimen examination


Submerged wood was mostly collected from lentic or lotic streams, lakes or rivers in China and Thailand as part of a north-south survey (Hyde et al. 2016a), and from the river Nile in Egypt between 2010 to 2019. The principle of processing samples, including collection, incubation, isolation and examination, following Shearer (1993a). The detailed procedure of single spore isolation follows Chomnunti et al. (2014). Pseudoparaphyses are observed at 100× using a Nikon ECLIPSE Ni compound microscope fitted with a Canon EOS 600D digital camera. Two types of pseudoparaphyses, cellular and trabeculate, are defined if they are clearly seen. Herbarium specimens are deposited in the herbarium of Mae Fah Luang University (MFLU), Chiang Rai, Thailand and the herbarium of Cryptogams Kunming Institute of Botany Academia Sinica (KUN-HKAS), Kunming, China. Living cultures are deposited in the Mae Fah Luang University Culture Collection (MFLUCC), Chiang Rai, Thailand and Kunming Institute of Botany culture collection (KUMCC), Kunming, China. Facesoffungi and Index Fungorum numbers are registered as in Jayasiri et al. (2015) and Index Fungorum (2020).


DNA extraction, PCR amplification and sequencing

Cultures were grown on PDA at room temperature (25–27 °C) and a Biospin Fungus Genomic DNA Extraction Kit (Bioer Technology Co., Hangzhou, P.R. China) was used to extract total genomic DNA from the fresh mycelium according to the manufacturer’s instructions. Fragments of five loci were amplified with polymerase chain reaction (PCR). Primer pairs LR0R/LR5, NS1/NS4, ITS5/ITS4, EF1-EF1-983F/EF1-2218R and fRPB2-5F/fRPB2-7cR are used for LSU, SSU, ITS, TEF and RPB2, respectively. The amplifications were carried out using the method described by Zhang et al. (2017a). The PCR products were viewed on 1% agarose electrophoresis gels stained with ethidium bromide. Sequencing of five loci were carried out by Shanghai Sangon Biological Engineering Technology & Services Co. Shanghai, P.R. China.


Phylogenetic analysis

The sequences generated in this study were supplemented with other Dothideomycetous sequences obtained from GenBank, based on blast searches and relevant literatures. The accession numbers used in the multigene analyses were provided in supplementary material 1 and newly obtained strains with their accession numbers were shown in supplementary material 2. Multiple sequence alignments were generated with MAFFT v. 7 (https://mafft.cbrc.jp/alignment/server/index.html) (Kuraku et al. 2013; Katoh et al. 2019). The alignments were visually improved with BioEdit (Hall 1999). Phylogenetic analyses of the combined aligned dataset consisted of maximum likelihood (ML) and Bayesian inference (BI). Maximum likelihood (ML) analysis was performed at the CIPRES Science Gateway v.3.3 (https://www.phylo.org/portal2/home.action) (Miller et al. 2010). The final tree was selected among suboptimal trees from each run by comparing likelihood scores under the GTRGAMMA substitution model. The best-fit models for the Bayesian analyses were selected with MrModeltest v. 2.2 (Nylander 2004). The best-fit model GTR+I+G for LSU, ITS and RPB2, and HKY+I+G for TEF. Six simultaneous Markov chains were run for one million generations and trees were sampled every 100th generation and 10000 trees were obtained. The first 2500 trees representing the burn-in phase of the analyses were discarded, while the remaining 7500 trees were used for calculating posterior probabilities in the majority rule consensus tree. Phylogenetic tree was visualized using FigTree v1.4.0 (http://tree.bio.ed.ac.uk/software/figtree/, Rambaut 2012). Sequences derived in this study are deposited in GenBank.

Results

Phylogenetic analysis of combined LSU, ITS, TEF and RPB2 sequence data

The combined LSU, ITS, TEF and RPB2 sequence dataset were employed for species of Dothideomycetes. The alignment comprised 714 strains with an alignment length of 5882 total characters including gaps. The RAxML analysis resulted in a best scoring likelihood tree selected with a final value for the combined dataset ln L = − 216610.299349. The matrix has 4462 distinct alignment patterns, with 70.36% of undetermined characters or gaps. Estimated base frequencies are as follows: A = 0.238070, C = 0.259088, G = 0.278997, T = 0.223845; substitution rates AC = 1.423061, AG = 3.547230, AT = 1.643052, CG = 1.083323, CT = 7.578123, GT = 1.000000; gamma distribution shape parameter a = 0.439170 (Fig. 1).

Fig. 1
figure 1

World distribution of freshwater Dothideomycetes. Twelve morphologically identified freshwater species which occur worldwide are shown with different symbols on the map. The number of freshwater dothideomycetous species in each country is shown with different colours, which indicates “richness” from zero to almost 150 species (in China)

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Taxonomy

In this section, each freshwater genus is treated with a generic description and notes. A list with freshwater distribution and a key to freshwater species within each genus are provided. A key to freshwater genera within each family is also provided. An illustration of new species, new collections or line-drawings is provided after each entry, where possible. The freshwater distribution mainly follows an online resource for freshwater fungi (http://fungi.life.illinois.edu/) and published papers. The species is marked with an asterisk if there are published sequences deposited in GenBank. Freshwater Dothideomycetes are distributed in six orders, viz. Jahnulales, Kirschsteiniotheliales, Minutisphaerales, Natipusillales, Pleosporales and Tubeufiales; a few are scattered in Dothideomycetes families/genera incertae sedis. Classification and list of freshwater Dothideomycetes are shown in Table 1.

Table 1 Classification and list of freshwater Dothideomycetes
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*Denotes species having sequence data in GenBank


Jahnulales K.L. Pang, Abdel-Wahab, El-Shar., E.B.G. Jones & Sivichai, Mycol. Res. 106(9): 1033 (2002)


Aliquandostipitaceae Inderb., Am. J. Bot. 88(1): 54 (2001)


Key to freshwater sexual genera of Aliquandostipitaceae

  1. 1.

    Ascomata hyaline or slightly pigmented………2

  2. 1.

    Ascomata dark………3

  3. 2.

    Ascospores with longitudinal sulcate striations, without appendages or sheath………Megalohypha

  4. 2.

    Ascospores lacking longitudinal sulcate striations, often with a large sheath………Aliquandostipite

  5. 3.

    Peridium composed of single row of cells………………………Neojahnula

  6. 3.

    Peridium not as above………4

  7. 4.

    Ascospores without sheath or appendage………Jahnula

  8. 4.

    Ascospores with sheath or appendage………5

  9. 5.

    Ascospores surrounded by a wavy mucilaginous sheath………Pseudojahnula

  10. 5.

    Ascospores mostly with a refractive mucilaginous pad at each end………Ascagilis


Aliquandostipite Inderbitzin, Am. J. Bot. 88(1): 54 (2001)

Saprobic on submerged wood or fallen, decorticated branch. Sexual morph: Mycelium visible on the substratum, of wide hyphae (> 10 µm and up to 50 µm wide), which may bear ascomata. Ascomata scattered to clustered or gregarious, immersed to erumpent or superficial, uniloculate, globose to broadly ellipsoidal, dimorphic, with or without stalk-like hypha attached to the base, hyaline or pale brown when young, turn to dark brown with age, membranous, papillate. Peridium thin, one-layered, composed of pale brown, thin-walled cells of textura angularis to globosa. Pseudoparaphyses numerous, cellular, sparsely branched, hyaline, septate. Asci 8-spored, bitunicate, fissitunicate, clavate, with thickened apical region. Ascospores bi- to multi-seriate, variably arranged, oval, pale brown, 1-septate, constricted at the septum, usually asymmetric, mostly with a well-developed hyaline sheath (Inderbitzin et al. 2001). Asexual morph: Undetermined.

Type species: Aliquandostipite khaoyaiensis Inderb.

Notes: Aliquandostipite was introduced to accommodate two species A. khaoyaiensis Inderb. and A. sunyatsenii Inderb., which form dimorphic ascomata (sessile and stalked) and the widest hyphae known from ascomycetes (Inderbitzin et al. 2001). However, with Jahnula sequences added in the phylogenetic tree, A. sunyatsenii clustered in Jahnula with high bootstrap support, which was therefore transferred to Jahnula based on limited data (Pang et al. 2002). Among six species recorded in Index Fungorum (2020), four have been confirmed in Aliquandostipite with molecular data (Huang et al. 2018; Hyde et al. 2019). Aliquandostipite is similar to Jahnula, but can be distinguished from the latter by its hyaline or less pigmented ascomata and larger asci and ascospores.

According to our observations and literature, the ascospores of A. crystallinus Raja et al., A. khaoyaiensis and A. minuta Raja & Shearer form acicular crystals when stored in lactic acid and glycerin (Raja et al. 2005; Raja and Shearer 2007). These three taxa, however, can be distinguished based on the gelatinous sheath and appendages on ascospores. Aliquandostipite siamensiae (Sivichai & E.B.G. Jones) J. Campb. et al. is the only species producing dimorphic ascospores. Aliquandostipite manochiae Sri-indr. et al. is distinct in that it has branched and anastomosing pseudoparaphyses, lacking arcicular crystals within the spores, and ascospore size (Liu et al. 2015). Aliquandostipite manochiae might be synonymous with A. khaoyaiensis (Inderbitzin et al. 2001) based on their similar morphological features, overlapping dimension of ascomata, asci, and ascospores, and their freshwater habitats. Molecular data are necessary to confirm A. manochii as a unique species (Fig. 2).

Fig. 2
figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2figure 2

Phylogram generated from maximum likelihood analysis of combined LSU, ITS, TEF and RPB2 sequence data for species of Dothideomycetes. Bootstrap values for maximum likelihood equal to or greater than 75% and Bayesian posterior probabilities equal to or greater than 0.95 are placed near the branches as ML/BYPP. Newly generated sequences are in red and ex-type strains are in bold. The new species introduced in this study are indicated with underline. Freshwater strains are indicated with a red letter “F”. Orders not treated in this study are compressed. The tree is rooted to Capronia pilosella AFTOL-ID 657 and Endocarpon pallidulum AFTOL-ID 661 (Eurotiomycetes)

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List of freshwater Aliquandostipite species


*Aliquandostipite crystallinus Raja, A. Ferrer & Shearer, Mycotaxon 91: 208 (2005); Fig. 3e, g–i

Fig. 3
figure 3

Aliquandostipite spp. (Material examined: USA, Tennessee, Great Smoky Mountains National Park, Elkmont Campground, a small stream between Jakes Creek and Little River, on submerged, decorticated wood, 5 July 2002, H.A. Raja & N. Hamburger, R76-1; Florida, Big Cypress National Preserve, Cypress Swamp Loop Road, on submerged decorticated wood, 22 March 2005, H.A. Raja, J.L. Crane & A.N. Miller, F89-1). a–d, f A. khaoyaiensis (F89-1). a Stalked ascoma. b Ascospore with slug-like gelatinous sheath in nigrosin. c, d Asci. f Ascospore with thick-walled gelatinous sheath. e, g–i A. crystallinus (R76-1). e Ascoma. g, h Asci. i Ascospores emerging from the ascus to form a large, C-shaped to fusiform, gelatinous sheath. Scale bars: a = 100 µm, b–d = 20 µm, e = 200 µm, f–i = 20 µm

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Freshwater distribution: Costa Rica (Raja et al. 2005), Panama (Raja et al. 2005), Peru (Shearer et al. 2015), USA (Raja et al. 2005, 2009b), Venezuela (Raja et al. 2005)


*Aliquandostipite khaoyaiensis Inderbitzin, Am. J. Bot. 88: 54 (2001)

Facesoffungi number: FoF09158; Figs. 3a–d, f, 4

Fig. 4
figure 4

Aliquandostipite khaoyaiensis a–h, l, m (MFLU 18-1573, new geographical record), i–k (MFLU 18-1554, new geographical record). a, b, i Ascomata on host surface. c Vertical section of ascoma. d Structure of peridium. e Pseudoparaphyses. f, j Bitunicate asci. g, h, k Ascospores. l, m Colony on PDA (left-front, right-reverse). Scale bars: c = 100 μm, d = 30 μm, e, g, h = 20 μm, f, j, k = 50 μm

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Freshwater distribution: Costa Rica (Raja et al. 2005), USA (Raja et al. 2009b), Thailand (this study)

Saprobic on submerged wood. Sexual morph: Ascomata 220–240 μm high, 190–210 μm diam., hyaline to pale brown, become dark brown with age, scattered, superficial, sometimes seated in a pseudostroma, globose or subglobose, sessile, membranous, with ostiolate papilla. Peridium 25–40 μm thick, membranous, composed of thin-walled, pale brown, compressed cells of textura angularis. Pseudoparaphyses 2 μm diam., numerous, cellular, sparsely branched, hyaline, septate. Asci 115–175 × 35–65 μm (\( {\bar{\text{x}}} \) = 145 × 45 μm, n = 20), 8-spored, bitunicate, fissitunicate, clavate, thickened at the apex, sessile or short pedicellate, with a well-developed ocular chamber. Ascospores 50–70 × 14.5–19.5 μm (\( {\bar{\text{x}}} \) = 63 × 17 μm, n = 20), variably arranged in asci, oval, hyaline to pale brown, 1-septate, constricted at the septum, asymmetric, upper cell slightly shorter and wider than lower cell, guttulate, straight or curved, thin-walled, smooth, sheathed. Sheath first appressed to the ascospore wall, expanding and detaching from the polar regions when mounted in water, becoming balloon-like at the two poles, finally surrounding the entire ascospore. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 30 mm in 45 days at 25 °C, grey from above, dark brown from below, surface rough, with dense mycelium, mostly immersed in culture, dry, edge entire.

Material examined: THAILAND, Songkhla Province, on submerged wood in a stream, 10 May 2018, W. Dong, hat827-1 (MFLU 18-1573), living culture MFLUCC 18-1249; ibid., hat827-2 (HKAS 105024), living culture KUMCC 19-0039; Chestnut Hill, on submerged wood in a stream, 10 May 2018, W. Dong, hat460-1 (MFLU 18-1554); ibid., hat460-2 (HKAS 105066), living culture KUMCC 19-0083.

Notes: Our two new isolates MFLUCC 18-1249 and KUMCC 19-0083 are identified as Aliquandostipite khaoyaiensis based on their identical LSU, SSU, ITS sequence data and morphology, although they lack a stalked ascomata. The new isolates cluster with A. khaoyaiensis and A. siamensis with high bootstrap support (Fig. 14), which appears to be conspecific. However, our isolates must be A. khaoyaiensis because of its typical sheath. Aliquandostipite khaoyaiensis has monomorphic, pale brown ascospores with well-developed sheath (Inderbitzin et al. 2001). While A. siamensis has dimorphic, hyaline or brown ascospores without sheath (Pang et al. 2002). Aliquandostipite siamensis lacks ITS sequence data and needs fresh material to confirm its phylogenetic difference with A. khaoyaiensis. This is a new geographical record for A. khaoyaiensis from parts of Thailand.


Aliquandostipite manochii Sri-indr., Boonyuen, Suetrong, K.L. Pang & E.B.G. Jones, Fungal Diversity 72: 103 (2015)

Freshwater distribution: Thailand (Liu et al. 2015)


Aliquandostipite minuta Raja and Shearer, Mycoscience 43: 395 (2007); Fig. 5a–e

Fig. 5
figure 5

Aliquandostipitaceae spp. (Material examined: USA, Florida, Monroe County, freshwater swamp at Big Cypress National Preserve, on submerged decorticated wood, 17 February 2006, H.A. Raja & J.L. Crane, ILL 40108 = F117-1, holotype; PANAMA, Colon, Soberania National Park, Juan Grande, on submerged, decorticated wood, 12 January 2003, A. Ferrer, AF005). a–e Aliquandostipite minuta (F117-1). a Squash mount of ascoma. b, c Asci. d Extended endoascus releasing ascospore. e Ascospore showing fusiform sheath and filamentous appendages. f–j Megalohypha aqua-dulces (AF005). f Ascoma on substrate. g Ascus. h, i Ascospores. j Broad hyphae in PDA. Scale bars: a–e, g–i = 20 µm, f, j = 200 µm

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Freshwater distribution: USA (Raja and Shearer 2007; Raja et al. 2009b)


Aliquandostipite separans (Abdel-Wahab & El-Sharouney) J. Campb., Raja A. Ferrer, Sivichai & Shearer, Can. J. Bot. 85: 881 (2007)

Basionym: Patescospora separans Abdel-Wahab & El-Sharouney, Mycol. Res. 106: 1033 (2002)

Freshwater distribution: Egypt (Pang et al. 2002)


*Aliquandostipite siamensiae (Sivichai & E.B.G. Jones) J. Campb., Raja A. Ferrer, Sivichai & Shearer, Can. J. Bot. 85: 879 (2007)

Basionym: Jahnula siamensiae Sivichai & E.B.G. Jones, Mycol. Res. 106: 1037 (2002)

Freshwater distribution: Thailand (Pang et al. 2002), USA (Raja et al. 2009b)


Key to freshwater Aliquandostipite species

  1. 1.

    Ascospores with sheath………2

  2. 1.

    Ascospores with narrow sheath or without sheath………A. separans

  3. 2.

    Ascospores dimorphic………A. siamensiae

  4. 2.

    Ascospores monomorphic………3

  5. 3.

    Ascospores without appendages………A. khaoyaiensis

  6. 3.

    Ascospores with appendages………4

  7. 4.

    Ascospores with small apical appendages………A. crystallinus

  8. 4.

    Ascospores with numerous filamentous appendages………A. minuta


Jahnula Kirschst., Annls mycol. 34(3): 196 (1936)

Saprobic on submerged wood. Sexual morph: Ascomata semi-immersed to erumpent, become superficial with base remaining immersed, solitary or clustered in small groups, globose to subglobose, unilocular, brown to dark brown, membranous, with a stalk-like strand or stoloniferous hyphae attached to the substratum, or covered by sparse hair-like projections, with ostiolate papilla. Peridium variable in thickness, comprising a few layers of relatively large, thin-walled, light brown cells of textura angularis. Pseudoparaphyses cellular, persistent, filiform or hypha-like, hyaline, sparsely septate. Asci 8-spored, bitunicate, fissitunicate, mostly cylindrical, short-pedicellate, with an ocular chamber and sometimes with a faint ring. Ascospores mostly uniseriate, ellipsoid-fusiform, apical cell slightly larger, both cells tapering with rounded ends, reddish brown or dark brown, l-septate, straight or curved (definition sensu stricto from Hawksworth (1984), Hyde and Goh (1999b) and Raja and Shearer (2006)). Asexual morph: Undetermined.

Type species: Jahnula aquatica (Kirschst.) Kirschst.

Notes: Jahnula is undoubtedly polyphyletic (Hyde et al. 2013, 2017, 2019; Huang et al. 2018) and Jahnula sensu stricto was mentioned in Huang et al. (2018), and Hyde et al. (2013). Hyde et al. (2013) suggested that taxonomic changes are needed for taxa in Jahnula sensu lato, but more jahnula-like species need to be collected and sequenced. We accept three species, J. aquatica, J. granulosa K.D. Hyde & S.W. Wong and J. rostrata Raja & Shearer in Jahnula sensu stricto, which have been collected from freshwater habitats (listed below). These three species share common characters in having dark ascomata with few appendages attached to the base, and ellipsoid-fusiform, brown, l-septate ascospores with wider apical cell (Hawksworth 1984; Hyde and Wong 1999; Raja and Shearer 2006). Asci of J. aquatica are cylindrical, those of J. granulosa are obclavate and those of J. rostrata are clavate. Jahnula aquatica has smooth-walled ascospores without a sheath, while J. granulosa has granular-walled ascospores with a thin mucilaginous sheath. Jahnula rostrata is similar to J. granulosa by rough-walled ascospores, but differs in having an irregularly striated pattern. These three species formed a well-supported and stable clade in previous publications (Huang et al. 2018; Hyde et al. 2019) and in this study (Fig. 14).

The type species, Jahnula aquatica commonly occurs in freshwater habitats as listed below. The lectotype of J. aquatica was examined, illustrated and described by Hawksworth (1984). Jahnula aquatica is characterized by ascomata attached to the substratum by subiculum-like hyphae, peridium uneven in thickness, cylindrical asci, elongate-ellipsoid to very broadly fusiform, reddish brown, smooth, moderately thick-walled ascospores without a distinct gelatinous sheath (Hawksworth 1984). The sequences of two isolates R68-1 and R68-2 (not type) were obtained by Raja and Shearer (2006), recognized and used for current phylogenetic analyses (Suetrong et al. 2011a; Huang et al. 2018; Hyde et al. 2019).

Xylomyces chlamydosporus Goos et al. was considered to be the asexual state of Jahnula aquatica based on fusiform chlamydospores produced in culture (Sivichai et al. 2011). However, the phylogenetic analyses did not support this conclusion because J. aquatica and X. chlamydosporus scattered in different clades within Jahnulales (Campbell et al. 2007; Suetrong et al. 2011a; this study, Fig. 14).

We introduce two new genera Neojahnula and Pseudojahnula to accommodate two known species Jahnula australiensis K.D. Hyde and J. potamophila K.D. Hyde & S.W. Wong, respectively. Seven species, i.e. J. bipolaris, J. guttulaspora, J. queenslandica, J. seychellensis, J. submersa, J. sunyatsenii and J. thailandensis, which are phylogenetically distant from J. aquatica, are transferred to Ascagilis K.D. Hyde. Nine species, i.e. J. apiospora, J. appendiculata, J. bipileata, J. dianchia, J. morakotii, J. poonythii, J. purpurea, J. sangamonensis and J. systyla, are placed in Jahnula sensu lato.

For convenience and practicality, we have excluded Jahnula systyla K.D. Hyde & S.W. Wong from the key, because we believe it may be phylogenetically related to Aliquandostipite, pending future phylogenetic studies (Fig. 6).

Fig. 6
figure 6

Aliquandostipitaceae spp. (Material examined: USA, Florida, on submerged wood, July 2006, J.L. Crane & H.A. Raja, F111; Costa Rica, La Selva Biological Station, Sura 60, on submerged, corticated wood, CMP, A492; Florida, Apalachicola National Forest, swampy area of Whitehead Lake, on submerged, decorticated wood, 13 July 2004, HAR and CB, F49-1 (ILL), holotype; CANADA, Prince Edward Island, on submerged decorticated wood, in a small creek beside dirt road near Bonshaw and Trans Canada Highway, 13 October 2007, A. Carter, AC-706, ILL40554, holotype). a, e, h Pseudojahnula potamophila (F111). a Ascomata attached to the wood with broad hyphae. e, h Ascospores. b Section of ascoma of Ascagilis seychellensis (A492). c, j Jahnula bipileata (F49-1). c Ascoma. j Ascospores. d, i Jahnula apiospora (AC-706). d Structure of peridium. i One-septate apiosporous ascospore. f, g Ascospores of Ascagilis bipolaris showing bipolar appendages. Scale bars: a, c = 200 µm, b, d–j = 20 µm

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List of freshwater Jahnula sensu stricto species

*Jahnula aquatica (Plöttner & Kirschst.) Kirschst., Ann., Mycol. 34: 196 (1936); Fig. 7

Fig. 7
figure 7

Jahnula aquatica (Material examined: USA, Illinois, Salt Fork Association Lake (SFA Lake), a manmade private lake, on submerged, decorticated wood, 15 October 2002, HAR, R68-1). a Squash mount of ascoma. b Ascus. c Pseudoparaphyses. d, e Ascospores. Scale bars: a = 100 µm, b, c, e = 20 µm, d = 10 µm

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Basionym: Melanopsamma aquatica Kirschst., Krypt.-Fl. Brandenburg (Leipzig) 7(2): 226 (1911)

Freshwater distribution: China (Ho 1998), France (Fournier et al. 2010), Germany (Kirschstein 1936; Hawksworth 1984), Malaysia (Ho et al. 2001), South Africa (Hyde and Wong 1999), Thailand (Sivichai et al. 2011), USA (Raja and Shearer 2006; Raja et al. 2009b)


*Jahnula granulosa K.D. Hyde & S.W. Wong, Nova Hedwig. 68: 497 (1999)

Freshwater distribution: China (Cai et al. 2002a), South Africa (Hyde and Wong 1999), Thailand (Suetrong et al. 2011a)


*Jahnula rostrata Raja & Shearer, Mycologia 98: 325 (2006)

Facesoffungi number: FoF09237; Fig. 8

Fig. 8
figure 8

Jahnula rostrata (MFLU 20-0435, new geographical record). a Appearance of black ascoma on host substrate. b Structure of ostiole. c Pseudoparaphyses. d Vertical section of ascoma. e Structure of peridium. f–i Ascospores. j–m Bitunicate asci. n Germinated ascospore. o, p Colony on PDA (left-front, right-reverse). Scale bars: b, e–n = 30 μm, c = 10 μm, d = 50 μm

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Freshwater distribution: China (this study), USA (Raja and Shearer 2006; Raja et al. 2009b)

Saprobic on submerged wood. Sexual morph: Ascomata 320–420 × 310–330 μm, superficial, solitary to scattered, subglobose or pyriform, brown to black, rough-walled, with ostiolate papilla. Neck 48–75 × 74–98 μm, composed of subglobose cells diverging from the ostiole. Peridium 25–40 μm wide, two-layered, outer layer composed of large, dark brown, thick-walled cells of textura angularis, inner layer composed of compressed, hyaline, thin-walled cells of textura angularis. Pseudoparaphyses 2.5–3.5 μm wide, cellular, hypha-like, hyaline, septate, unbranched. Asci 135–200 × 21–32 μm (\( {\bar{\text{x}}} \) = 160 × 28 μm, n = 15), 8-spored, bitunicate, fissitunicate, clavate, short pedicellate, with an ocular chamber. Ascospores 28–40 × 11–17 μm (\( {\bar{\text{x}}} \) = 35 × 13.5 μm, n = 15), overlapping biseriate, ellipsoidal or broadly fusiform, with rounded or occasionally acute ends, upper cell slightly broader than lower cell, hyaline to brown when young, dark brown at maturity, 1-septate, constricted at the septum, with thick and dark band at septum when mature, rough-walled, with minute striated pattern, guttulate, with or without sheath. Asexual morph: Undetermined.

Culture characteristics: On PDA, colonies irregular, reaching 35 mm in 30 days at 25 °C, dark from above and below, dry, hairy, dense in the centre, becoming sparse at the edge.

Material examined: CHINA, Yunnan Province, Kunming, Liangwang National Wetland Park, on submerged wood in a stream, 19 March 2019, C.X. Li, L15 (MFLU 20-0435).

Notes: Our collection MFLU 20-0435 is identified as Jahnula rostrata based on very similar morphology and only one and one nucleotide difference in LSU and SSU sequence data between MFLU 20-0435 and F4-3, respectively. They form a well-supported clade in our phylogenetic analysis (Fig. 14). MFLU 20-0435 has thinner asci than the holotype (135–200 × 21–32 μm vs. 152–190 × 32–40 μm) (Raja and Shearer 2006). Our collection is a new geographical record for J. rostrata from China. Jahnula rostrata (F4-3) lacks ITS sequence data, we supplement ITS sequence for this species in this study.


Key to freshwater Jahnula sensu stricto species

  1. 1.

    Ascospores smooth-walled………J. aquatica

  2. 1.

    Ascospores rough-walled………2

  3. 2.

    Ascospores granular-walled………J. granulosa

  4. 2.

    Ascospores with an irregularly striated pattern………J. rostrata


List of freshwater Jahnula sensu lato species

Jahnula apiospora A. Carter, Raja & Shearer, Mycoscience 49: 326 (2008); Fig. 6d, i

Freshwater distribution: Canada (Raja et al. 2008)


*Jahnula appendiculata Pinruan, K.D. Hyde & E.B.G. Jones, Sydowia 54: 243 (2002); Fig. 9

Fig. 9
figure 9

Jahnula appendiculata (Material examined: PERU, on submerged wood, S. Zelski & H.A. Raja, PE0010). a Stalked ascoma. b Broad hypha attached to the ascomatal sides and bases. c Young ascus. d, e Ascospores with broad gelatinous sheath and polar appendages indicated by arrows. Scale bars: a = 50 µm, b–e = 20 µm

Full size image

Freshwater distribution: Peru (Shearer et al. 2015), Thailand (Pinruan et al. 2002; Sivichai and Boonyuen 2010)


*Jahnula bipileata Raja & Shearer, Mycologia 98: 321 (2006); Fig. 6c, j

Freshwater distribution: USA (Raja and Shearer 2006)


Jahnula morakotii Sivichai & Boonyuen, Mycotaxon 112: 476 (2012)

Freshwater distribution: Thailand (Sivichai and Boonyuen 2010)


Jahnula poonythii K.D. Hyde & S.W. Wong, Nova Hedwig. 68: 499 (1999)

Freshwater distribution: China (Cai et al. 2002a; Luo et al. 2004), Mauritius (Hyde and Wong 1999), Mexico (Gonzalez and Chavarria 2005)


Jahnula purpurea J. Fourn., Raja & Shearer, Mycokeys 9: 30. (2015)

Freshwater distribution: Martinique (Fournier et al. 2015)


*Jahnula sangamonensis Shearer & Raja, Mycologia 98: 327 (2006)

Freshwater distribution: USA (Raja and Shearer 2006; Raja et al. 2009b)


Jahnula systyla K.D. Hyde & S.W. Wong, Nova Hedwig. 68: 506 (1999)

Freshwater distribution: Australia (Hyde and Wong 1999), China (Ho 1998), Malaysia (Ho et al. 2001)


Key to freshwater Jahnula sensu lato species

  1. 1.

    Ascospores without modification (no appendages/sheath/caps)………2

  2. 1.

    Ascospores equipped with modification (appendages/sheath/caps)………6

  3. 2.

    Ascospores asymmetric………J. apiospora

  4. 2.

    Ascospores symmetric………3

  5. 3.

    Ascomata stain substrate purple………J. purpurea

  6. 3.

    Ascomata not as above………4

  7. 4.

    Ascomata with a short but prominent neck, endoasci extend up to 500 µm long in water………J. sangamonensis

  8. 4.

    Ascomata with a short papilla, endoasci not extending………5

  9. 5.

    Ascomata with stalk-like strands attached to the base, with a few sparse hair-like projections………J. poonythii

  10. 5.

    Ascomata without any appendages………J. dianchia

  11. 6.

    Ascospores without unfurling bipolar appendages………J. bipileata

  12. 6.

    Ascospores with long unfurling bipolar appendages………7

  13. 7.

    Ascospores 18–20 × 5–6 μm………J. morakotii

  14. 7.

    Ascospores 48–55 × 23–26 μm………J. appendiculata


Ascagilis K.D. Hyde, Aust. Syst. Bot. 5(1): 109 (1992)

Saprobic on submerged wood. Sexual morph: Ascomata semi-immersed or superficial, solitary, scattered or gregarious, globose or subglobose, coriaceous, black, ostiolate. Peridium comprising several layers of thin-walled, brown, angular cells. Pseudoparaphyses numerous, cellular, hyaline, thin, septate, branched. Asci 8-spored, clavate, obclavate, broadly or narrowly cylindrical, fissitunicate, sometimes expanding 4–5 times longer than original length, with an ocular chamber and ring. Ascospores bi- to tri-seriate, 1-septate, brown, mostly with a refractive mucilaginous pad at each end. Asexual morph: Undetermined.

Type species: Ascagilis bipolaris K.D. Hyde

Notes: Ascagilis was introduced to accommodate a jahnula-like species, A. bipolaris, characterized by clavate to cylindrical asci with an ocular chamber and ring and fusiform to ellipsoidal, 1-septate, brown ascospores with a pad at each end (Hyde 1992b). Ascagilis was suppressed since the type species A. bipolaris was transferred to Jahnula (Hyde and Wong 1999) and they thought the fusiform to ellipsoidal ascospores with bipolar pads of Ascagilis was insufficient to separate these two genera. Jahnula has been shown to be polyphyletic in previous publications (Hyde et al. 2013, 2017, 2019; Huang et al. 2018) and in this study. The clade comprising A. bipolaris and six other species is phylogenetically distant from Jahnula sensu stricto (Fig. 14). Additionally, most species including our new species, A. submersa and A. thailandensis, have a refractive mucilaginous pad at each end, which is lacking in Jahnula sensu stricto. All species in this clade cluster with strong bootstrap support (Fig. 14). We therefore, resurrect Ascagilis and transfer four species and introduce two new species in the genus (see list below). The mucilaginous pad of ascospores is treated as a noticeable characteristic of Ascagilis.


List of freshwater Ascagilis species

*Ascagilis bipolaris K.D. Hyde, Aust. Syst. Bot. 5(1): 111 (1992); Fig. 6f, g

Synonymy: Jahnula bipolaris (K.D. Hyde) K.D. Hyde, Nova Hedwigia 68(3–4): 494 (1999)

Freshwater distribution: Australia (Hyde 1992b; Hyde and Wong 1999), China (Tsui et al. 2000; Tsui and Hyde 2004), Costa Rica (Raja and Shearer 2006), Malaysia (Ho et al. 2001), Peru (Shearer et al. 2015), Thailand (Suetrong et al. 2011a), USA (Raja et al. 2009b)


*Ascagilis guttulaspora (Qing Tian, Y.Z. Lu & K.D. Hyde) W. Dong, Doilom & K.D. Hyde, comb. nov.

Index Fungorum number: IF557893; Facesoffungi number: FoF09228

Basionym: Jahnula guttulaspora Qing Tian, Y.Z. Lu & K.D. Hyde, Fungal Diversity 87: 6 (2017)

Freshwater distribution: China (Hyde et al. 2017)


*Ascagilis queenslandica (Dayarathne, Fryar & K.D. Hyde) W. Dong, Doilom & K.D. Hyde, comb. nov.

Index Fungorum number: IF557894; Facesoffungi number: FoF09229

Basionym: Jahnula queenslandica Dayarathne, Fryar & K.D. Hyde, Fungal Diversity 96: 91 (2019)

Freshwater distribution: Australia (Hyde et al. 2019)


*Ascagilis seychellensis (K.D. Hyde & S.W. Wong) W. Dong, Doilom & K.D. Hyde, comb. nov.

Index Fungorum number: IF557895; Facesoffungi number: FoF09230

Basionym: Jahnula seychellensis K.D. Hyde & S.W. Wong, Nova Hedwig. 68: 504 (1999); Fig. 6b

Freshwater distribution: Brazil (Barbosa et al. 2013), Costa Rica (Raja and Shearer 2006), Seychelles (Hyde and Wong 1999)


*Ascagilis submersa W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557896; Facesoffungi number: FoF09238; Fig. 10

Fig. 10
figure 10

Ascagilis submersa (MFLU 18-1527, holotype). a Ascomata on host with associated appendages (arrow). b Vertical section of ascoma. c Structure of peridium. d Pseudoparaphyses. e–h Bitunicate asci. i Ascospores. j Ascospore in Indian Ink. k Germinated ascospore. l, m Colony on PDA (left-front, right-reverse). Scale bars: b = 50 μm, c–k = 20 μm

Full size image

Etymology: in reference to the submerged habitats of the fungus

Holotype: MFLU 18-1527

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 160–250 μm diam., black, solitary or scattered, superficial, globose or subglobose, with several long, brown to dark brown appendages which are procumbent on the substrate surface, membranous. Peridium 30–35 μm thick, composed of 4–5 layers of thick-walled, olive to dark brown, compressed cells of textura angularis, with 2 layers of flattened, compressed, hyaline cells inwardly. Pseudoparaphyses 3 μm diam., numerous, cellular, hypha-like, hyaline, septate, embedded in a gelatinous matrix. Asci 110–160 × 35–42 μm (\( {\bar{\text{x}}} \) = 135 × 38 μm, n = 10), 8-spored, bitunicate, thick-walled, broadly obclavate, with thick-walled apex and well-developed ocular chamber, completely filled by ascospores, short pedicel observed only when young. Ascospores 37.5–41 × 17–19 μm (\( {\bar{\text{x}}} \) = 40 × 18 μm, n = 20), overlapping biseriate and uniseriate near the apex, straight or slightly curved, hyaline when young, pale brown to brown when mature, 1-septate, constricted at the septum, ellipsoidal, guttulate, thin-walled, smooth, with bipolar, helmet-shaped appendages. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 10 mm in 10 days at 25 °C, dark brown from above and below, surface rough, with sparse mycelium, dry, edge entire.

Material examined: THAILAND, Songkhla Province, on submerged wood in a stream, 10 May 2018, W. Dong, 20180513-1 (MFLU 18-1527, holotype), ex-type living culture MFLUCC 18-1143; ibid., 20180513-2 (HKAS 104996, isotype), ex-type living culture KUMCC 19-0013.

Notes: Ascagilis submersa clusters with A. queenslandica, A. sunyatsenii and A. thailandensis (Fig. 14). Ascagilis submersa is similar to A. sunyatsenii in having ellipsoidal, pale brown ascospores with bipolar helmet-shaped appendages (Inderbitzin et al. 2001). However, A. sunyatsenii has saccate, ovoid to elongate asci and longer ascospores ((39–)46–52 × 16–23 μm) contrasting with broadly obclavate asci and shorter ascospores (37.5–41 × 17–19 μm) in A. submersa (Inderbitzin et al. 2001). Ascagilis submersa has olive peridium and ascospores, which are unique in the genus and different from the dark pigmented peridium and light brown ascospores of A. sunyatsenii. Additionally, A. sunyatsenii has two types of ascomata, sessile and stalked, while A. submersa has only sessile ascomata and with several appendages which are procumbent on the substrate surface (Inderbitzin et al. 2001). Because of scarcity of some nucleic acid genes, only ITS sequence can be compared. There are two nucleotide differences in ITS sequence data, but only crossing 207 nucleotides. The distinct morphology of A. submersa provides strong evidence to be a new species in Ascagilis.


*Ascagilis sunyatsenii (Inderb.) W. Dong, Doilom & K.D. Hyde, comb. nov.

Index Fungorum number: IF557897; Facesoffungi number: FoF09239

Basionym: Aliquandostipite sunyatsenii Inderb., Am. J. Bot. 88(1): 57 (2001)

Synonymy: Jahnula sunyatsenii (Inderb.) K.L. Pang, E.B.G. Jones & Sivichai, Mycol. Res 106: 1037 (2002)

Freshwater distribution: China (Inderbitzin et al. 2001; Pang et al. 2002)


*Ascagilis thailandensis W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557898; Facesoffungi number: FoF09240; Fig. 11

Fig. 11
figure 11

Ascagilis thailandensis (MFLU 18-1514, holotype). a, b Ascomata on host tissue. c Vertical section of ascoma. d Structure of peridium. e–h Bitunicate asci. i Pseudoparaphyses. j–m Ascospores. n Germinated ascospore. o, p Colony on PDA (left-front, right-reverse). Scale bars: c, n = 50 μm, d–m = 20 μm

Full size image

Etymology: in reference to Thailand, where the holotype was collected

Holotype: MFLU 18-1514

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 280–320 μm high, 200–250 μm diam., black, scattered, superficial with base immersed in host tissue, conical, membranous, coriaceous, with short ostiolate papilla. Peridium 30–40 μm thick, composed of 4–5 layers of thin-walled, pale brown, large cells of textura angularis. Pseudoparaphyses 3.5 μm diam., numerous, cellular, hypha-like, hyaline, septate. Asci 125–185 × 24–30 μm (\( {\bar{\text{x}}} \) = 162 × 27.5 μm, n = 5), 8-spored, bitunicate, broadly or narrowly cylindrical, with an ocular chamber, short pedicellate. Ascospores 33–48.5 × 11–16 μm (\( {\bar{\text{x}}} \) = 41 × 13 μm, n = 30), biseriate or uniseriate above ascus center and overlapping biseriate near the center or base, brown, 1-septate, slightly constricted at the septum, irregularly fusiform, mostly curved, tapering at apical cell and slightly flattened or rounded at the basal cell, guttulate, thin-walled, smooth, with small, inconspicuous pads at one or both apices. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 5 mm in 5 days at 25 °C, olive brown from above, brown from below, surface rough, with sparse mycelium, dry, edge entire.

Material examined: THAILAND, Songkhla Province, on submerged wood in a stream, 10 May 2018, W. Dong, 20180524 (MFLU 18-1514, holotype), ex-type living culture MFLUCC 18-1149; ibid., Chestnut Hill, on submerged wood in a stream, 10 May 2018, W. Dong, hat454-1 (MFLU 18-1546), living culture MFLUCC 18-1247; ibid., hat454-2 (HKAS 105022), living culture KUMCC 19-0037.

Notes: Ascagilis thailandensis clusters with A. queenslandica, A. submersa and A. sunyatsenii with low bootstrap support (Fig. 14). The phylogenetically closest species A. sunyatsenii only has ITS sequence data (323 nucleotides) in GenBank, which has three nucleotide differences with A. thailandensis. Due to the limited sequence data, A. sunyatsenii and A. thailandensis appear to represent the same species in our phylogenetic tree (Fig. 14). However, their morphological characteristics strongly support them to be the different species. Ascagilis sunyatsenii has two types of ascomata (sessile and stalked), saccate, ovoid to elongate asci and ellipsoidal ascospores with two conspicuous helmet-shaped appendages (Inderbitzin et al. 2001). Whereas, A. thailandensis has sessile ascomata, cylindrical asci and irregularly fusiform ascospores with small, inconspicuous pads. Ascagilis thailandensis can be easily distinguished from the other two phylogenetically related species A. queenslandica and A. submersa by ascospore shape, size and appendages (Hyde et al. 2019).

Ascagilis thailandensis is morphologically similar to A. guttulaspora in having nearly fusiform, smooth-walled, guttulate ascospores with tapering apical cell and slightly flattened basal cell. However, A. guttulaspora has smaller ascospores (29–33 × 8.5–10.5 μm vs. 33–48.5 × 11–16 μm) and without pads (Hyde et al. 2017). The ascomata of A. guttulaspora are attached to the substratum by wide, brown hyphae, while they are lacking in A. thailandensis (Hyde et al. 2017). Our phylogenetic analysis supports them to be the different species (Fig. 14).


Key to freshwater Ascagilis species

  1. 1.

    Ascospores without mucilaginous pads………2

  2. 1.

    Ascospores with mucilaginous pads at one or both ends………3

  3. 2.

    Ascospores aseptate………A. queenslandica

  4. 2.

    Ascospores 1-septate………A. guttulaspora

  5. 3.

    Ascomata sessile………4

  6. 3.

    Ascomata sessile and stalked………6

  7. 4.

    Ascospores with inconspicuous pads at one or both ends………A. thailandensis

  8. 4.

    Ascospores with conspicuous bipolar pads………5

  9. 5.

    Ascospores 37.5–41 × 17–19 μm………A. submersa

  10. 5.

    Ascospores 42–52 × 16–23 μm………A. bipolaris

  11. 6.

    Ascospores monomorphic………A. sunyatsenii

  12. 6.

    Ascospores dimorphic………A. seychellensis


Neojahnula W. Dong, H. Zhang & K.D. Hyde, gen. nov.

Index Fungorum number: IF557823; Facesoffungi number: FoF07688

Etymology: named refers to its morphological similarity to Jahnula

Saprobic on submerged wood. Sexual morph: Ascomata scattered or clustered in small groups, superficial, subglobose or obpyriform, brown to black, membranous, with scarcely projecting papilla, ostiolate, with few setae. Peridium thin, one-layered. Pseudoparaphyses numerous, cellular, hypha-like, hyaline, sparsely septate. Asci 8-spored, bitunicate, fissitunicate, cylindrical, sessile or short pedicellate, with an ocular chamber and faint apical ring. Ascospores uni- to bi-seriate, elongate, ellipsoidal, tapering towards the apices, brown, 1-septate, upper cell slightly wider than lower cell. Asexual morph: Undetermined.

Type species: Neojahnula australiensis (K.D. Hyde) W. Dong, H. Zhang & K.D. Hyde

Notes: Jahnula australiensis K.D. Hyde was collected from submerged wood in freshwater in Australia (Hyde 1993a). It was initially placed in Jahnula because the peridium comprises massive pseudoparenchymatous cells, which is the remarkable feature of Jahnula (Hyde 1993a). However, J. australiensis was shown to not belong to Jahnula sensu stricto and clustered basal to two asexual genera Brachiosphaera Nawawi and Speiropsis Tubaki with strong bootstrap support (Prihatini et al. 2008; Hyde et al. 2017; Huang et al. 2018; this study, Fig. 14). Jahnula australiensis is not supported in Jahnula because it has few setae surrounding the ascomata, a thin peridium comprising single row of cells, asci with an ocular chamber and faint apical ring and uni- to bi-seriate ascospores, these features are different from Jahnula. As suggested by Hyde et al. (2013), we transfer J. australiensis to a new genus, Neojahnula based on phylogenetic analyses and morphology.


List of freshwater Neojahnula species


*Neojahnula australiensis (K.D. Hyde) W. Dong, H. Zhang & K.D. Hyde, comb. nov.

Index Fungorum number: IF557899; Facesoffungi number: FoF09241; Fig. 12

Fig. 12
figure 12

Neojahnula australiensis (MFLU 15-2711). a Ascomata on host surface. b Vertical section of ascoma. c Pseudoparaphyses. d Structure of peridium. e, f Bitunicate asci. g Ocular chamber and faint apical ring. h Ascospore. i, j Colony on PDA (up-front, down-reverse). Scale bars: b, e, f = 20 μm, c = 15 μm, d = 5 μm, g, h = 10 μm

Full size image

Basionym: Jahnula australiensis K.D. Hyde, Aust. Syst. Bot. 6(2): 161 (1993)

Freshwater distribution: Australia (Hyde 1993a), Brunei (Ho et al. 2001), China (Ho et al. 2001), Peru (Shearer et al. 2015), Thailand (Suetrong et al. 2011a; this study) USA (Raja et al. 2009b)

Saprobic on submerged wood. Sexual morph: Ascomata 80–110 μm high, 100–130 μm diam., scattered or gregarious, superficial, subglobose or lenticulate when compressed, black, membranous, with scarcely projecting papilla, ostiolate, with few hyaline to pale brown hyphae. Peridium thin, 5–10 μm thick, composed of one row of large, thin-walled, brown to dark brown cells of textura angularis. Pseudoparaphyses 3 μm diam., numerous, cellular, hypha-like, hyaline, sparsely septate. Asci 135–175 × 17–19 μm (\( {\bar{\text{x}}} \) = 155 × 17.5 μm, n = 5), 8-spored, bitunicate, fissitunicate, cylindrical, sessile or short pedicellate, with an ocular chamber and faint apical ring. Ascospores 23–27.5 × 6–7.5 μm (\( {\bar{\text{x}}} \) = 25.5 × 7 μm, n = 15), uni- to bi-seriate, elongate, ellipsoidal, tapering towards the apices, rounded at the base, brown, 1-septate, constricted at the septum, upper cell slightly wider than lower cell, guttulate, mostly curved, smooth, thin-walled, without sheath. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 20 mm in 25 days at 25 °C, reddish brown from above, dark brown to black from below, surface rough, with dense mycelium, fluffy, raised as a annulus around the margin, dry, edge entire.

Material examined: THAILAND, Prachuap Khiri Khan Province, on submerged wood in a stream, 30 July 2015, K.D. Hyde, 66D (MFLU 15-2711), living culture MFLUCC 15-0975.

Notes: Our collection MFLUCC 15-0975 clusters with two strains of Neojahnula australiensis (SS3613 and SS0665) with high bootstrap support (Fig. 14). SS3613 only has ITS and SS0665 only has SSU sequence data in GenBank, which are identical with MFLUCC 15-0975. The morphology of SS3613 and SS0665 cannot be compared as they were not provided in Prihatini et al. (2008) and Pang et al. (2002), respectively. The sequence data from ex-type strain of N. australiensis is lacking, but our collection MFLUCC 15-0975 has identical morphological characteristics with the holotype (BRIP 19208), except for longer asci (135–175 × 17–19 μm vs. 90–140 × 14–18 μm) (Hyde 1993a). Based on morphological characteristics and available sequence data, MFLUCC 15-0975 is identified as N. australiensis.


Pseudojahnula W. Dong, H. Zhang & K.D. Hyde, gen. nov.

Index Fungorum number: IF557824; Facesoffungi number: FoF07689

Etymology: named refers to its morphological similarity to Jahnula

Saprobic on submerged wood. Sexual morph: Ascomata solitary or gregarious, erumpent, become superficial with base remaining immersed, subglobose, obpyriform or almost conical, hyaline and then metallic grey, coriaceous, with ostiolate papilla, with algal associations. Peridium comprising several layers of relatively large, hyaline, thin-walled, angular cells and covered with sparse hyaline hairs. Pseudoparaphyses numerous, trabeculate, filamentous, hyaline, septate, unbranched between the asci, branching and anastomosing above. Asci 8-spored, bitunicate, fissitunicate, obclavate, pedicellate, with an ocular chamber and faint ring. Ascospores bi- to tri-seriate near the base, overlapping uniseriate near the apex, l-septate, light brown, ellipsoid-fusiform, surrounded by a mucilaginous sheath, which is wavy in outline. Asexual morph: Undetermined.

Type species: Pseudojahnula potamophila (K.D. Hyde & S.W. Wong) W. Dong, H. Zhang & K.D. Hyde

Notes: Jahnula potamophila is morphologically closest to Ascagilis bipolaris in ascospore size, but differs by narrow mucilaginous sheath which is wavy in outline (Hyde and Goh 1999b; Hyde and Wong 1999). Both species cluster distantly in Jahnulales (Fig. 14). The phylogenetic placement of J. potamophila is unstable, it clustered with Jahnula sensu stricto clade with low bootstrap support (Huang et al. 2018), but had good affinity in Hyde et al. (2017). With Megalohypha aqua-dulces and our new sequences added in the order, J. potamophila clustered with M. aqua-dulces with moderate bootstrap support (Hyde et al. 2019; this study, Fig. 14).

Jahnula potamophila has hyaline to metallic grey ascomata, a several-layered peridium comprising hyaline cells and covered with sparse hyaline hairs, obclavate asci with an ocular chamber and faint ring, and ascospores surrounded by a wavy mucilaginous sheath (Hyde and Wong 1999). These characteristics differ from Jahnula, therefore, Pseudojahnula is introduced to accommodate J. potamophila.


List of freshwater Pseudojahnula species

*Pseudojahnula potamophila (K.D. Hyde & S.W. Wong) W. Dong, H. Zhang & K.D. Hyde, comb. nov.

Index Fungorum number: IF557900; Facesoffungi number: FoF09242; Fig. 6a, e, h

Basionym: Jahnula potamophila K.D. Hyde & S.W. Wong, Nova Hedwigia 68(3–4): 499 (1999)

Freshwater distribution: Australia (Hyde and Wong 1999), Costa Rica (Raja and Shearer 2006), USA (Raja et al. 2009b)


Megalohypha A. Ferrer & Shearer, Ferrer, Sivichai & Shearer, Mycologia 99: 456 (2007)

Saprobic on submerged wood. Sexual morph: Ascomata superficial, uniloculate, globose to obpyriform, hyaline, membranous, with ostiolate papilla, stalked or sessile. Peridium composed of large, hyaline, thin-walled cells. Pseudoparaphyses numerous, trabeculate, filamentous, hyaline, septate, branched, anastomosing above the asci. Asci 8-spored, bitunicate, fissitunicate, broadly clavate or fusiform, short pedicellate, with or without an ocular chamber. Ascospores irregularly arranged, ellipsoidal, acutely tapered at apices, straight, brown to dark brown, 1-septate, symmetrical, rough-walled, longitudinally striate, without appendages or gelatinous sheath (Ferrer et al. 2007). Asexual morph: Undetermined.

Type species: Megalohypha aqua-dulces A. Ferrer & Shearer

Notes: Megalohypha is monotypic with M. aqua-dulces occurring on submerged decorticated wood in Panama and Thailand (Ferrer et al. 2007). Megalohypha can be easily distinguished from Aliquandostipite and Jahnula based on its pale to dark brown, 1-septate, rough-walled ascospores with longitudinally sulcate striations and without appendages or gelatinous sheath (Ferrer et al. 2007). It formed a moderately supported clade with Pseudojahnula (Hyde et al. 2019) and this study (Fig. 14).


List of freshwater Megalohypha species

*Megalohypha aqua-dulces A. Ferrer & Shearer, Mycologia 99(3): 458 (2007); Fig. 5f–j

Freshwater distribution: Panama (Ferrer et al. 2007), Thailand (Ferrer et al. 2007)


Key to freshwater asexual genera of Aliquandostipitaceae

  1. 1.

    Chlamydospores long, multi-septate………Xylomyces

  2. 1.

    Conidia tetraradiate with 4–8 appendages………Brachiosphaera


Brachiosphaera Nawawi, Descals, Nawawi & Webster, Trans. Br. Mycol. Soc. 67: 213 (1976)

Saprobic on submerged wood, wet wood and river foam. Sexual morph: Undetermined. Asexual morph: Colonies effuse, mycelium mostly immersed. Hyphae varying in width, septate, hyaline at first, turning olivaceous brown with age. Conidiophores macronematous, erect, unbranched, hyaline, septate, of variable length. Conidiogenous cells holoblastic, sympodial. Conidia acrogenous, tetraradiate, clustered in chains, ellipsoid or round-shaped, with 4–10 conidial arms, each 1–4-septate (Nawawi and Webster 1976; Suetrong et al. 2011a).

Type species: Brachiosphaera tropicalis Nawawi

Notes: Brachiosphaera species are commonly recorded in freshwater habitats as listed below. The genus is characterized by tetraradiate conidia with a round central cell with 4–10 arms, each of them 1–4-septate (Nawawi and Webster 1976; Suetrong et al. 2011a). The conidia of B. tropicalis consist of a brown, spherical body and furnished with 4–5 arms, which are longer than 1.5 times diam. of the central part (Nawawi and Webster 1976; Suetrong et al. 2011a). Currently, B. tropicalis is phylogenetically related to Jahnulales (Fig. 14). Sequence data for B. jamaicensis is necessary to shed light on its phylogenetic affinities.


List of freshwater Brachiosphaera species

Brachiosphaera jamaicensis (J.L. Crane & Dumont) Nawawi, Trans. Br. mycol. Soc. 67(2): 216 (1976)

Basionym: Actinospora jamaicensis J.L. Crane & Dumont, Can. J. Bot. 53(9): 843 (1975)

Freshwater distribution: Jamaica (Crane and Dumont 1975), China (Chang 1994), Poland (Suetrong et al. 2011a)


*Brachiosphaera tropicalis Nawawi, Trans. Br. mycol. Soc. 67(2): 213 (1976); Fig. 13c–f

Fig. 13
figure 13

Aliquandostipitaceae spp. (Material examined: USA, Tennessee, Great Smoky Mountains National Park, on submerged decorticated wood, 21 June 2005, H.A. Raja, A.N. Miller & E.B. Lickey, H58-4; PANAMA, Colon Province, Barro Colorado Island, small river, January 2003, on submerged decorticated wood, A. Ferrer, E192-1). a, b Conidia of Xylomyces chlamydosporus (H58-4). c–f Brachiosphaera tropicalis (E192-1). c–e Conidia in PDA. f Broad hyphae in PDA. Scale bars: a–d = 200 µm, e = 50 µm, f = 100 µm

Full size image

Freshwater distribution: China (Chang 1994; Cai et al. 2002a), Malaysia (Nawawi and Webster 1976), Panama (Campbell et al. 2007), Peru (Shearer et al. 2015), Puerto Rico (Nieves-Rivera and Santos-Flores 2005), Thailand (Tubaki et al. 1983; Suetrong et al. 2011a), USA (Raja et al. 2009b), Venezuela (Smits et al. 2007)


Key to freshwater Brachiosphaera species

  1. 1.

    Conidia with 4–5 arms………B. tropicalis

  2. 1.

    Conidia with 6–10, up to 10–13 arms………B. jamaicensis

Xylomyces Goos, Brooks & Lamore, Mycologia 69(2): 282 (1977)

Saprobic on submerged wood, leaves in freshwater or rotten leaves, root in terrestrial habitats. Sexual morph: Undetermined. Asexual morph: Colonies on natural substrate thin, effuse, reddish brown. Mycelium immersed and superficial, composed of branched, septate, fuscous hyphae. Stroma absent. Conidiophores and conidia absent. Chlamydospores abundant, broadly fusiform, brown to blackish, intercalary, solitary or catenate, occasionally branched, straight or curved, uniform in colour or sometimes end cells paler, with thick septate, distinctly constricted at the septa, thick-walled (Goos et al. 1977).

Type species: Xylomyces chlamydosporus Goos, R.D. Brooks & Lamore

Notes: The chlamydospores of Xylomyces are often found on submerged wood in freshwater habitats (Goos et al. 1977; Goh et al. 1997). The type species X. chlamydosporus was shown to belong in Aliquandostipitaceae based on molecular data (Campbell et al. 2007; Huang et al. 2018). Freshwater species X. aquaticus (Dudka) K.D. Hyde & Goh and X. elegans Goh et al. clustered in Pleosporales, therefore they were excluded from Xylomyces (Prihatini et al. 2008; Suetrong et al. 2011a; Tanaka et al. 2015). Five species are accepted in the genus, however, only X. chlamydosporus has been confirmed with molecular data. The species can be distinguished by chlamydospores size, septation and ornamentation of wall (see key below). This genus is in need of additional molecular studies to establish if other described species are phylogenetically related to X. chlamydosporus within Jahnulales (Fig. 14).

Fig. 14
figure 14

Phylogram generated from maximum likelihood analysis of combined LSU, SSU and ITS sequence data for species of Jahnulales. Bootstrap values for maximum likelihood equal to or greater than 70% and Bayesian posterior probabilities equal to or greater than 0.95 are placed near the branches as ML/BYPP. Newly generated sequences are in red and ex-type strains are in bold. The new species introduced in this study are indicated with underline. Freshwater strains are indicated with a red letter “F”. The tree is rooted to Cladosporium allicinum AFTOL-ID 1591 (Capnodiales)

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List of freshwater Xylomyces species


Xylomyces acerosisporus M.S. Oliveira, Malosso & R.F. Castañeda, Mycotaxon 130: (2015)

Freshwater distribution: Brazil (Oliveira et al. 2015)


*Xylomyces chlamydosporus Goos, R. D. Brooks & Lamore [as ‘chlamydosporis’], Mycologia 69(2): 282 (1977); Fig. 13a, b

Freshwater distribution: Australia (Hyde and Goh 1997), Brunei (Goh et al. 1997; Fryar et al. 2004), China (Luo et al. 2004), China (Goh et al. 1997; Tsui et al. 2001c), Seychelles (Hyde and Goh 1998b), USA (Goos et al. 1977; Lamore and Goos 1978; Raja et al. 2009b)


Xylomyces giganteus Goh, W.H. Ho, K.D. Hyde & K.M. Tsui, Mycol. Res. 101(11): 1324 (1997)

Freshwater distribution: Australia (Goh et al. 1997), China (Jiang et al. 2008), South Africa (Hyde et al. 1998)


Xylomyces punctatus Goh, W.H. Ho, K.D. Hyde & K.M. Tsui, Mycol. Res. 101(11): 1328 (1997)

Freshwater distribution: China (Goh et al. 1997; Tsui et al. 2001c)


Xylomyces pusillus Goh, W.H. Ho, K.D. Hyde & K.M. Tsui, Mycol. Res. 101(11): 1328 (1997)

Freshwater distribution: China (Goh et al. 1997; Tsui et al. 2001c; Cai et al. 2002a)


Key to freshwater Xylomyces species

  1. 1.

    Chlamydospores > 25 µm wide………2

  2. 1.

    Chlamydospores < 20 µm wide………3

  3. 2.

    Chlamydospores 5–9-septate………X. chlamydosporis

  4. 2.

    Chlamydospores 6–26-septate………X. giganteus

  5. 3.

    Chlamydospores with distinct punctate wall………X. punctatus

  6. 3.

    Chlamydospores lack distinct punctate wall………4

  7. 4.

    Chlamydospores 42–56 × 7–11 µm………X. pusillus

  8. 4.

    Chlamydospores 95–180 × 8–10 µm………X. acerosisporus


Kirschsteiniotheliales Hern.-Restr., R.F. Castañeda, Gené & Crous, Stud. Mycol. 86: 72 (2017)


Kirschsteiniotheliaceae Boonmee & K.D. Hyde, Mycologia 104(3): 705 (2012)


Kirschsteiniothelia D. Hawksw., J. Linn. Soc., Bot. 91: 182 (1985)


synonymy: Dendryphiopsis S. Hughes, Can. J. Bot. 31:655 (1953)

Saprobic mostly on dead wood in terrestrial or submerged wood in freshwater habitats. Sexual morph: Ascomata superficial, solitary or clustered, subglobose to globose, membranaceous, dark brown to black, with a central papilla. Peridium composed of several layers of cells of textura angularis. Pseudoparaphyses numerous, trabeculate, filiform, hyaline. Asci 8-spored, bitunicate, fissitunicate, cylindric-clavate, long pedicellate, apically rounded, with an ocular chamber. Ascospores biseriate, ellipsoidal, dull green, olive brown to dark brown at maturity, 1–2-septate, smooth-walled (Hawksworth 1985; Boonmee et al. 2012). Asexual morph: Hyphomycetous. Conidiophores macronematous, mononematous, erect, gregarious, elongate and thick-walled, straight and slightly curved, apically branched, septate, brown to dark brown, smooth-walled. Conidiogenous cells holoblastic, monoblastic, terminal, constricted at delimiting septa. Conidia broadly ellipsoid-obovoid, fusiform to obclavate, rounded at both ends, initially 1-septate, later becoming 2-septate, occasionally 3-septate, constricted and darkly pigmented at the septa, reddish brown to dark brown, grayish brown, smooth-walled (Hyde et al. 2013; Su et al. 2016b).

Type species: Kirschsteiniothelia atra (Corda) D. Hawksw.

Notes: Kirschsteiniothelia is characterized by superficial, subglobose to globose ascomata, cylindric-clavate asci and ellipsoidal, dull green, olive-brown to dark brown ascospores (Boonmee et al. 2012). The asexual morph of Kirschsteiniothelia has been connected to Dendryphiopsis, typified by D. atra (Corda) S. Hughes with molecular data (Boonmee et al. 2012; Hyde et al. 2013; Su et al. 2016b). For convenience, Wijayawardene et al. (2014) proposed to use Kirschsteiniothelia over Dendryphiopsis and named K. atra as the type species. All Kirschsteiniothelia species formed a well-supported clade and were placed in Kirschsteiniotheliaceae by Boonmee et al. (2012). Six freshwater species with molecular data have been accepted in the genus and all of which are asexual morphs (see list below). All freshwater species have unbranched conidiophores and mostly clavate conidia (slender conidia in K. fluminicola Z.L. Luo et al.), except K. aethiops (Sacc.) D. Hawksw. producing branched conidiophores and cylindrical conidia (Su et al. 2016b).

Fig. 15
figure 15

Kirschsteiniothelia spp. (Material examined: CHINA, Yunnan Province, Dali, Cangshan Mountain, Huadianba, saprobic on decaying wood submerged in stream, May 2014, Y. Su, S-144, HKAS 84022; ibid., on decaying wood submerged in Wanhua Stream, March 2014, X.Y. Liu, S-043, HKAS 93066, holotype). a–h Kirschsteiniothelia atra (HKAS 84022). a, b Colonies on the substratum. c Conidiophores with conidia. d, e Conidiogenous cells and conidia. f–h Conidia. i–o Kirchsteiniothelia submersa (HKAS 93066). i, j Conidiophores with conidia. k Conidiogenous cells with conidia. l–o Conidia. Scale bars: c = 130 μm, d, e = 50 μm, f–h, l–o = 20 μm, i, j = 100 μm, k = 30 μm

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List of freshwater Kirschsteiniothelia species


*Kirschsteiniothelia aquatica Z.L. Luo, K.D. Hyde & H.Y. Su, Mycosphere 9(4): 759 (2018)

Freshwater distribution: China (Bao et al. 2018)

*Kirschsteiniothelia atra (Corda) D. Hawksw., Fungal Diversity 69:37 (2014); Fig. 15a–h

Basionym: Dendryphion atrum Corda, Icon. fung. (Prague) 4: 33 (1840)

Synonymy: Amphisphaeria aethiops Sacc., Syll. fung. (Abellini) 1: 722 (1882)

Dendryphiopsis atra (Corda) S. Hughes, Can. J. Bot. 31: 655 (1953)

Sphaeria aethiops Berk. & Curtis, Grevillea 4: 143 (1876)

Kirschsteiniothelia aethiops (Sacc.) D. Hawksw., J. Linn. Soc., Bot. 91(1 -2): 185 (1985)

Freshwater distribution: China (Su et al. 2016b)


*Kirschsteiniothelia cangshanensis Z.L. Luo, D.F. Bao, K.D. Hyde & H.Y. Su, Mycosphere 9(4): 760 (2018)

Freshwater distribution: China (Bao et al. 2018)


*Kirschsteiniothelia fluminicola Z.L. Luo, K.D. Hyde & H.Y. Su, Mycosphere 9(4): 760 (2018)

Freshwater distribution: China (Bao et al. 2018)


*Kirschsteiniothelia rostrata J Yang & K.D. Hyde, Fungal Diversity 87: 45 (2017)

Freshwater distribution: China (Bao et al. 2018), Thailand (Hyde et al. 2017)


*Kirschsteiniothelia submersa H.Y. Su & K.D. Hyde, Fungal Diversity (2016); Fig. 15i–o

Freshwater distribution: China (Su et al. 2016b)


Key to freshwater Kirschsteiniothelia species

  1. 1.

    Conidiophores branched………K. atra

  2. 1.

    Conidiophores unbranched………2

  3. 2.

    Conidia slender, solitary to short-catenate………K. fluminicola

  4. 2.

    Conidia not slender, solitary………3

  5. 3.

    Conidia with sheath………4

  6. 3.

    Conidia without sheath………5

  7. 4.

    Conidia 33–43 μm long………K. cangshanensis

  8. 4.

    Conidia 80–150 μm long………K. rostrata

  9. 5.

    Conidiophores 114–151 × 7–8 μm………K. aquatica

  10. 5.

    Conidiophores 220–280 × 6–7 μm………K. submersa


Minutisphaerales Raja, Oberlies, Shearer & A.N. Mill., Mycologia 107(4): 854 (2015)


Acrogenosporaceae Jayasiri & K.D. Hyde, Mycosphere 9(4): 809 (2018)


Acrogenospora M.B. Ellis, Dematiaceous Hyphomycetes (Kew): 114 (1971)

Saprobic on decaying wood, bark, culms of bamboo in freshwater or terrestrial habitats. Sexual morph: Hysterothecia laterally compressed, with a prominent sunken slit, solitary to gregarious, erect and elevated, presenting an almost stipitate appearance, thick-walled. Pseudoparaphyses branched. Asci 8-spored, cylindrical, fusoid or obovate, obtuse at the apex, short pedicellate, thin-walled. Ascospores oval, aseptate, hyaline or moderately pigmented (Mason 1941; Jayasiri et al. 2018). Asexual morph: Hyphomycetous. Colonies effuse, dark brown to black, glistening, hairy. Mycelium mostly immersed, consist of septate, thin-walled, smooth, hyaline to pale brown hyphae. Conidiophores macronematous, mononematous, solitary, erect, subulate or cylindrical, straight or slightly flexuous, pale brown to dark brown, septate, unbranched, smooth. Conidiogenous cells holoblastic, monoblastic, integrated, terminal or intercalary, with percurrent proliferations, cylindrical. Conidia solitary, dry, acrogenous, simple, spherical, subspherical, olive to brown, aseptate, truncate at the base, smooth or verrucose (Ellis 1971; Hyde et al. 2019).

Type species: Acrogenospora sphaerocephala (Berk. & Broome) M.B. Ellis

Notes: Acrogenospora is a holomorphic genus, characterized by laterally compressed hysterothecia with a prominent sunken slit, oval, aseptate, hyaline ascospores and spherical, olive to brown conidia (Hyde et al. 2019). Acrogenospora is used to suppress Farlowiella Sacc. (Rossman et al. 2015) and their sexual-asexual connections were linked based on molecular data (Jayasiri et al. 2018). Thus, Acrogenospora is the only genus in Acrogenosporaceae (Jayasiri et al. 2018; Hyde et al. 2019). The sequence data of the type species A. sphaerocephala were generated based on a freshwater collection from Thailand (Hyde et al. 2019). Bao et al. (2020) introduced seven new species collected from a small area of Yunnan Province, China, which indicated Acrogenospora is a speciose genus. 20 species were reported in Acrogenospora and 13 have been confirmed with molecular data (Bao et al. 2020). Freshwater Acrogenospora species are morphologically very similar and the sequence data are main evidence to separate them.


List of freshwater Acrogenospora species

*Acrogenospora aquatica D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 3 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


*Acrogenospora basalicellularispora D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 8 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


*Acrogenospora ellipsoidea D.M. Hu, L. Cai & K.D. Hyde, Sydowia 62(2): 194 (2010)

Freshwater distribution: Yunnan, China (Hu et al. 2010a)


*Acrogenospora guttulatispora D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 8 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


*Acrogenospora obovoidspora D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 8 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


*Acrogenospora olivaceospora D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 10 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


Acrogenospora ovalis Goh, K.D. Hyde & C.K.M. Tsui [as ‘ovalia’], Mycol. Res. 102(11): 1312 (1998)

Freshwater distribution: Hong Kong (China) (Goh et al. 1998b)


*Acrogenospora sphaerocephala (Berk. & Broome) M.B. Ellis, Dematiaceous Hyphomycetes (Kew): 114 (1971)

Basionym: Monotospora sphaerocephala Berk. & Broome, Ann. Mag. nat. Hist., Ser. 3 3: 361 (1859)

Synonymy: Halysium sphaerocephalum (Berk. & Broome) Vuill., Bull. Séanc. Soc. Sci. Nancy, Sér. 3 11: 167 (1911)

Monosporella sphaerocephala (Berk. & Broome) S. Hughes, Can. J. Bot. 31: 654 (1953)

Monotosporella sphaerocephala (Berk. & Broome) S. Hughes, Can. J. Bot. 36: 787 (1958)

Freshwater distribution: China (Goh and Hyde 1999; Tsui et al. 2000), Philippines (Cai et al. 2003a), Seychelles (Hyde and Goh 1998b), Thailand (Sivichai et al. 2000; Hyde et al. 2019), USA (Lamore and Goos 1978; Shearer and Crane 1986)


*Acrogenospora submersa D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 12 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


*Acrogenospora subprolata Goh, K.D. Hyde & C.K.M. Tsui, Mycol. Res. 102(11): 1314 (1998)

Freshwater distribution: Australia (Goh et al. 1998b), China (Goh et al. 1998b; Tsui et al. 2001c; Bao et al. 2020), Seychelles (Goh et al. 1998b), South Africa (Goh et al. 1998b), UK (Goh et al. 1998b)


*Acrogenospora thailandica J. Yang & K.D. Hyde, Fungal Diversity 96: 78 (2019)

Freshwater distribution: Thailand (Hyde et al. 2019)


*Acrogenospora verrucispora Hong Zhu, L. Cai & K.Q. Zhang [as ‘verrucospora’], Mycotaxon 92: 384 (2005)

Freshwater distribution: Yunnan, China (Zhu et al. 2005; Bao et al. 2020)


*Acrogenospora yunnanensis D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Frontiers in Microbiology 11: 18 (2020)

Freshwater distribution: Yunnan, China (Bao et al. 2020)


Key to freshwater Acrogenospora species

  1. 1.

    Conidiophores 850–950 × 3.5–8 µm………A. thailandica

  2. 1.

    Conidiophores < 800 µm long………2

  3. 2.

    Conidiophores have wide range of length, 100–730 × 7.2–10.5 µm………A. sphaerocephala

  4. 2.

    Conidiophores not as above………3

  5. 3.

    Conidia with a small, hyaline basal cell………4

  6. 3.

    Conidia without basal cell………5

  7. 4.

    Conidiophores 202–250 × 7.5–9.5 µm………A. aquatica

  8. 4.

    Conidiophores 259–395 × 8–12 µm………A. basalicellularispora

  9. 5.

    Conidia hyaline when young, darker when mature………6

  10. 5.

    Conidia colour almost consistent………8

  11. 6.

    Conidiophores 163–223 × 6.7–10 µm………A. submersa

  12. 6.

    Conidiophores > 260 µm long………7

  13. 7.

    Conidiophores 7.5–8.6 µm wide………A. guttulatispora

  14. 7.

    Conidiophores 8.6–12 µm wide………A. yunnanensis

  15. 8.

    Conidia verrucose………A. verrucispora

  16. 8.

    Conidia smooth………9

  17. 9.

    Conidia ellipsoidal………A. ellipsoidea

  18. 9.

    Conidia not as above………10

  19. 10.

    Conidia 24–33 × 18–22 µm………A. ovalia

  20. 10.

    Conidia > 32 µm long………11

  21. 11.

    Conidia 39–46 × 30–39 µm………A. subprolata

  22. 11.

    Conidia < 39 µm long………12

  23. 12.

    Conidiophores 209–277 × 7.5–10 µm………A. obovoidspora

  24. 12.

    Conidiophores 102–172 × 5.8–9 µm………A. olivaceospora


Minutisphaeraceae Raja, Oberlies, Shearer & A.N. Mill., Mycologia 107(4): 854 (2015)


Minutisphaera Shearer, A.N. Mill. & A. Ferrer, Mycologia 103(2): 415 (2011)

Saprobic on submerged wood. Sexual morph: Ascomata scattered or clustered in groups, erumpent to superficial, uniloculate, globose to subglobose, brown, with central, rounded ostiole surrounded by black hairs. Peridium thin, composed of textura angularis to globulosa cells. Pseudoparaphyses sparse in young ascomata, becoming abundant with age, cellular, hyaline, septate, simple or branched, embedded in a gelatinous matrix. Asci 8-spored, bitunicate, fissitunicate, ovoid to obclavate, lacking a pedicel, rounded at the apex. Ascospores irregularly arranged, clavate, mostly straight, hyaline to pale brown, 1(–3)-septate, constricted at the septum, multiguttulate, asymmetrical, smooth- or rough-walled when mature, with sheath and filamentous appendages (Ferrer et al. 2011). Asexual morph: Undetermined.

Type species: Minutisphaera fimbriatispora Shearer, A.N. Mill. & A. Ferrer

Notes: All species reported within Minutisphaera were collected from freshwater habitats (Ferrer et al. 2011; Raja et al. 2013b, 2015; Bao et al. 2019a). The ascospores of Minutisphaera are unusual in being asymmetrical and often equipped with a sheath and filamentous appendages (Ferrer et al. 2011; Raja et al. 2015). The shape and size of ascospores, and features of appendages and sheaths are used for distinguishing all species within the genus (see key below).

In the analysis of Raja et al. (2015) BLAST search of ITS sequences revealed that two isolates, Pleosporales sp. 39g (JX244063) and Didymosphaeria sp. TS_04_050 (HQ713763), shared 99% similarity with Minutisphaera aspera. Both strains were isolated as fungal endophytes in previous studies (Grünig et al. 2011). Thus, it was hypothesized that members of Minutisphaerales may lead a dual life as saprobes on submerged wood and as fungal endophytes of trees living in close proximity to rivers and streams. However, additional studies are needed to shed light on the dual ecological life style of Minutisphaera spp.


List of freshwater Minutisphaera species

*Minutisphaera aquaticum D.F. Bao, L. Xu & H.Y. Su, Asian Journal of Mycology 2(1): 309 (2019)

Freshwater distribution: Thailand (Bao et al. 2019a)


*Minutisphaera aspera Raja, Oberlies, Shearer & A.N. Mill, Mycologia 107(4): 854 (2015); Fig. 16a–c

Fig. 16
figure 16

Minutisphaera spp. (Material examined: USA, North Carolina: Rockingham County, Piedmont Plateau, Big Beaver Island Creek, Madison, on submerged decorticated wood, 26 April 2013, H.A. Raja & N.H. Oberlies, G427-1, ILLS 72340, holotype; ibid., G156-4, ILLS 72342, paratype). a–c M. aspera (G427-1). a Ascoma on wood. b Ascus. c Ascospores. d–f M. parafimbriatispora (G156-4). d Ascoma in culture. e Asci. f Ascospore with gelatinous sheath constricted at the mid-septum. Scale Bars: a = 100 µm, b, c = 20 µm, d = 50 µm, e, f = 10 µm

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Freshwater distribution: USA (Raja et al. 2015)


*Minutisphaera fimbriatispora Shearer, A.N. Mill. & A. Ferrer, Mycologia 103(2): 415 (2011)

Freshwater distribution: USA (Ferrer et al. 2011; Raja et al. 2013b)


*Minutisphaera japonica Kaz. Tanaka, Raja & Shearer, Mycologia 105(4): 966, (2013); Fig. 17

Fig. 17
figure 17

Minutisphaera japonica (Material examined: FRANCE, Ariège, Vernajoul, ruisseaude Vernajoul, on submerged wood of Fraxinus excelsior, 2 July 2007, J. Fournier, JF07132, associated with Pseudohalonectria lutea and Aniptodera sp., leg det JF). a Ascomata on the wood surface. b Squash mount of ascoma, note dark-walled hyphae on the peridial wall. c Longitudinal section of apothecioid ascoma. d–f Asci. g, h Ascospores. Scale bars: a = 200 µm, b, d, f, h = 20 µm, c = 50 µm, e = 30 µm, g = 10 µm

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Freshwater distribution: Japan (Raja et al. 2013b), France (Raja et al. 2015)


*Minutisphaera parafimbriatispora Raja, Oberlies, Shearer & A.N. Mill, Mycologia 107(4): 855 (2015); Fig. 16d–f

Freshwater distribution: USA (Raja et al. 2015)


Key to freshwater Minutisphaera species

  1. 1.

    Ascospores with sheath, but lacking appendages………2

  2. 1.

    Ascospores with sheath and appendages………4

  3. 2.

    Ascospores with rough-walled………M. aspera

  4. 2.

    Ascospores smoth-walled………3

  5. 3.

    Ascospores broadly fusiform, with acute apex………M. japonica

  6. 3.

    Ascospores fusiform to clavate, with rounded apex………M. aquaticum

  7. 4.

    Ascospores 24–36 × 6–8 µm………M. fimbriatispora

  8. 4.

    Ascospores 18–25 × 4–7 µm………M. parafimbriatispora


Natipusillales Raja, Shearer, A.N. Mill. & K.D. Hyde, Fungal Diversity 63: 9 (2013)


Natipusillaceae Raja, Shearer & A.N. Mill., Mycologia 104(2): 570 (2012)


Natipusilla A. Ferrer, A.N. Mill. & Shearer, Mycologia 103(2): 417 (2011)

Saprobic on submerged wood. Sexual morph: Ascomata scattered or aggregated, erumpent to superficial, uniloculate, globose to subglobose, hyaline to brown, lacking ostioles, occasionally with minute papilla. Peridium thin, membranous, glabrous, forming a textura angularis in surface view. Pseudoparaphyses sparse or absent. Asci 8-spored, bitunicate, fissitunicate, globose to obclavate, lacking a pedicel, with or without an ocular chamber. Ascospores overlapping tri- to multi-seriate, arranged in a group, fusiform or clavate, straight, the upper cell often shorter and broader than the lower one, mostly 1-septate, hyaline, occasionally becoming 3-septate, pale brown at age, guttulate when young, smooth, thin-walled, with or without a gelatinous sheath (Ferrer et al. 2011). Asexual morph: Undetermined.

Type species: Natipusilla decorospora A. Ferrer, A.N. Mill. & Shearer

Notes: All species described in this genus were reported from freshwater habitats in tropical and subtropical regions (Ferrer et al. 2011; Raja et al. 2012). Phylogenetic analysis using combined SSU and LSU sequence dataset placed this genus in a distinct clade within Dothideomycetes. Therefore, a novel family, Natipusillaceae (Raja et al. 2012), and new order Natipusillales (Hyde et al. 2013) were established for Natipusilla.

Ascominuta Ranghoo & K.D. Hyde was thought to be an earlier name of Natipusilla (Hyde et al. 2013) due to the bitunicate, globose asci, sheathed ascospores and freshwater habitat (Ranghoo and Hyde 1999; Hu et al. 2010a; Ferrer et al. 2011; Raja et al. 2012). However, Ascominuta differs from Natipusilla in the numbers of ascospores per asci (4 vs. 8) and colour of ascomata (hyaline to brown vs. dark brown to black). Only LSU sequence (AF132335) of A. lignicola Ranghoo & K.D. Hyde (generic type) was deposited in Genbank, and it cannot be aligned with Natipusilla. Further molecular and morphological data are needed to clarify these two genera (Raja et al. 2012). All Natipusilla species are surrounded by a complex gelatinous sheath, except N. naponensis A. Ferrer et al. lacks a sheath (Ferrer et al. 2011; Raja et al. 2012).


List of freshwater Natipusilla species

*Natipusilla decorospora A. Ferrer et al., Mycologia 103(2): 417 (2011)

Freshwater distribution: Ecuador (Ferrer et al. 2011)


*Natipusilla limonensis A. Ferrer, A.N. Mill. & Shearer, Mycologia 103(2): 417 (2011); Fig. 18

Fig. 18
figure 18

Natipusilla limonensis (Material examined: PERU, Cusco, Camanti, Stream at Quince Mil Trail 1, on submerged wood, 26 May 2010, S. Zelski & H.A. Raja, PE0003). a Section of ascoma. b Globose to subglobose asci. c–f Ascospores. Arrows indicating complex gelatinous sheath. Scale bars: a = 50 µm, b–f = 20 µm

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Freshwater distribution: Costa Rica (Ferrer et al. 2011), Peru (Shearer et al. 2015)


*Natipusilla naponensis A. Ferrer, A.N. Mill. & Shearer, Mycologia 103(2): 420 (2011)

Freshwater distribution: Ecuador (Ferrer et al. 2011)


*Natipusilla bellaspora Raja, Shearer & A.N. Mill., Mycologia 104(2): 570 (2012)

Freshwater distribution: Peru (Raja et al. 2012; Shearer et al. 2015)


Key to freshwater Natipusilla species

  1. 1.

    Ascospores without sheath………N. naponensis

  2. 1.

    Ascospores with complex gelatinous sheaths………2

  3. 2.

    Ascospores 40–48 × 10–13 µm………N. bellaspora

  4. 2.

    Ascospores < 40 µm long………3

  5. 3.

    Ascospores 30–37 × 6–9 µm, with a single scythe shaped appendage at the apex………N. decrospora

  6. 3.

    Ascospores 35–40 × 9–11 µm, without appendage………N. limonensis


Pleosporales Luttrell ex M.E. Barr


Acrocalymmaceae Crous & Trakun., IMA Fungus 5(2): 404 (2014)


Acrocalymma Alcorn & J.A.G. Irwin, Trans. Br. mycol. Soc. 88(2): 163 (1987)

Saprobic on submerged wood, leaf litter or other plants. Sexual morph: (from culture) Ascomata scattered, erumpent becoming superficial, uniloculate, globose, covered with light gray, gnarled, warted hairs, black, coriaceous, with ostiolate papilla. Neck central, terete, composed of 2–5 layers of brown, polygonal cells around ostiole with hyaline periphyses. Peridium thin, composed of polygonal, brown, pseudoparenchyma cells. Pseudoparaphyses numerous, septate, anastomosing. Asci bitunicate, 8-spored, cylindrical, short pedicellate, apically rounded, with an distinct ocular chamber. Ascospores uni- to bi-seriate, narrowly fusiform, straight or slightly curved, hyaline first, pale reddish brown with lighter coloured end cells, 1–3-septate, constricted at first submedian septum, basically symmetrical, smooth to rough-walled, thin-walled, with 4 guttules, with a thin sheath (Shoemaker et al. 1991). Asexual morph: Coelomycetous. Pycnidia scattered or aggregated, semi-immersed to superficial, unilocular, globose, brown to black, with ostiolate papilla or rostrate. Conidiophores reduced. Conidiogenous cells holoblastic, determinate, discrete, cylindrical, lageniform to ampulliform, hyaline, smooth. Conidia cylindrical to fusoid with subobtuse apex, straight, hyaline at first, becoming pigmented with age, 0–3-septate, not constricted at septa, smooth, thin-walled, with or without helmet-shaped appendages (Zhang et al. 2012a).

Type species: Acrocalymma medicaginis Alcorn & J.A.G. Irwin

Notes: Acrocalymma is well-studied because all species have sequences in GenBank. Acrocalymma species are easy to produce pycnidia in culture (Trakunyingcharoen et al. 2014). The sexual morph was linked to Massarina walkeri Shoemaker et al. which formed in the pure culture of Acrocalymma medicaginis (Shoemaker et al. 1991). However, these two species are phylogenetically distinct (Trakunyingcharoen et al. 2014). Therefore, Trakunyingcharoen et al. (2014) transferred M. walkeri to Acrocalymma and treated A. medicaginis and A. walkeri as distinct species. The type material of A. medicaginis was re-examined and described by Li et al. (2020). Acrocalymma aquatica is the first species reported from freshwater habitats, and it is characterized by cylindrical to fusoid, 0(–1)-septate conidia with an apical, mucilaginous, helmet-shaped appendage (Zhang et al. 2012a). In this study, we report a new freshwater species A. bipolare, which is collected from Egypt.


List of freshwater Acrocalymma species

*Acrocalymma aquatica H. Zhang & K.D. Hyde, Cryptog. Mycol. 33(3): 337 (2012); Fig. 19

Fig. 19
figure 19

Acrocalymma aquatica (Material examined: THAILAND, Chiang Mai Province, Doi Inthanon, on submerged wood, 16 November 2010, H. Zhang, d67, MFLU 11-1113, holotype). a Pycnidia on substrate surface. b–d Conidia with single apical appendage. Scale bars: b–d = 5 μm

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Freshwater distribution: Thailand (Zhang et al. 2012a)


*Acrocalymma bipolare Abdel-Aziz & Abdel-Wahab, sp. nov.

Index Fungorum number: IF557901; Facesoffungi number: FoF09243; Fig. 20

Fig. 20
figure 20

Acrocalymma bipolare (CBS H-22673, holotype). a Vertical section of a pycnidium. b Section through the peridium. c–f Variously shaped conidia with mucoid polar appendages. Note the apical appendages formed long filament (arrowed in d, stained in toluidine blue). Scale bars: a = 50 μm, b, c = 10 μm, d–f = 5 μm

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Etymology: in reference to the polar mucoid appendages of the conidia

Holotype: CBS H-22673

Saprobic on submerged wood in freshwater habitats. Sexual morph: Undetermined. Asexual morph: Pycnidia 175–230 µm diam., globose or subglobose, erumpent to superficial, solitary, dark brown to black, without ostiolate papilla. Peridium 8–17 µm thick, composed of 8–11 layers of dark brown to black, thick walled, melanized cells of textura angularis, inwardly with narrow lumens, hyaline and thin walled cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells 2.5–7 × 2–3.5 μm, holoblastic, determinate, discrete, cylindrical or lageniform, hyaline, formed from the inner cells of the pycnidial wall. Conidia 9–12 × 3–5 µm (\( {\bar{\text{x}}} \) = 10.8 × 4.3, n = 50), cylindrical to fusiform, straight, unicellular, hyaline, thin-walled, guttulate, with rounded apex and slightly narrow, truncate base, with mucoid polar appendages that filled with oil droplets, appendages elongate in water to form filaments. Apical appendages 4–6 μm diam., rounded. Lower appendages 1–2 μm in length, sometimes split in the middle.

Culture characteristics: Colonies on PDA up to 22 mm diam. after 2 weeks at 23 °C, with grey to dark brown aerial and immersed mycelium; reverse dark brown.

Material examined: EGYPT, Sohag City, River Nile, 26°33′32ʺN, 31°41′44ʺE, on decaying wood submerged in River Nile water, 14 August 2012, F.A. Abdel-Aziz, (CBS H-22673, holotype), ex-type living culture MD1321.

Notes: Acrocalymma bipolare clusters with the type species A. medicaginis and A. pterocarpi Jayasiri et al. with high bootstrap support (Fig. 2). Morphologically, Acrocalymma bipolare is different from A. medicaginis by its appendages that are filled with oil droplets and become filaments when mounted in water, while A. medicaginis processes globose to hemispherical or helmet-shaped appendages which do not form filaments in water (Alcorn and Irwin 1987). In addition, A. bipolare has shorter conidia than A. medicaginis (9–12 × 3–5 µm vs. 11–21 × 3.5–5 µm). The phylogenetically close species A. pterocarpi did not produce asexual morph in culture, thus their morphology can not be compared (Jayasiri et al. 2019).

Acrocalymma bipolare only has LSU sequence data, and it has four and six nucleotide differences with A. medicaginis and A. pterocarpi, respectively, which indicates them to be different species. This is also supported by our phylogenetic analysis (Fig. 2).

Acrocalymma bipolare differs from the other freshwater species A. aquatica in having shorter conidia (9–12 × 3–5 μm vs. 12–17 × 3–4 μm) and bipolar appendages compared with a single appendage in A. aquatica (Zhang et al. 2012a).


Key to freshwater Acrocalymma species

  1. 1.

    Conidia 12–17 × 3–4 μm, with a single, helmet-shaped appendage………A. aquatica

  2. 1.

    Conidia 9–12 × 3–5 μm, with bipolar appendages filled with oil droplets, appendages elongate in water to form filaments………A. bipolare


Aigialaceae Suetrong, Sakay., E.B.G. Jones, Kohlm., Volkm.-Kohlm. & C.L. Schoch, Stud. Mycol. 64: 166 (2009)


Key to freshwater genera of Aigialaceae

  1. 1.

    Asci cylindro-clavate, ascospores often hyaline, smooth………Fissuroma

  2. 1.

    Asci obclavate, ascospores often brown, verrucose………Neoastrosphaeriella


Fissuroma J.K. Liu, Phook., E.B.G. Jones & K.D. Hyde, Fungal Diversity 51(1): 145 (2011)

Saprobic on submerged wood, dead stem of bamboo or other plant materials. Sexual morph: Ascomata scattered, rarely clustered, solitary to gregarious, immersed beneath host epidermis, becoming raised, uniloculate, rarely biloculate joined at the base, hemispherical domes, glabrous, black, coriaceous or carbonaceous, opening with a slit-like ostiole. Peridium thick, uneven in thickness, poorly developed at the base, thick at sides towards the apex, composed of dark brown pseudoparenchymatous cells, with host cells, plus fungal tissue, arranged in a textura angularis to textura epidermoidea, carbonaceous at slit-like opening. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, embedded in a gelatinous matrix and anastomosing between and above the asci. Asci 8-spored, bitunicate, fissitunicate, obclavate to cylindrical, pedicellate, apically rounded, with a small ocular chamber. Ascospores overlapping uni- to tri-seriate, fusiform, straight to slightly curved, hyaline, 1-septate, slightly constricted at the septum, smooth, thin-walled, surrounded by a mucilaginous sheath (Liu et al. 2011). Asexual morph: Coelomycetous, reported as pleurophomopsis-like from culture of Fissuroma aggregata (KT 767). Conidiomata globose to subglobose. Conidiophores cylindrical, simple to branched, septate. Conidiogenous cells phialidic. Conidia globose, hyaline (Tanaka and Harada 2005).

Type species: Fissuroma maculans (Rehm) J.K. Liu, E.B.G. Jones & K.D. Hyde

Notes: Fissuroma comprises 12 species in Index Fungorum (2020). Eight species have been confirmed with molecular data (Liu et al. 2011; Phookamsak et al. 2015b; Tennakoon et al. 2018; Wanasinghe et al. 2018a). However, Phookamsak et al. (2015b) indicated that F. aggregata (I. Hino & Katum.) Phook. et al. is a species complex which might comprise at least three distinct species based on their phylogenetic analyses. After re-examination of the holotype specimen of F. aggregata (≡ Melanopsamma aggregata), Phookamsak et al. (2015b) considered the collection MFLU 11-0146 of Liu et al. (2011) as a distinct species from F. aggregata. Therefore, F. neoaggregatum Phook. & K.D. Hyde was introduced based on the specimen MFLU 11-0146 (Phookamsak et al. 2015b). The asexual morph was developed in the culture of F. aggregata (KT 767) (Tanaka and Harada 2005). However, the real identity of KT 767 needs confirmation to separate F. aggregata KT 984, as shown in the phylogenetic tree (Fig. 2). We provide a description and illustration for F. neoaggregatum collected from submerged wood in Thailand.


List of freshwater Fissuroma species

*Fissuroma neoaggregatum Phook. & K.D. Hyde [as ‘neoaggregata’], Fungal Diversity 74: 158 (2015); Fig. 21

Fig. 21
figure 21

Fissuroma neoaggregatum (MFLU 11-1140, new habitat record). a Appearance of ascomata on wood. b Asci and pseudoparaphyses. c–e Asci. Note the bitunicate ascus in e. f Pseudoparaphyses. g–i Ascospores. (i in Indian Ink). j Front view of colony on PDA. Scale bars: b–f = 50 μm, g–i = 20 μm

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Freshwater distribution: Thailand (this study)

Saprobic on submerged wood. Sexual morph: Ascostromata dark brown to black, solitary to gregarious, immersed in host epidermis, visible as raised, dome-shaped areas on the host surface, hemisphaerical to depressed conical, with flattened base, uniloculate, coriaceous, ostiolate, with slit-like opening. Peridium of unequal thickness, poorly developed at the base. Pseudoparaphyses dense, trabeculate, filiform, embedded in a hyaline gelatinous matrix. Asci 140–155 × 16–21 μm (\( {\bar{\text{x}}} \) = 149 × 19 μm, n = 5), 8-spored, bitunicate, obclavate, with furcate to truncate pedicel, apically rounded, with an ocular chamber. Ascospores 36–43 × 6–8 μm (\( {\bar{\text{x}}} \) = 40.5 × 7 μm, n = 10), overlapping biseriate and uniseriate at the apex, hyaline, fusiform, with acute ends, 1-septate, constricted at the septum, smooth-walled, guttulate, surrounded by a gelatinous sheath. Asexual morph: Undetermined.

Material examined: THAILAND, Chiang Rai Province, on submerged wood, 18 January 2010, H. Zhang, a45 (MFLU 11-1140), living culture MFLUCC 10-0917.

Notes: Our new collection MFLUCC 10-0917 is identified as Fissuroma neoaggregatum based on nearly identical ITS sequence data with ex-type strain MFLUCC 10-0554 (two nucleotide differences) and morphological characters. Our collection has shorter asci (140–155 × 16–21 μm vs. 155–197 × 15–18) μm) and ascospores (36–43 × 6–8 μm vs. 39–54 × 7–9 μm) than the holotype (Phookamsak et al. 2015b). Moreover, our collection shows a broad, mucilaginous sheath in Indian Ink, while the holotype has only a thin sheath (Phookamsak et al. 2015b). Other sequences are expected for MFLUCC 10-0917 to further clarify this species. This is a new habitat record for F. neoaggregatum from freshwater.


Neoastrosphaeriella J.K. Liu, E.B.G. Jones & K.D. Hyde, Fungal Diversity 51(1): 148 (2011)

Saprobic on decaying twigs, petiole and submerged wood in terrestrial and freshwater habitats. Sexual morph: Ascomata scattered, immersed to semi-immersed, with a slit-like ostiole, black. Peridium uneven in thickness, carbonaceous, composed of dark brown, thick-walled cells. Pseudoparaphyses trabeculate, hyaline, anastomosing, embedded in a gelatinous matrix. Asci 8-spored, bitunicate, fissitunicate, obclavate to cylindrical, pedicellate, with a small ocular chamber. Ascospores fusiform, hyaline to dark brown, mostly verrucose when mature, 1-septate, surrounded by a mucilaginous sheath (Liu et al. 2011). Asexual morph: Undetermined.

Type species: Neoastrosphaeriella krabiensis J.K. Liu, E.B.G. Jones & K.D. Hyde

Notes: Neoastrosphaeriella was established by Liu et al. (2011) to accommodate species having ascomata with slit-like ostioles, obclavate asci and brown, verrucose ascospores. Neoastrosphaeriella aquatica D.F. Bao et al. was the only species found in freshwater habitats and differed from the others by hyaline to greyish brown, verrucose ascospores and distinct molecular characters (Bao et al. 2019b). Neoastrosphaeriella nested well in Aigialaceae (Liu et al. 2011; Wanasinghe et al. 2018a; Bao et al. 2019b; Zhang et al. 2020; this study, Fig. 2).


List of freshwater Neoastrosphaeriella species

*Neoastrosphaeriella aquatica D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Phytotaxa 391(3): 201 (2019)

Freshwater distribution: Thailand (Bao et al. 2019b)


Amniculicolaceae Y. Zhang ter, C.L. Schoch, J. Fourn., Crous & K.D. Hyde, Stud. Mycol. 64: 95 (2009)


Key to freshwater genera of Amniculicolaceae

  1. 1.

    Asexual morph………Vargamyces

  2. 1.

    Sexual morph………2

  3. 2.

    Ascospores hyaline, 1-septate………Amniculicola

  4. 2.

    Ascospores pigmented, multi-septate………3

  5. 3.

    Ascospores golden yellow to brown, dictyoseptate………Murispora

  6. 3.

    Ascospores reddish brown, phragmoseptate………Neomassariosphaeria


Amniculicola Y. Zhang ter & K.D. Hyde, Mycological Research 112 (10): 1189 (2008)

Saprobic on submerged wood or in plant humus. Sexual morph: Ascomata solitary to gregarious, immersed to nearly superficial, black, uniloculate, subglobose to conical, glabrous, ostiolate, with or without two tuberculate flared lips surrounding a slit-like ostiole, sometimes with a flattened base not easily removed from the substrate, usually staining the woody substrate purple. Peridium 2-layered, outer layer composed of heavily pigmented thick-walled cells of textura angularis, inner layer composed of hyaline thin-walled cells of textura angularis. Pseudoparaphyses dense, trabeculate, filiform, persistent, hyaline, embedded in mucilage, anastomosing between and above the asci. Asci 8-spored, bitunicate, fissitunicate, cylindrical to narrowly fusoid, short pedicellate. Ascospores mostly uniseriate, fusoid, hyaline, septate, symmetrical, smooth, thin-walled, surrounded by a hyaline, gelatinous sheath (Zhang et al. 2008c, 2009b, 2012b). Asexual morph: Hyphomycetous. Conidiophores usually simple. Conidia curved or sigmoid, tapering to the end, septate, hyaline (illustration and description see Ingold (1942)).

Type species: Amniculicola lignicola Y. Zhang ter & K.D. Hyde

Notes: The freshwater genus Amniculicola is characterized by staining the woody substrate purple (Zhang et al. 2008c, 2009b). However, two other species, A. aquatica Z.L. Luo et al. and A. guttulata et al., did not produce pigmentation on the host substrate (Hyde et al. 2019; this study, Fig. 22). A comprehensive account of Amniculicola was provided by Zhang et al. (2008c, 2012b). Amniculicola immersa Y. Zhang ter, J. Fourn., A. lignicola and A. parva Y. Zhang ter, J. Fourn. were collected from Europe (Denmark and France), A. longissima (Sacc. & P. Syd.) Nadeeshan & K.D. Hyde was collected from Latin America (Costa Rica), while A. aquatica and A. guttulata were collected from Asia (China). Amniculicola longissima is the only asexual morphic species in the genus and confirmed by phylogenetic analyses (Hyde et al. 2019; this study, Fig. 2).

Fig. 22
figure 22

Amniculicola guttulata (MFLU 18-1160). a Appearance of black ascomata on host. b Vertical section of ascoma. c Structure of peridium. d Pseudoparaphyses. e–h Bitunicate asci. i–n Ascospores. o Germinated ascospore. p Colony on PDA (from front and reverse). Scale bars: b = 50 μm, c, e–h = 20 μm, d, i–o = 10 μm

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The paraphyletic nature of Amniculicola was shown in Wanasinghe et al. (2015), as Amniculicola species clustered in three different sister clades. This nature is not well resolved as one asexual genus Vargamyces Tóth nested within Amniculicola in Hyde et al. (2019) and this study (Fig. 2).


List of freshwater Amniculicola species

*Amniculicola aquatica Z.L. Luo, K.D. Hyde & H.Y. Su, Fungal Diversity 96: 13 (2019)

Freshwater distribution: China (Hyde et al. 2019)


*Amniculicola guttulata Z.L. Luo, K.D. Hyde & H.Y. Su, Fungal Diversity 96: 13 (2019); Fig. 22

Freshwater distribution: China (Hyde et al. 2019, this study)

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 160–180 μm high, 170–190 μm diam., black, solitary, scattered or occasionally gregarious, superficial, subglobose to conical, with ostiolate papilla. Peridium 35–45 μm thick, comprising several layers of hyaline to pale brown, thin-walled, large cells of textura angularis or irregular cells. Pseudoparaphyses 2.2 μm diam., dense, trabeculate, filiform, persistent, hyaline, embedded in mucilage, anastomosing between and above the asci. Asci 105–141(–256) × 8.5–10.5 μm (\( {\bar{\text{x}}} \) = 158.5 × 10 μm, n = 10), 8-spored, bitunicate, fissitunicate, cylindrical to narrowly fusiform, with a twisted, bifurcate, short pedicel, 7.5–18.5 μm long, apically rounded or slightly obtuse, endoascus elongate to 256 μm long when mature, with a small inconspicuous apical apparatus barely seen in water. Ascospores 21.5–27 × 5.2–6.2 μm (\( {\bar{\text{x}}} \) = 23.7 × 5.6 μm, n = 15), uni-seriate or overlapping uni-seriate, straight or slightly curved, hyaline, 1-septate, obscure when young, visible when germinated, deeply constricted at the septa, slightly constricted at a quarter, the upper cell slightly broader than the lower one, broadly fusiform, with two prominent big guttules in the middle cell and four gradually smaller ones beside, thin-walled, smooth, with a thin, hyaline, bipolar narrowed, mucilaginous sheath which is 1–2 μm thick. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 25 mm diam. in 25 days at 25 °C, grey to white from above, pale grey from below, surface rough, fluffy, with dense mycelium, dry, raised, edge entire, with white margin.

Material examined: CHINA, Yunnan Province, Dehong, on submerged wood in a stream, 25 November 2017, G.N. Wang, H55A-1 (MFLU 18-1160), living culture MFLUCC 18-1038; ibid., H55A-2 (HKAS 101736), living culture KUMCC 18-0088.

Notes: Our new collection MFLUCC 18-1038 is identified as Amniculicola guttulata based on morphological characters; identical LSU and TEF sequence data and phylogenetic analysis (Fig. 2). The asci in the holotype are up to 127 μm long, while in our collection they are up to 256 μm long when mature (Hyde et al. 2019).


*Amniculicola immersa Y. Zhang ter, J. Fourn., Crous & K.D. Hyde, Persoonia 23: 50 (2009)

Freshwater distribution: Denmark (Zhang et al. 2009b)


*Amniculicola lignicola Y. Zhang ter & K.D. Hyde, Mycological Research 112 (10): 1189 (2008)

Freshwater distribution: France (Zhang et al. 2008c)


*Amniculicola longissima (Sacc. & P. Syd.) Nadeeshan & K.D. Hyde, IMA Fungus 7(2): 301 (2016)

Freshwater distribution: Leicester, UK (Ingold 1942)


*Amniculicola parva Y. Zhang ter, J. Fourn., Crous & K.D. Hyde, Persoonia 23: 52 (2009)

Freshwater distribution: France (Zhang et al. 2009b)


Key to freshwater Amniculicola species

  1. 1.

    Asexual morph………A. longissima

  2. 1.

    Sexual morph………2

  3. 2.

    Ascomata superficial………3

  4. 2.

    Ascomata immersed………A. immersa

  5. 3.

    Asci longer than 130 μm………A. lignicola

  6. 3.

    Asci shorter than 130 μm………4

  7. 4.

    Substrate stained purple………A. parva

  8. 4.

    Substrate natural colour………5

  9. 5.

    Peridium 35–50 µm thick, ascospores 24–32 × 6–8 µm………A. aquatica

  10. 5.

    Peridium 27–35 µm thick, ascospores 23–27 × 5–7 µm………A. guttulata


Murispora Yin. Zhang, C.L. Schoch, J. Fourn., Crous & K.D. Hyde, Studies in Mycology 64: 95 (2009)

Saprobic on submerged wood or dead stems or branches of plant materials. Sexual morph: Ascomata scattered to gregarious, immersed, uniloculate, lenticular, slightly protruding at the apex, opening through a small rounded pore, with ostiolate papilla, stain substrate purple. Peridium thin, composed of a few layers cells of textura angularis, thicker at the apex with pseudoparenchymatous cells. Pseudoparaphyses cellular, embedded in mucilage. Asci 8-spored, bitunicate, fissitunicate, cylindro-clavate, short pedicellate. with a small ocular chamber. Ascospores biseriate to uniseriate near the base, fusoid with narrowly rounded ends, slightly curved, golden yellow or hyaline, turning brown when senescent, muriform, with one, rarely two longitudinal septa in all cells except end cells, 7–9-transversally septate, basically symmetrical, smooth or finely verruculose, surrounded by a wide mucilaginous sheath (Zhang et al. 2009c, 2012b). Asexual morph: Coelomycetous. Pycnidia solitary, mainly immersed, uniloculate, dark brown to black. Peridium reddish brown cells of textura angularis, with inner most layer thin, hyaline. Conidiophores reduced to conidiogenous cells. Conidiogenous cells phialidic, determinate, hyaline, smooth, formed from the inner most layer of pycnidium wall. Conidia ellipsoidal, hyaline, aseptate, smooth-walled, thin-walled (Wanasinghe et al. 2015).

Type species: Murispora rubicunda (Niessl) Y. Zhang ter, J. Fourn. & K.D. Hyde

Notes: Zhang et al. (2009c) introduced Murispora based on Pleospora rubicunda Niessl which clustered in a robust clade with Amniculicola species in Amniculicolaceae. Wanasinghe et al. (2015) introduced another six species to Murispora from terrestrial habitats and they found the asexual morph of M. hawksworthii Wanas. et al. from single ascospore isolates. Murispora species stain substrate purple (Zhang et al. 2009c, 2012b; Wanasinghe et al. 2015; Bao et al. 2019d) and this character should be a common feature to the genus. Four freshwater species have been reported in Murispora, three of which were collected from China (see list below).


List of freshwater Murispora species

*Murispora aquatica D.F. Bao, Z.L. Luo, K.D. Hyde & H.Y. Su, Phytotaxa 416(1): 5 (2019)

Freshwater distribution: China (Bao et al. 2019d)


*Murispora cicognanii Wanas., Camporesi, E.B.G. Jones & K.D. Hyde, Cryptog. Mycol. 36(4): 437 (2015)

Freshwater distribution: China (Hyde et al. 2019)


*Murispora fagicola Wanas., Camporesi, E.B.G. Jones & K.D. Hyde, Cryptog. Mycol. 36(4): 429 (2015)

Freshwater distribution: China (Bao et al. 2019d)


*Murispora rubicunda (Niessl) Yin. Zhang, J. Fourn. & K.D. Hyde, Studies in Mycology 64: 96 (2009); Fig. 23

Fig. 23
figure 23

Murispora rubicunda (Material examined: FRANCE, Haute-Garonne, Avignonet, on submerged dead herbaceous stem, 16 January 2007, M. Delpont, IFRD 2017). a Herbarium label and specimen of Murispora rubicunda. b Appearance of ascomata on host surface. c Vertical section through ascoma. d Peridium. e, f Asci. g Apex of ascus. h Pedicel. j Pseudoparaphyses. i, k Ascospores with wide sheath in Indian Ink. Scale bars: c = 100 μm, d = 20 μm, e, f = 25 μm, g–k = 10 μm

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Basionym: Pleospora rubicunda Niessl, Verh. nat. Ver. Brünn 14: 191 (1876)

Synonymy: Sphaeria rubicunda Schwein., Transactions of the American Philosophical Society 4 (2): 222 (1832)

Massariosphaeria rubicunda (Niessl) Crivelli, Über die heterogene Ascomycetengattung Pleospora Rabh.: 144 (1983)

Karstenula rubicunda (Niessl) M.E. Barr, N. Amer. Fl., Ser. 2 (New York) 13: 52 (1990)

Freshwater distribution: Austria (Magnes and Hafellner 1991), France (Zhang et al. 2012b)


Key to freshwater Murispora species

  1. 1.

    Ascospores > 31 μm long………2

  2. 1.

    Ascospores < 31 μm long………3

  3. 2.

    Ascospores more than 10 transverse septa………M. cicognanii

  4. 2.

    Ascospores 7–9 transverse septa………M. rubicunda

  5. 3.

    Ascospores 21–25 × 8–10 μm………M. aquatica

  6. 3.

    Ascospores 27–31 × 11–13 μm………M. fagicola


Neomassariosphaeria Y. Zhang ter, J. Fourn. & K.D. Hyde, Stud. Mycol. 64: 96 (2009)

Saprobic on submerged culm of Phragmites. Sexual morph: Ascomata scattered or in small groups, deeply immersed, lenticular or depressed ellipsoidal, with ostiolate papilla, stain substrate purple, visible as black, slit-like spot on the surface of substrate. Peridium comprising several layers of rounded to angular, brown to dark brown, thick-walled cells. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, embedded in a matrix. Asci 8-spored, bitunicate, fissitunicate, cylindro-clavate to broadly clavate, with an ocular chamber and short pedicel. Ascospores bi- to tri-seriate, narrowly to broadly fusoid, straight or curved, multi-septate, reddish brown, minutely verruculose, usually with a thin gelatinous sheath. Asexual morph: Undetermined.

Type species: Neomassariosphaeria typhicola Y. Zhang ter, J. Fourn. & K.D. Hyde

Notes: Neomassariosphaeria was established in Amniculicolaceae to accommodate Massariosphaeria typhicola (P. Karst.) Leuchtm (Zhang et al. 2009c). Later, it was transferred to Lindgomycetaceae based on phylogenetic analysis (Ariyawansa et al. 2015). However, we found that CBS 123126 is a problematic strain, which possesses two different LSU sequences and clusters in different families, i.e. FJ795504 (Zhang et al. 2009c) in Lindgomycetaceae and GU301844 (Schoch et al. 2009) in Amniculicolaceae. We substantiate GU301844 is the real sequence of Neomassariosphaeria typhicola (P. Karst.) Y. Zhang ter et al., while FJ795504, listed in Zhang et al. (2009c), is invalid (pers. comm. Zhang Y.). Ariyawansa et al. (2015) only included the invalid sequence FJ795504 in their phylognetic tree, which resulted in the classification of Neomassariosphaeria in Lindgomycetaceae.

Neomassariosphaeria often stains the woody substrate purple, and has reddish brown, multi-septate ascospores with an irregular, hyaline, gelatinous sheath, which morphologically fit well with the familial concept of Amniculicolaceae (Zhang et al. 2009c; Hyde et al. 2013). Based on morphology and our phylogenetic tree (Fig. 2), we follow Zhang et al. (2009c), to retain Neomassariosphaeria in Amniculicolaceae. Accession number FJ795504 bearing CBS123126 is treated as an invalid sequence.

Neomassariosphaeria typhicola (IFRD 2018) was considered as a synonym of Massariosphaeria typhicola (Zhang et al. 2009c). Although they are similar in the ascomata that staining the substrate purple, re-examination of IFRD 2018 (Fig. 24) shows that it is different from M. typhicola in having lenticular ascomata with an elongate papilla on the substrate, dark brown to black, reddish brown ascospores (Leuchtmann 1984). Phylogeny also placed them in different families, i.e. N. typhicola in Amniculicolaceae, M. typhicola in Lindgomycetaceae (Zhang et al. 2009c; this study, Fig. 2). We therefore, treat Neomassariosphaeria typhicola and Massariosphaeria typhicola as different species in this study.

Fig. 24
figure 24

Neomassariosphaeria typhicola (IFRD 2018). a Herbarium packet and specimen. b Ascomata immersed in host substrate. c Vertical section of ascoma. d Structure of peridium. e Pseudoparaphyses. f, g Mature asci. h Immature ascus. i An ocular chamber in mature ascus. j Verruculose ascospores. k Ascospore in Indian Ink. l Ascospore in ascus. m Ascospore with a thin, gelatinous sheath. n Mature ascospore. Scale bars: c = 100 μm, d, f–h = 20 μm, e = 3 μm, i–n = 10 μm

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List of freshwater Neomassariosphaeria species


*Neomassariosphaeria typhicola Y. Zhang ter, J. Fourn. & K.D. Hyde, Stud. Mycol. 64: 96 (2009)

Facesoffungi number: FoF09244; Fig. 24

Saprobic on submerged culm of Phragmites. Sexual morph: Ascomata 150–250 μm high, 300–500 μm diam., scattered or gregarious, immersed, lenticular or depressed ellipsoidal, with ostiolate papilla, stain the substrate purple, visible as a black, slightly protruding elongate papilla on the substrate. Peridium 20–35 μm thick, comprising several layers of rounded to angular, brown to dark brown, thick-walled cells. Pseudoparaphyses 1–2 μm wide, numerous, uneven in width or tuberculed, trabeculate, hyaline, septate, embedded in hyaline to reddish matrix. Asci 90–120 × 14–20 μm (\( {\bar{\text{x}}} \) = 97 × 16.5 μm, n = 10), 8-spored, bitunicate, fissitunicate, cylindro-clavate to broadly clavate, apically rounded, with a well-developed ocular chamber and short pedicel. Ascospores 30–40 × 6–8 μm (\( {\bar{\text{x}}} \) = 34.5 × 6.5 μm, n = 20), bi- to tri-seriate, narrowly to broadly fusoid, with subacute or rounded ends, straight or curved, 7-septate, constricted at the septa, dark brown to black or reddish brown, minutely verruculose, somewhat thick-walled, usually with a thin, broadly fusoid or irregular, gelatinous sheath. Asexual morph: Undetermined.

Material examined: DENMARK, Sjaeland, Frederikskilde, Suserup Skove, Tystrup Lake, on submerged culm of Phragmites, 25 May 2007, leg. & det. J. Fournier (IFRD 2018).

Notes: We re-examined the specimen Neomassariosphaeria typhicola IFRD 2018 collected by Zhang et al. (2009c) to confirm its generic placement in Amniculicolaceae. The sequence data (LSU: GU301844, SSU: GU296174, RPB2: GU371795) and morphological characters of N. typhicola support it as a distinct genus in Amniculicolaceae. The reddish brown, phragmoseptate ascospores of Neomassariosphaeria differs from hyaline, 1-septate ascospores of Amniculicola and golden yellow to brown, dictyoseptate ascospores of Murispora (see key to freshwater genera of Amniculicolaceae).

Vargamyces Tóth, Acta bot. hung. 25: 403 (1980) [1979]

Saprobic on submerged wood or leaves. Sexual morph: Undetermined. Asexual morph: Mycelium mostly immersed, composed of septate, hyaline to pale brown, smooth hyphae. Conidiophores micronematous to semi-macronematous, solitary, erect, straight to flexuous, cylindrical, subhyaline to pale brown, with percurrent proliferations. Conidiogenous cells integrated, terminal, cylindrical to cupuliforme, hyaline to pale brown. Conidia solitary, fusiform, truncate at the base, apically rounded, paler toward the ends, multi-septate, smooth-walled. Microconidia blastic growing on undifferentiated hyphae, solitary, terminal, lateral or intercalary, globose to obovoid, unicellular, hyaline, smooth (Hernández-Restrepo et al. 2017).

Type species: Vargamyces aquaticus (Dudka) Tóth

Notes: The monotypic genus Vargamyces was introduced by Tóth (1979) to accommodate V. aquaticus which was collected from submerged leaves and wood (Dudka 1966; Hyde and Goh 1999a). Vargamyces aquaticus was transferred to Xylomyces based on detachable, fusiform, dematiaceous and multiseptate chlamydospores (Hyde and Goh 1999a). However, Hernández-Restrepo et al. (2017) showed that V. aquaticus clustered in Amniculicolaceae, which is phylogenetically distant from the type species of Xylomyces, X. chlamydosporus (this study, Fig. 2). Therefore, Vargamyces is distinct and the only asexual genus in Amniculicolaceae, as listed in Wijayawardene et al. (2018, 2020). The lectotype (Fig. 1 in Dudka (1966)) and epitype (CBS H-22992, culture ex-epitype CBS 636.91) were also designated by Hernández-Restrepo et al. (2017).


List of freshwater Vargamyces species

*Vargamyces aquaticus (Dudka) Tóth, Acta Mus. Silesiae, Ser. A 25(3–4): 403 (1980) [1979]

Basionym: Camposporium aquaticum Dudka, Ukr. Bot. Zh. 23:91 (1966)

Synonymy: Sporidesmium ontariense Matsush., Matsush. Mycol. Mem. 3: 16 (1983)

Xylomyces aquaticus (Dudka) K.D. Hyde & Goh, Mycol. Res. 103(12): 1573 (1999)

Repetophragma ontariense (Matsush.) W.P. Wu, Fungal Diversity Res. Ser. 15: 82 (2005)

Freshwater distribution: England (Hyde and Goh 1999a), Spain (Hernández-Restrepo et al. 2017), Ukraine (Dudka 1966), USA (Tóth 1979)

Anteagloniaceae K.D. Hyde, J.K. Liu & A. Mapook, Fungal Diversity 63: 33 (2013)


Key to freshwater sexual genera of Anteagloniaceae

  1. 1.

    Ascomata orange-pigmented at the apex, asci clavate to cylindric-clavate, ascospores hyaline, fusiform………Flammeascoma

  2. 2.

    Ascomata stain the substrate purple, asci cylindrical, ascospores olivaceous, ellipsoidal………Purpureofaciens


Flammeascoma Phook. & K.D. Hyde, Fungal Diversity 72(1): 63 (2015)

Saprobic on submerged bamboo or dead wood. Sexual morph: Ascostromata solitary to gregarious, erumpent to superficial, visible as dull, black, dark at the basal region and orange brown at the apex, conical to lenticular, with a flattened base, uni- to bi-loculate, glabrous, coriaceous, ostiolate, with pore-like opening. Peridium thick-walled, of unequal thickness, poorly developed at the base, composed of several layers of dark, pseudoparenchymatous cells, with fungal tissue intermixed with the host cells. Pseudoparaphyses dense, trabeculate, filiform, hyaline, septate. Asci 8-spored, bitunicate, fissitunicate, clavate to cylindric-clavate, short pedicellate, apically rounded with well-developed ocular chamber. Ascospores uni- to bi-seriate, fusiform, hyaline, becoming brown when released from asci, septate, smooth-walled, with or without mucilaginous sheath (Liu et al. 2015). Asexual morph: Undetermined.

Type species: Flammeascoma bambusae Phook. & K.D. Hyde

Notes: Flammeascoma was introduced to accommodate F. bambusae, which was collected from submerged bamboo (Liu et al. 2015). Flammeascoma is similar to familial type Anteaglonium Mugambi & Huhndorf in the shape of asci and ascospores (Mugambi and Huhndorf 2009, Liu et al. 2015). However, Flammeascoma is unique in producing reddish orange pigmentation at the apex of ascostromata, while Anteaglonium lacks this character and always sits on a thin darkened crust with or without sparse, dark brown, septate subiculum (Mugambi and Huhndorf 2009; Ariyawansa et al. 2015; Liu et al. 2015).

Flammeascoma appears to be polyphyletic as F. bambusae (type) clusters with Anteaglonium gordoniae Jayasiri et al. and F. lignicola Boonmee & K.D. Hyde clusters with our new genus Purpureofaciens, but without bootstrap support (Fig. 2). More species are expected to clarify whether Flammeascoma is polyphyletic.


List of freshwater Flammeascoma species

*Flammeascoma bambusae Phookamsak & K.D. Hyde, Fungal Diversity 72(1): 64 (2015); Fig. 25

Fig. 25
figure 25

Flammeascoma bambusae (Material examined: THAILAND, Chiang Mai Province, Chom Tong District, Doi Inthanon, on dead stem of submerged bamboo (Bambusae), 5 September 2009, R. Phookamsak, RP0013, MFLU 11-0143, holotype). a Appearance of ascostromata on the host surface. b Section through an ascostroma. c Peridium. d–f Bitunicate asci. g Pseudoparaphyses. h–m Ascospores. Scale bars: b = 200 μm, c–f = 50 μm, g–m = 20 μm

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Freshwater distribution: Thailand (Liu et al. 2015)


Purpureofaciens W. Dong, H. Zhang & K.D. Hyde, gen. nov.

Index Fungorum number: IF557804; Facesoffungi number: FoF08717

Etymology: referring to staining the substrate purple

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata black, scattered or gregarious, semi-immersed to superficial, conical, flattened at the basal region, stain the substrate purple and produce reddish pigmentation at the apex, coriaceous, with a large, protuberant, ostiolate papilla. Peridium of unequal thickness, poorly developed at the base, composed of reddish brown, large cells of textura angularis. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, embedded in a hyaline gelatinous matrix. Asci 8-spored, bitunicate, cylindrical, pedicellate, apically rounded. Ascospores uniseriate, curved, olivaceous, septate, ellipsoidal with acute ends, guttulate, thin-walled, smooth, surrounded by a thin mucilaginous sheath. Asexual morph: Undetermined.

Type species: Purpureofaciens aquatica W. Dong, H. Zhang & K.D. Hyde

Notes: Purpureofaciens is unique by semi-immersed to superficial, conical ascomata staining woody substrate purple, cylindrical asci, ellipsoidal, curved, olivaceous ascospores with acute ends and white to reddish colonies in culture (Mugambi and Huhndorf 2009; Liu et al. 2015). The reddish culture is also observed in a strain of Anteaglonium parvulum (MFLUCC 14-0821), which has similar shape of ascospores, but the characters of ascomata, asci and ascospores can easily separate them (Jayasiri et al. 2016). Flammeascoma has reddish pigmentation at the apex of ascomata, which resembles Purpureofaciens. However, the conical ascomata with a large, protuberant, ostiolate papilla, cylindrical asci and ellipsoidal, olivaceous ascospores of Purpureofaciens are different from those of Flammeascoma (Liu et al. 2015). Purpureofaciens clusters with Flammeascoma species with low bootstrap support (Fig. 2) and morphology separates them as distinct genera. Purpureofaciens comprises one freshwater species P. aquatica which was collected from Thailand.


List of freshwater Purpureofaciens species

*Purpureofaciens aquatica W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557902; Facesoffungi number: FoF08718; Fig. 26

Fig. 26
figure 26

Purpureofaciens aquatica (MFLU 18-1569, holotype). a–c Appearance of black ascomata on host surface. d, e Vertical section of ascomata. f Structure of peridium. g Pseudoparaphyses. h, i Bitunicate asci. j–l Ascospores. m Germinated ascospores. n, o Colony on PDA (left-front, right-reverse). Scale bars: d, e = 100 μm, f–i, m = 20 μm, j–l = 10 μm

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Etymology: in reference to aquatic habitat of the fungus

Holotype: MFLU 18-1569

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 200–300 μm high, 300–400 μm diam., dark brown to black, scattered or gregarious, semi-immersed to superficial, conical, flattened at the basal region, stain the substrate purple and produce reddish pigmentation at the apex, coriaceous, with a large, protuberant, ostiolate papilla, 110–130 μm long, 110–130 μm wide. Peridium unequal thickness, poorly developed at the base, up to 45 μm at the sides and 15 μm at the base, composed of reddish brown, large cells of textura angularis. Pseudoparaphyses 2 μm diam., numerous, trabeculate, filiform, hyaline, embedded in a hyaline gelatinous matrix. Asci 135–150 × 8.5–11 μm (\( {\bar{\text{x}}} \) = 143 × 9.7 μm, n = 10), 8-spored, bitunicate, cylindrical, pedicellate, apically rounded. Ascospores 15–22 × 7–10 μm (\( {\bar{\text{x}}} \) = 18.5 × 8 μm, n = 20), uniseriate, curved, olivaceous, 1-septate, constricted at the septum, ellipsoidal with acute ends, with two prominent guttules when immature, thin-walled, smooth, surrounded by a thin mucilaginous sheath. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 50 mm diam. in 25 days at 25 °C, annulate, white to reddish to purplish red from center to edge, surface rough, dry, raised, edge entire.

Material examined: THAILAND, Satun Province, on submerged wood in a stream, 10 May 2018, W. Dong, hat266-1 (MFLU 18-1569, holotype), ex-type living culture MFLUCC 18-1241; ibid. hat266-2 (HKAS 105016, isotype), ex-type living culture KUMCC 19-0032.


Aquasubmersaceae A. Hashim. & Kaz. Tanaka, Persoonia 39: 56 (2017)


Aquasubmersa K.D. Hyde & H. Zhang, Cryptog. Mycol. 33(3): 340 (2012)

Saprobic on submerged or dead wood. Sexual morph: Ascomata scattered to grouped, immersed to semi-immersed, uniloculate, subglobose, glabrous, dark brown to black, with ostiolate papilla. Peridium composed of flattened, thin-walled cells of textura angularis. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, septate. Asci 8-spored, bitunicate, cylindrical, short pedicellate, apically rounded, with an ocular chamber. Ascospores biseriate, broadly fusiform with rounded ends, straight, hyaline, with one mostly median septum, with gelatinous sheath (Ariyawansa et al. 2015). Asexual morph: Coelomycetous. Pycnidia solitary to scattered, semi-immersed to superficial, subglobose to ellipsoidal, dark brown to black, ostiolate, minute or without papilla. Ostiole circular, dark-brown, central. Peridium composed of polygonal, hyaline to brown cells. Conidiophores reduced. Conidiogenous cells holoblastic, determinate, lageniform, hyaline, smooth. Conidia ellipsoidal, some with papillate base, hyaline, aseptate, smooth, thin-walled, without sheath or appendage (Zhang et al. 2012a).

Type species: Aquasubmersa mircensis Huang Zhang & K.D. Hyde

Notes: Aquasubmersa was established to accommodate a freshwater coelomycetous species, A. mircensis (Zhang et al. 2012a). It was placed in the family Lophiotremataceae in previous studies (Ariyawansa et al. 2015; Doilom et al. 2017). A comprehensive study on Lophiotremataceae based on morphological observations and multigene analyses showed that Aquasubmersa should be transferred to a newly established family Aquasubmersaceae (Hashimoto et al. 2017). The second species, A. japonica A. Hashim. & Kaz. Tanaka with sexual morph, was also collected from freshwater habitats and conidiomata were produced in culture (Ariyawansa et al. 2015). The asexual morph of A. japonica matches the generic concept of Aquasubmersa. It differs from A. mircensis by pycnidia with minute papilla, thicker peridium and conidia lacking papillate bases. The type material of A. mircensis was re-examined by Li et al. (2020) and this study.


List of freshwater Aquasubmersa species

*Aquasubmersa japonica A. Hashim. & Kaz. Tanaka, Fungal Diversity 75: 87 (2015)

Freshwater distribution: Japan (Ariyawansa et al. 2015)


*Aquasubmersa mircensis H. Zhang & K.D. Hyde, Cryptog. Mycol. 33(3): 340 (2012); Fig. 27

Fig. 27
figure 27

Aquasubmersa mircensis (Material examined: THAILAND, Chiang Mai Province, Mushroom Research Centre, on submerged wood, 21 April 2011, H. Zhang, m3, MFLU 11-1001, holotype). a Pycnidium on wood surface. b Section of pycnidium. c Peridium with conidiogenous cells. d Conidiogenous cells with conidia. e, f Conidia. Scale bars: b = 100 µm, c = 20 µm, d = 15 µm, e, f = 10 µm

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Freshwater distribution: Thailand (Zhang et al. 2012a)


Key to freshwater Aquasubmersa species

1. Pycnidia up to 215 μm high, 115–195 μm diam., with minute papilla, peridium > 20 µm thick, conidia without papillate base………A. japonica


1. Pycnidia 130–170 μm high, 150–250 μm diam., without papilla, peridium < 20 μm thick, conidia with papillate base………A. mircensis


Astrosphaeriellaceae Phook. & K.D. Hyde, Fungal Diversity 74: 161 (2015)


Key to freshwater genera of Astrosphaeriellaceae

  1. 1.

    Asci mostly broadly clavate………2

  2. 1.

    Asci cylindrical………3

  3. 2.

    Ascospores hyaline, cylindrical………Aquatospora

  4. 2.

    Ascospores mostly dark brown to black, fusiform to broad-ellipsoid or biconic………Caryospora

  5. 3.

    Producing hyphomycetous asexual morph………Pithomyces

  6. 3.

    Producing coelomycetous asexual morph or unknown………4

  7. 4.

    Ascospores brown to reddish brown, with paler end cells………Xenoastrosphaeriella

  8. 4.

    Ascospores hyaline to brown, without paler end cells………Astrosphaeriella


Aquatospora W. Dong, H. Zhang & K.D. Hyde, gen. nov.

Index Fungorum number: IF557903; Facesoffungi number: FoF09245

Etymology: referring to aquatic habitat of this fungus

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata black, scattered or gregarious, superficial, hemisphaerical to conical, base flattened, carbonaceous, with ostiolate papilla. Ostiole central, relatively broad, circular, black. Peridium uneven in width, carbonized, composed of black amorphous layer, whose cells are obscured. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, sparsely septate. Asci 8-spored, bitunicate, clavate to narrowly ellipsoidal, with a short pedicel, apically rounded. Ascospores 2–4-seriate, hyaline, septate, subcylindrical, thin-walled, smooth. Asexual morph: Undetermined.

Type species: Aquatospora cylindrica W. Dong, H. Zhang & K.D. Hyde

Notes: Aquatospora clusters with Acrocordiopsis Borse & K.D. Hyde and Xenoastrosphaeriella Jayasiri et al. with low bootstrap support (Fig. 2). Acrocordiopsis is a problematic genus, which was placed in Salsugineaceae by Hyde et al. (2013) and Caryosporaceae by Ariyawansa et al. (2015). Acrocordiopsis was referred to Salsugineaceae (Wijayawardene et al. 2018, 2020) based on some common morphology between Acrocordiopsis and Salsuginea K.D. Hyde (Borse and Hyde 1989; Hyde 1991). Acrocordiopsis clustered distantly from Salsuginea in the phylogenetic tree of Zhang et al. (2018) and close to Aquatospora in our study (Fig. 2). Xenoastrosphaeriella differs from Aquatospora in having paler end cells of ascospores (Hawksworth and Boise 1985; Phookamsak et al. 2015b; Jayasiri et al. 2019). Aquatospora shares similar characters with Astrosphaeriellaceae members in having large, erumpent to superficial, hemisphaerical or conical, carbonaceous ascomata with nearly flattened bases (Borse and Hyde 1989; Ariyawansa et al. 2015; Phookamsak et al. 2015b). However, the unique combination of morphology of clavate to narrowly ellipsoidal asci, hyaline, cylindrical ascospores and molecular characters warrant Aquatospora as a new genus (more details see key to freshwater genera of Astrosphaeriellaceae).


List of freshwater Aquatospora species

*Aquatospora cylindrica W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557904; Facesoffungi number: FoF09246; Fig. 28

Fig. 28
figure 28

Aquatospora cylindrica (MFLU 18-1534, holotype). a Appearance of black ascomata on host. b, c Vertical section of ascoma. d Structure of peridium. e Pseudoparaphyses. f–h Bitunicate asci. i–k Ascospores. k Ascospore in Indian Ink. l Germinated ascospore. m, n Colony on PDA (left-front, right-reverse). Scale bars: c = 200 μm, d, f–l = 50 μm, e = 20 μm

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Etymology: referring to cylindrical ascospores of this fungus

Holotype: MFLU 18-1534

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 600–650 μm high, 800–900 μm diam., black, scattered or gregarious, superficial, hemisphaerical to conical, base flattened, carbonaceous, with ostiolate papilla. Ostiole central, relatively broad, circular, black. Peridium up to 155 μm at the sides and 65 μm at the base, carbonized, composed of black amorphous layer, whose cells are obscured. Pseudoparaphyses 0.5 μm diam., numerous, trabeculate, filiform, hyaline, sparsely septate. Asci 175–270 × 50–65 μm (\( {\bar{\text{x}}} \) = 225 × 58 μm, n = 10), 8-spored, bitunicate, clavate to narrowly ellipsoidal, straight or slightly curved, with a short pedicel, apically rounded. Ascospores 70–85 × 20–25 μm (\( {\bar{\text{x}}} \) = 77 × 23 μm, n = 10), 2–4-seriate, cylindrical, slightly tapering at two sides, 1-septate, constricted at the septum, hyaline, straight, with four prominent big guttules, thin-walled, smooth, without sheath. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 40 mm in 35 days at 25 °C, brown to white from above, dark brown from below, surface rough, dry, raised, edge entire.

Material examined: THAILAND, Songkhla Province, on submerged wood in a stream, 10 May 2018, W. Dong, hat146-1 (MFLU 18-1534, holotype), ex-type living culture MFLUCC 18-1287; ibid., hat146-2 (HKAS 105042, isotype), ex-type living culture KUMCC 19-0060.


Astrosphaeriella Syd. & P. Syd., Annls mycol. 11(3): 260 (1913)

Saprobic or parasitic on bamboo, palm or stout grasses in freshwater or terrestrial habitats. Sexual morph: Ascostromata solitary to gregarious, erumpent to superficial, conical or mammiform, at maturity with stellate appearance from above, dark opaque, uniloculate, glabrous, brittle, carbonaceous, ostiolate. Peridium relatively thick, poorly developed at the base, composed of thick, opaque and melanized cells, arranged in a textura angularis with palisade-like cells at the rim. Pseudoparaphyses dense, trabeculate, filiform, hyaline, anastomosing, embedded in a hyaline gelatinous matrix. Asci 8-spored, bitunicate, fissitunicate, cylindrical to cylindric-clavate, subsessile to short-pedicellate, apically rounded with ocular chamber. Ascospores overlapping uni- to bi-seriate, hyaline to brown, fusiform with rounded to acute ends, septate, smooth, with or without appendages or mucilaginous sheaths (Phookamsak et al. 2015b). Asexual morph: Coelomycetous. Conidiomata pycnidial, scattered or solitary, immersed to superficial, conical or hemisphaerical to globose. Pycnidial walls composed of several layers of dark brown to black, textura angularis to textura intricata of cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic or phialidic, integrated, cylindrical or cylindric-clavate or ampulliform, septate or aseptate, hyaline, smooth. Conidia globose to subglobose, or oblong, aseptate, hyaline, smooth (Phookamsak et al. 2015b).

Type species: Astrosphaeriella fusispora Syd. & P. Syd.

Notes: Astrosphaeriella is characterized by carbonaceous, conical to mammiform ascostromata with ruptured, reflexed, stellate, host remnants and hyaline to brown, fusiform ascospores (Liu et al. 2011; Phookamsak et al. 2015b). Astrosphaeriella aquatica K.D. Hyde, A. seychellensis K.D. Hyde & Goh and A. thailandensis J. Ren et al. were only found from freshwater habitats (Hyde 1994; Hyde and Frohlich 1998; Hyde and Goh 1998b; Ren et al. 2013). Other freshwater Astrosphaeriella species were found from both freshwater and terrestrial habitats (see list below). Astrosphaeriella aquatica resembles A. seychellensis and A. thailandensis in having hyaline, fusiform ascospores, but differs in having a wide spreading mucilaginous sheath which is drawn out at the poles.

Phookamsak et al. (2015b) transferred Astrosphaeriella sensu lato, which have coriaceous ascomata with short to long neck and striate ascospores, to a newly established family Pseudoastrosphaeriellaceae. Astrosphaeriella papuana Aptroot has striate ascospores which are typical characters of Pseudoastrosphaeriellaceae (Aptroot 1995; Hyde and Frohlich 1998; Phookamsak et al. 2015b). However, molecular data for A. papuana is unavailable, thus this species is retained in Astrosphaeriellaceae. Astrosphaeriella papuana might be transferred to Pseudoastrosphaeriellaceae when the type specimen is recollected and recircumscribed.

Astrosphaeriella stellata (Pat.) Sacc. was treated as an earlier name of A. fusispora and followed by some studies (Hyde and Frohlich 1998; Hyde et al. 2000; Tanaka et al. 2009; Liu et al. 2011). Liu et al. (2011) named one collection MFLUCC 10-0555 as A. stellata, which was later considered as a reference specimen of A. fusispora (Phookamsak et al. 2015b). Therefore, A. stellata isolated from freshwater habitats is probably a collection of A. fusispora.

The asexual morph was developed in the culture of Astrosphaeriella bambusae Phookamsak& K.D. Hyde (Phookamsak et al. 2015b).


List of freshwater Astrosphaeriella species


Astrosphaeriella aquatica K.D. Hyde, Mycol. Res. 98(7): 719 (1994)

Freshwater distribution: Papua New Guinea (Hyde 1994), Ecuador (Hyde and Frohlich 1998)


Astrosphaeriella exorrhiza Boise, Sydowia 38: 117 (1986) [1985]

Freshwater distribution: Ecuador (Hyde and Frohlich 1998)


Astrosphaeriella maquilingiana (Rehm) K.D. Hyde & J. Fröhl., Sydowia 50(1): 103 (1998)

Basionym: Trematosphaeria maquilingiana Rehm, Leafl. of Philipp. Bot. 8: 2952 (1916)

Synonymy: Trematosphaeria maquilingiana var. schizostachyi Rehm, Leafl. of Philipp. Bot. 8: 2952 (1916)

Trematosphaeria maquilingiana Rehm, Leafl. of Philipp. Bot. 8: 2952 (1916) var. maquilingiana

Freshwater distribution: Australia (Vijaykrishna and Hyde 2006)


Astrosphaeriella papuana Aptroot, Nova Hedwigia 60(3–4): 333 (1995)

Freshwater distribution: Papua New Guinea (Hyde and Frohlich 1998), Philippines (Cai et al. 2003a)


Astrosphaeriella seychellensis K.D. Hyde & Goh, S. Afr. J. Bot. 64: 332 (1998)

Freshwater distribution: Seychelles (Hyde and Goh 1998b)


*Astrosphaeriella stellata (Pat.) Sacc., Syll. fung. (Abellini) 24(2): 938 (1928); Fig. 29

Fig. 29
figure 29

Astrosphaeriella stellata (Material examined: VIETNAM, Hanoi, on Bamboo (sur Bambou), 17 April 1911, M. Duport 451, FH 00290284, Pat 552, holotype). a Herbarium label and specimen. b Ascostromata on host surface. c Section through ascostroma. d Section through peridium. e Pseudoparaphyses. f–h Bitunicate asci. i Ocular chamber. j–p Ascospores. Scale bars: d, e, i–p = 20 μm, f–h = 50 μm

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Basionym: Amphisphaeria stellata Pat., Bull. Soc. mycol. Fr. 29: 223 (1913)

Synonymy: Astrosphaeriella stellata var. palmicola F. San Martín & P. Lavín, Lavin & Lavín, Acta Bot. Mexicana 46: 22 (1999)

Astrosphaeriella bambusella Höhn., Sber. Akad. Wiss. Wien, Math.-naturw. Kl., Abt. 1 129: 168 (1920)

Astrosphaeriella fuscomaculans W. Yamam., Sci. Rep. Hyogo Univ. Agric., Ser. 2, Agr. Biol. 1: 63 (1954)

Microthelia fuscomaculans (W. Yamam.) E. Müll., Beitr. Kryptfl. Schweiz 11(no. 2): 286 (1962)

Freshwater distribution: China (Tsui et al. 2001d; Wong and Hyde 2001; Luo et al. 2004; Hu et al. 2013), Philippines (Cai et al. 2003a)


Astrosphaeriella thailandensis J. Ren, C.Y. Jie, Y.L. Jiang, K.D. Hyde & Yong Wang bis, Sydowia 65(1): 33–43 (2013)

Freshwater distribution: Thailand (Ren et al. 2013)


Key to freshwater Astrosphaeriella species

  1. 1.

    Ascospores surrounded by sheath or with appendage………2

  2. 1.

    Ascospores without sheath or appendage………6

  3. 2.

    Ascospores sometimes slightly verrucose………A. maquilingiana

  4. 2.

    Ascospores smooth-walled………3

  5. 3.

    Ascospores with minute appendages at both ends………A. exorrhiza

  6. 3.

    Ascospores surrounded by a mucilaginous sheath………4

  7. 4.

    Ascospores pigmented………A. stellata

  8. 4.

    Ascospores hyaline………5

  9. 5.

    Ascospores 30–42 × 7–8 μm, sheath drawn out at the poles………A. aquatica

  10. 5.

    Ascospores 16–22 × 4–4.5 μm, sheath appears as remnants in drided material………A. seychellensis

  11. 6.

    Ascospores with striate ornamentation………A. papuana

  12. 6.

    Ascospores smooth-walled………A. thailandensis


Caryospora De Not., Micr. Ital. Novi 9: 7 (1855)

Saprobic on submerged wood, decaying fruit pericarp or old stone of Amygdalus persica. Sexual morph: Ascomata pseudothecioid, medium to large, scattered, erumpent to superficial, uniloculate, broadly conical or hemispherical, with flattened base, brown to black, carbonaceous, with ostiolate papilla. Ostiole relatively broad, central, circular. Peridium thicker at sides and thinner at base, strongly carbonized. Pseudoparaphyses numerous, trabeculate, filiform, hyaline, sparsely septate. Asci 2(8)-spored, bitunicate, fissitunicate, broadly cylindrical to clavate or obclavate, pedicellate. Ascospores large, bi- to tri-seriate, fusiform to broad-ellipsoid or biconic, straight, usually hyaline when young, becoming pigmented at maturity, septate, most symmetrical, smooth or rough, often with polar germ pores at each end, thin- to thick-walled, with or without sheath. Asexual morph: Undetermined.

Type species: Caryospora putaminum (Schwein.) Fuckel

Notes: Caryospora was placed in Caryosporaceae based on sequence data of C. aquatica and one putative strain of C. minima (Ariyawansa et al. 2015). Caryospora clustered with Acrocordiopsis with weak bootstrap support (Ariyawansa et al. 2015). Jayasiri et al. (2019) placed Caryospora in Astrosphaeriellaceae based on the similarity of carbonaceous ascostromata and trabeculate pseudoparaphyses with other genera in Astrosphaeriellaceae. However, Astrosphaeriellaceae members, including Caryospora, did not form a well-supported clade (Jayasiri et al. 2019; this study, Fig. 2). Zopfia rhizophila Rabenh. (Zopfiaceae) also clustered in Astrosphaeriellaceae, but with low bootstrap support (Jayasiri et al. 2019; this study, Fig. 2). In this study, we follow Jayasiri et al. (2019) and accept Caryospora in Astrosphaeriellaceae based on morphology.

Caryospora obclavata was reported from freshwater habitat and characterized by obclavate asci with ellipsoidal to biconic ascospores uniseriately arranged at the apex and base (Raja and Shearer 2008). The taxonomic placement of this species needs to be confirmed with molecular data. We introduce one new species C. submersa based on morphology and phylogeny. Additionally, C. aquatica and C. quercus are redescribed and reported as new geographical record in Thailand and new habitat record in freshwater, respectively.


List of freshwater Caryospora species

*Caryospora aquatica H. Zhang, K.D. Hyde & Ariyawansa, Fungal Diversity 75: 54 (2015); Figs. 30, 31

Fig. 30
figure 30

Caryospora aquatica (MFLU 18-1202, new geographical record). a–c Ascomata on host surface. d, f Vertical section of ascomata. e Structure of peridium. g Pseudoparaphyses. h–k Bitunicate asci. l–q Immature and mature ascospores. r, s Ascospores in Indian Ink. t, u Colony on PDA (up-front, down-reverse). Scale bars: d = 100 μm, e–g, l–s = 20 μm, h–k = 40 μm

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Fig. 31
figure 31

Caryospora aquatica (Material examined: THAILAND, Chiang Rai Province, on submerged wood, 18 January 2010, H. Zhang, MFLU 11-1083, holotype). a Ascomata on the host surface. b Section of an ascoma on wood. c Section of ascoma. d, e Different stages of asci. f Ascospores. Scale bars: a = 700 µm, b, c = 350 µm, d, e = 60 µm, f = 50 µm

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Freshwater Distribution: China (this study), Thailand (Ariyawansa et al. 2015)

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 270–310 μm high, 250–330 μm diam., black, scattered or gregarious, superficial, hemisphaerical, flattened at the base, carbonaceous, ostiolate, always covered with mass of black, glistening ascospores. Peridium 50–60 μm at the sides, poorly developed at the base, easily cracked, two-layered, outer layer strongly carbonized, amorphous, composed of several layers of black cells that cannot be differentiated and often occluded, inner layer composed of several layers of hyaline cells of textura angularis. Pseudoparaphyses 2 μm diam., numerous, trabeculate, filiform, hyaline, septate. Asci 160–210(–270) × 45–58 μm (\( {\bar{\text{x}}} \) = 190 × 52 μm, n = 10), 8-spored, bitunicate, narrowly to broadly clavate, with short pedicel, apically rounded. Ascospores 35–48 × 18–28 μm (\( {\bar{\text{x}}} \) = 42 × 22 μm, n = 40), biseriate, straight, broadly fusiform, hyaline to pale brown, 1-septate when young, becoming irregularly diamond-shaped, dark brown to black, 3-septate when mature, with rounded ends, constricted and darker at the median septum, not constricted and inconspicuous at two terminal septa, asymmetric, with two large central cells and two small end cells, guttulate, thick-walled, smooth, sometimes with polar germ pores at each end, surrounded by an irregular, mucilaginous sheath. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 15 mm diam. in 50 days at 25 °C, grey from above, black from below, surface rough, dry, with dense mycelium, raised, edge entire.

Material examined: CHINA, Yunnan Province, on submerged wood in a stream, 25 November 2017, G.N. Wang, H3A-1 (MFLU 18-1202), living culture MFLUCC 18-1030; ibid., H3A-2 (HKAS 101729), living culture KUMCC 18-0079.

Notes: Our new collection MFLUCC 18-1030 clusters with Caryospora aquatica H. Zhang et al. (MFLUCC 11-0008) and one putative strain of C. minima Jeffers with high bootstrap support (Fig. 2). MFLUCC 18-1030 has identical LSU sequence data with C. aquatica MFLUCC 11-0008 and seven (including three gaps) nucleotide differences in ITS sequence data between the two isolates, which indicate them to be the same species. Morphologically, our collection has very similar morphological characters with C. aquatica (Figs. 30, 31), except the slightly thinner asci (45–58 μm wide vs. 60–80 μm wide) and irregular sheaths (Ariyawansa et al. 2015). Given this, we identify our collection MFLUCC 18-1030 as C. aquatica. This is a new geographical record of C. aquatica in China.

Caryospora minima was initially collected from putrescent putamina of Amygdalus persica in Maryland, USA (Jeffers 1940). Later, several freshwater collections of C. minima were reported in China, Philippines and South Africa (see freshwater distribution of this species), but they were not confirmed by molecular data. Cai and Hyde (2007b) provided LSU (EU196550) and SSU (EU196551) sequence data for C. minima without strain code, description, illustration and herbarium information. Thus, these two sequences are doubtful. A specimen (IFRD 083-010) under name C. minima is illustrated in this study (Fig. 32). However, this species should be treated with caution based on the holotype BPI 71132. It would be good to establish an epitype for C. minima with reliable sequence data.

Fig. 32
figure 32

Caryospora minima (Material examined: CHINA, Hong Kong, on submerged wood in Lam Tsuen River, 1 December 1996, C. Tsui, IFRD 083-010). a Herbarium label and specimen of Caryospora minima. b Ascomata on host surface. c, d Vertical section through ascoma. e Structure of peridium. f Pseudoparaphyses. g, h Bitunicate asci. i Ocular chamber. j Ascospore. k Ascospore in Indian Ink. Scale bars: c = 100 μm, e, g, h = 20 μm, f, j–k = 10 μm, i = 5 μm

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*Caryospora minima Jeffers, Mycologia 32(4): 561 (1940); Fig. 32

Freshwater Distribution: China (Tsui et al. 2000; Luo et al. 2004), Philippines (Cai et al. 2003a), South Africa (Hyde et al. 1998)


Caryospora obclavata Raja & Shearer, Mycologia 100(3): 479 (2008)

Freshwater Distribution: USA (Raja and Shearer 2008)


*Caryospora quercus Jayasiri, E.B.G. Jones & K.D. Hyde, Mycosphere 10(1): 34 (2019); Fig. 33

Fig. 33
figure 33

Caryospora quercus (MFLU 17-1705, new habitat record). a, b Ascomata on host surface. c Vertical section of ascoma. d Structure of peridium. e–g, i–k Bitunicate asci. h Pseudoparaphyses. l Immature ascospore, with two-layered sheath (arrows). m–o Mature ascospores. p Germinated ascospore. q Ascospore in Indian Ink. r, s Colony on PDA (left-front, right-reverse). Scale bars: c = 100 μm, d, h, m–o = 20 μm, e–g, i–l, p, q = 50 μm

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Freshwater Distribution: Thailand (this study)

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 400–450 μm high, 410–460 μm diam., black, scattered, semi-immersed to superficial, conical, flattened at the base, carbonaceous, ostiolate, 70 μm diam., often covered with mass of black, glistening ascospores. Peridium 60–70 μm at the sides, poorly developed at the base, easily cracked, two-layered, outer layer strongly carbonized, composed of several layers of black cells that cannot be differentiated and often occluded, inner layer composed of 8–10 layers of hyaline, compressed cells of textura angularis. Pseudoparaphyses 2 μm diam., numerous, trabeculate, filiform, hyaline, distantly septate. Asci (165–)205–270(–305) × (58–)65–80 μm (\( {\bar{\text{x}}} \) = 235 × 70 μm, n = 10), 8-spored, bitunicate, narrowly to broadly clavate, indistinctly pedicellate which become tapered and disappeared with age, 30–60 μm long, apically rounded. Ascospores 47–60 × 19–33 μm (\( {\bar{\text{x}}} \) = 54 × 27 μm, n = 30), overlapping bi- to tri-seriate, straight, broadly fusiform to ellipsoidal, hyaline to pale brown, 1-septate when young, becoming irregularly diamond-shaped, dark brown to black, 3-septate when mature, with acute ends, constricted and darker at the median septum, not constricted and inconspicuous at two terminal septa, asymmetric, with two large central cells and two small end cells, thick-walled, smooth, sometimes with polar germ pores at each end, surrounded by a large, ellipsoidal, gelatinous, mucilaginous sheath, 3–37 μm thick, sheath two-layered and large when young, becoming one-layered and smaller after mature. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular, reaching 5 mm diam. in 20 days at 25 °C, grey from above and brown from below, surface rough, dry, raised, edge entire.

Material examined: THAILAND, Chiang Mai Province, on submerged wood in a stream, 1 September 2017, X.D. Yu, 1A (MFLU 17-1705, holotype), ex-type living culture MFLUCC 17-2342; ibid., G.N. Wang, 4.2 (MFLU 17-1668), living culture MFLUCC 17-2323.

Notes: Our new collections MFLUCC 17-2342 and MFLUCC 17-2323 cluster with Caryospora quercus with strong bootstrap support (Fig. 2). They have identical LSU sequence data and there are two nucleotide differences in SSU sequence data between MFLUCC 17-2342 and MFLU 18-2151 (type). No other genes can be compared.

Our new collections have overlapping ascospore size (47–60 × 19–33 μm vs. 41–54 × 18–28 μm) and similar morphological characters with C. quercus, i.e. broadly fusiform to ellipsoidal and hyaline to pale brown when young, becoming irregularly diamond-shaped and dark brown to black when mature, with acute ends, with one primary median septum and two inconspicuous septa near both ends, asymmetric, and with a sheath (Jayasiri et al. 2019). However, the ascospore sheaths in our collections are up to 37 μm thick, while they are 5.5 μm thick in the holotype of C. quercus. This might be a result of only measuring the mature ascospores in Jayasiri et al. (2019), because they are 37 μm thick in young ascospores and 3 μm thick in mature ascospores in our collections. Additionally, our collections have longer and wider asci ((165–)205–270(–305) × (58–)65–80 μm vs. (69–)110–147 × 30–35 μm) than the holotype of C. quercus. It is very likely that Jayasiri et al. (2019) only measured the immature asci as shown in their photo plate. We identify our collections to be C. quercus based on current phylogenetic analysis (Fig. 2), single gene comparison and morphology. We supplement ITS sequence data for C. quercus. This is a new habitat record for C. quercus from freshwater in Thailand.


*Caryospora submersa W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557905; Facesoffungi number: FoF09248; Figs. 34, 35

Fig. 34
figure 34

Caryospora submersa (MFLU 18-1539, holotype). a, b Ascomata on host surface. c Vertical section of ascoma. d Structure of peridium. e–g Bitunicate asci. h Asci embedded in pseudoparaphyses. i–k Ascospores. l Ascospore from fresh specimen mounted in Indian Ink. m Germinated ascospore. n Colony on PDA (up-front, down-reverse). Scale bars: c = 100 μm, d, l, m = 50 μm, e–k = 20 μm

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Fig. 35
figure 35

Caryospora submersa (MFLU 18-1541). a, b Ascomata on host surface. c Vertical section of ascoma. d Vertical section of ostiolate papilla. e Pseudoparaphyses. f–h Bitunicate asci. i, j Ascospores. k, l Ascospores from dried specimen mounted in Indian Ink. m Colony on PDA (left-front, right-reverse). Scale bars: c = 100 μm, d = 50 μm, e–l = 20 μm

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Etymology: in reference to submerged habitat of the fungus

Holotype: MFLU 18-1539

Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata 400–450 μm high, 450–540 μm diam., black, scattered or gregarious, superficial, conical, flattened at the base, carbonaceous, with ostiolate papilla. Peridium 45–85 μm at the sides, poorly developed at the base, easily cracked, strongly carbonized, composed of several black, amorphous layers of cells that cannot be differentiated and often occluded. Pseudoparaphyses 2 μm diam., numerous, trabeculate, filiform, hyaline, sparsely septate. Asci 130–180 × 17–26 μm (\( {\bar{\text{x}}} \) = 160 × 21 μm, n = 20), 8-spored, bitunicate, narrowly obclavate, with rounded apex, short pedicellate, with a well-developed ocular chamber. Ascospores 38–50 × 8–12 μm (\( {\bar{\text{x}}} \) = 44 × 9 μm, n = 20), mostly uniseriate near the apex, becoming biseriate towards the base, straight or slightly curved, hyaline, 1-septate, constricted at the septum, narrowly fusiform, guttulate, thin-walled, smooth, surrounded by a large, oblong or ellipsoidal, mucilaginous sheath, 40–80 μm wide, sheath becoming thin and dumbbell-like in dried specimen, 1.5–2.5 μm wide at the sides and 5–15 μm wide at two ends. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony circular or ellipsoidal, reaching 10 mm diam. in 10 days at 25 °C, grey to black from above, dark brown from below, surface rough, dry, raised, edge entire.

Material examined: THAILAND, Songkhla Province, Chestnut hill, on submerged wood in a stream, 10 May 2018, W. Dong, hat458-1 (MFLU 18-1539, holotype), ex-type living culture MFLUCC 18-1283; ibid., hat458-2 (HKAS 105039, isotype), ex-type living culture KUMCC 19-0056; Satun Province, on submerged wood in a stream, 10 May 2018, W. Dong, hat286-1 (MFLU 18-1541), living culture MFLUCC 18-1409; ibid., hat286-2 (HKAS 105070).

Notes: Caryospora submersa is similar to other Caryospora species in having superficial, dark, carbonaceous ascomata with flattened bases. Caryospora submersa is very different from other species by obclavate asci and narrowly fusiform, hyaline, thin-walled ascospores contrasting with broadly cylindrical to clavate asci and broadly fusiform, ellipsoidal or biconic, mostly pigmented, relatively thick-walled ascospores (Jeffers 1940; Hawksworth 1982; Abdel-Wahab and Jones 2000; Raja and Shearer 2008; Zhao and Zhao 2012; Ariyawansa et al. 2015). Caryospora submersa does not fit the generic concept of Caryospora except for the large, superficial, carbonaceous ascomata, which are similar to those in other genera within the family. However, C. submersa cannot be placed in other genera due to its distinct morphology and phylogenetically nests in Caryospora with strong bootstrap support (Fig. 2). Thus, Caryospora submersa sp. nov. is introduced.


Key to freshwater Caryospora species

  1. 1.

    Ascospores rough-walled………C. obclavata

  2. 1.

    Ascospores smooth-walled………2

  3. 2.

    Ascospores thin-walled………C. submersa

  4. 2.

    Ascospores thick-walled………3

  5. 3.

    Ascospores pale brown when mature………C. minima

  6. 3.

    Ascospores dark brown to black when mature………4

  7. 4.

    Ascospores 35–48 × 18–28 μm………C. aquatica

  8. 4.

    Ascospores 47–60 × 19–33 μm………C. quercus


Pithomyces Berk. & Broome, J. Linn. Soc., Bot. 14(no. 74): 100 (1873) [1875]

Saprobic in terrestrial or freshwater habitats. Sexual morph: Ascomata dark opaque, solitary to gregarious, erumpent to superficial, mammiform to conical, with ruptured, reflexed, stellate, host remnants around the base, uniloculate, glabrous, brittle, carbonaceous, with an indistinct ostiole. Peridium unequal thickness, poorly developed at the base, composed of thick, dark brown to black cells. Pseudoparaphyses dense, trabeculate, filiform, hyaline, sparsely septate. Asci 8-spored, bitunicate, fissitunicate, cylindrical, with furcate pedicel, apically rounded. Ascospores overlapping biseriate, pale brown to dark brown, fusiform with acute ends, septate, smooth-walled (Pratibha and Prabhugaonkar 2015; Wanasinghe et al. 2018a). Asexual morph: Hyphomycetous. Conidiophores mononematous, micronematous, mostly intercalary, sometimes denticulate, aseptate. Conidiogenous cells mono or polyblastic, hyaline to light brown, smooth. Conidia solitary, brown to black, obovate to oblong, verruculose to spinulose, transversely septate (Pratibha and Prabhugaonkar 2015; Wanasinghe et al. 2018a).

Type species: Pithomyces flavus Berk. & Broome

Notes: Pithomyces is similar to Astrosphaeriellaceae members in having dark, carbonaceous ascomata with a poorly developed base and fusiform ascospores (Liu et al. 2011; Phookamsak et al. 2015b; Wanasinghe et al. 2018a; Jayasiri et al. 2019). Pithomyces clustered in Astrosphaeriellaceae with low bootstrap support (Jayasiri et al. 2019; this study, Fig. 2), but morphologically belongs to Astrosphaeriellaceae.

Over 40 epithets are listed in Index Fungorum (2020), but only three have sequence data, viz. Pithomyces caryotae Wanas. et al., P. flavus and P. licualae Wanas. et al. Pithomyces flavus was found on the natural substratum associated with its sexual morph Astrosphaeriella vesuvius, which was confirmed in the phylogenetic analysis (Pratibha and Prabhugaonkar 2015). Wanasinghe et al. (2018a) found the asexual morphs of P. caryotae and P. licualae from culture, respectively. The asexual morphs of three species share similar morpholigical characters in having obovate, septate, pigmented and rough-walled conidia (Pratibha and Prabhugaonkar 2015; Wanasinghe et al. 2018a). Pithomyces flavus was recorded from freshwater habitats (Cai et al. 2002a; Hu et al. 2013), but not confirmed by molecular data.


List of freshwater Pithomyces species

*Pithomyces flavus Berk. & Broome, J. Linn. Soc., Bot. 14(no. 74): 100 (1873) [1875]

Basionym: Sphaeria vesuvius Berk. & Broome, J. Linn. Soc., Bot. 14(no. 74): 127 (1873) [1875]

Synonymy: Astrosphaeriella vesuvius (Berk. & Broome) D. Hawksw. & Boise, Sydowia 38: 122 (1986) [1985]

Freshwater distribution: China (Cai et al. 2002a; Hu et al. 2013)


Xenoastrosphaeriella Jayasiri, E.B.G. Jones & K.D. Hyde, Mycosphere 10(1): 36 (2019)

Saprobic on bamboo and palms in freshwater or terrestrial habitats. Sexual morph: Ascostromata dark opaque, gregarious, erumpent to superficial, mammiform to conical with ruptured, reflexed, stellate, host remnants around the base, uniloculate, glabrous, brittle, carbonaceous, central ostiolate, with pore-like opening. Peridium unequal thickness, poorly developed at the base, thick at the sides towards the apex, composed of thick, opaque and melanized cells. Pseudoparaphyses dense, trabeculate, hyaline, distinctly septate, anastomosing between the asci, embedded in a hyaline gelatinous matrix. Asci 8-spored, bitunicate, cylindrical, short pedicellate, apically rounded with an ocular chamber. Ascospores overlapping uni- to bi-seriate, brown to reddish brown, with paler end cells, fusiform with acute ends, septate, smooth-walled (Hawksworth and Boise 1985; Phookamsak et al. 2015b; Jayasiri et al. 2019). Asexual morph: Undetermined.

Type species: Xenoastrosphaeriella tornata (Cooke) Jayasiri & K.D. Hyde

Notes: Xenoastrosphaeriella was established by Jayasiri et al. (2019) to accommodate Astrosphaeriella tornata (Cooke) D. Hawksw. & Boise which formed a weakly supported clade with other genera in Astrosphaeriellaceae (Phookamsak et al. 2015b; this study, Fig. 2). Xenoastrosphaeriella is distinct in the family in having paler end cells of ascospores (Hawksworth and Boise 1985; Phookamsak et al. 2015b; Jayasiri et al. 2019). The second species Xenoastrosphaeriella trochus (D. Hawksw.) Phookamsak et al. was recently introduced based on morphology and phylogeny (Hongsanan et al. 2020a). Both species X. tornata and X. trochus were ever collected from freshwater habitats (see list below), but not confirmed by molecular data.


List of freshwater Xenoastrosphaeriella species

*Xenoastrosphaeriella tornata (Cooke) Jayasiri & K.D. Hyde, Mycosphere 10(1): 36 (2019)

Basionym: Trematosphaeria tornata Cooke, Grevillea 16(no. 79): 91 (1888)

Synonymy: Sphaeria tornata Berk. & M.A. Curtis, J. Acad. nat. Sci. Philad., N.S. 2(6): 290 (1854)

Astrosphaeriella tornata (Cooke) D. Hawksw. & Boise, Sydowia 38: 119 (1986) [1985]

Freshwater distribution: Philippines (Cai et al. 2003a)

*Xenoastrosphaeriella trochus (D. Hawksw.) Phookamsak, H.B. Jiang & K.D. Hyde, Mycosphere 11(1): 1762 (2020)

Basionym: Melanomma trochus Penz. & Sacc., Malpighia 11(9–10): 401 (1897)

Synonymy: Leptosphaeria trochus (Penz. & Sacc.) Höhn., Sber. Akad. Wiss. Wien, Math.-naturw. Kl., Abt. 1 118: 328 [54 repr.] (1909)

Asterella trochus (Penz. & Sacc.) Hara, Nippon-gaikingaku: 187 (1936)

Asterotheca trochus (Penz. & Sacc.) I. Hino, Bull. Miyazaki Coll. Agric. Forest. 10: 57 (1938)

Trematosphaeria trochus (Penz. & Sacc.) Teng, Sinensia, Shanghai 9: 257 (1938)

Astrosphaeriella trochus (Penz. & Sacc.) D. Hawksw., J. Linn. Soc., Bot. 82: 46 (1981)

Freshwater distribution: China (Tsui et al. 2000; Cai et al. 2002a, 2006b; Hu et al. 2013), Australia (Vijaykrishna and Hyde 2006), South Africa (Hyde and Frohlich 1998; Hyde et al. 1998)

Key to freshwater Xenoastrosphaeriella species

  1. 1.

    Ascospores 3-septate ········································ X. tornata

  2. 1.

    Ascospores mostly 5-septate ································· X. trochus


Bambusicolaceae D.Q. Dai & K.D. Hyde, Fungal Diversity 63: 49 (2013)


Bambusicola D.Q. Dai & K.D. Hyde, Cryptog. Mycol. 33(3): 367 (2012)

Saprobic on decaying bamboo or parasitic on living plant materials in terrestrial or freshwater habitats. Sexual morph: Ascostromata solitary to gregarious, immersed or semi-immersed and becoming erumpent, variable in shape, conical, convex or dome-shaped to ampulliform, globose to subglobose, with centrally located ostiole, with or without red pigment in the ostiole, uni- to multi- loculate, coriaceous, visible as raised, dark spots or elongate structures on host surface. Peridium composed of dark cells of textura angularis or textura epidermoidea or pseudoparenchymatous cells, normally with the flattened base comprising of thinner, hyaline, smaller cells or intermingling with host cells. Pseudoparaphyses dense, trabeculate, filiform, hyaline, indistinctly septate, anastomosing and branching above the asci. Asci 8-spored, bitunicate, cylindrical or cylindric-clavate, short pedicellate, with a shallow or well-developed chamber. Ascospores uni- to tri-seriate, fusiform, septate, mostly hyaline, rerely pale brown, mostly surrounded by a gelatinous sheath. Asexual morph: Coelomycetous. Conidiomata pycnothyrial, solitary to gregarious, scattered, immersed to semi-immersed, acerose or subglobose, pyriform or irregular, uni- to multi-loculate. Pycnidial wall composed of several layers, thin- to thick-walled, hyaline to dark brown cells of textura globulosa or textura angularis. Conidiophores indistinct. Conidiogenous cells holoblastic, annellidic, discrete, cylindrical, smooth. Conidia sometimes two types, macro and microconidia. Macroconidia cylindrical to ellipsoidal, pale brown to brown, septate, smooth, guttulate. Microconidia globose, oblong to ellipsoidal, hyaline to pale brown, aseptate, smooth, guttulate.

Type species: Bambusicola massarinia D.Q. Dai & K.D. Hyde

Notes: Bambusicola is well-studied because all species have sequence data in GenBank. Bambusicola is distinct in Bambusicolaceae in having fusiform, hyaline, mostly smooth-walled ascospores and cylindrical to ellipsoidal or globose conidia (Dai et al. 2012). The asexual morphs can be found in natural substrate (Bambusicola splendida D.Q. Dai & K.D. Hyde and B. irregulispora D.Q. Dai & K.D. Hyde) or in culture (B. massarinia D.Q. Dai & K.D. Hyde) (Dai et al. 2012). Bambusicola sometimes produces two types of conidia, such as B. dimorpha Thambugala et al. producing macro and micro conidia (Thambugala et al. 2017). Other Bambusicola species only formed macroconidia characterized as cylindrical to ellipsoidal, pale brown to brown, septate conidia. Bambusicola species are saprobic on decaying bamboo from Thailand or parasitic on living leaves or branches of Phyllostachys heteroclada Oliv. from China (Dai et al. 2012, 2015, 2017; Thambugala et al. 2017; Yang et al. 2019a). Herein, we report a new Bambusicola species isolated from submerged wood in China.


List of freshwater Bambusicola species

*Bambusicola aquatica W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557906; Facesoffungi number: FoF09249; Fig. 36

Fig. 36
figure 36

Bambusicola aquatica (MFLU 18-1164, holotype). a, b Appearance of black ascomata on host. c, d Vertical sections of ascomata. e Structure of peridium. f Pseudoparaphyses. g–i Bitunicate asci. j–l Ascospores. m Germinated ascospore. n Colony on PDA (from front). Scale bars: c, d = 100 μm, e = 50 μm, f, j–m = 10 μm, g–i = 20 μm

Full size image

Etymology: referring to aquatic habitat of this fungus

Holotype: MFLU 18-1164

Saprobic on decaying bamboo submerged in freshwater. Sexual morph: Ascomata 180–210 μm high, 250–370 μm diam., black, gregarious, densely clustered in a large group, semi-immersed to superficial, conical or dome-shaped in section. Peridium comprising host and fungal tissues, laterally 85–95 μm thick, composed of dark brown to black, thick-walled cells of textura angularis intermingled with host cells, with thinner basal part 10–35 μm thick, composed of subhyaline, rim-like cells. Pseudoparaphyses 1.4–1.8 μm diam., numerous, trabeculate, filiform, hyaline, indistinctly septate. Asci 80–126.5(–202) × 7–9.5(–10.5) μm (\( {\bar{\text{x}}} \) = 99.5 × 9 μm, n = 15), 8-spored, bitunicate, cylindrical, apically rounded, with a shallow chamber and a short, twisted or straight pedicel, easily and rapidly elongate when mounted in water, up to 202 μm long. Ascospores 18–22.5 × 4–5 μm (\( {\bar{\text{x}}} \) = 20.5 × 4.5 μm, n = 40), overlapping biseriate or partially uniseriate, narrowly fusiform, the upper cell slightly broader than the lower one, sharply narrowed towards to ends which are narrowly rounded, mostly slightly curved, rarely straight, hyaline, 1-septate, deeply constricted at the septum, slightly constricted at a quarter, guttulate, the guttula in upper cell larger than the lower cell, thin-walled, smooth, with a thin, hyaline, inconspicuous, mucilaginous sheath which is 2 μm thick when mounted in water, but 5–10 μm thick when mounted in Indian Ink. Asexual morph: Undetermined.

Culture characteristics: On PDA, colony irregular, reaching 30 mm diam. in 30 days at 25 °C, brown with pale green margin from above, dark brown from below, surface rough, with dense mycelium, dry, mucoid after old, edge undulate.

Material examined: CHINA, Yunnan Province, Dehong, on submerged wood in a stream, 25 November 2017, G.N. Wang, H29A-1 (MFLU 18-1164, holotype), ex-type living culture MFLUCC 18-1031; ibid., H29A-2 (HKAS 101730, isotype), ex-type living culture KUMCC 18-0080.

Notes: Bambusicola aquatica is most similar to B. loculata D.Q. Dai & K.D. Hyde in having narrowly fusiform, 1-septate, hyaline ascospores with narrowly rounded ends and an inconspicuous mucilaginous sheath (Dai et al. 2015). However, they can be distinguished by their ascus characteristics (cylindric-clavate, bi- to tri-seriate, 80–105 × 8–13 μm in B. loculata vs. cylindrical, overlapping biseriate or partially uniseriate, 80–126.5(–202) × 7–9.5(–10.5) μm in B. aquatica). Additionally, the ascomata of B. loculata are solitary or clustered in a small group, while those of B. aquatica are gregarious and densely clustered in a large group.

Bambusicola aquatica forms a strongly supported clade with B. irregulispora (Fig. 2). Bambusicola aquatica is an ascomycetous species, while B. irregulispora is a coelomycetous species. There are respectively 5, 6, 30 and 14 nucleotide differences in the LSU, ITS, TEF and RPB2 sequence data between B. aquatica (MFLUCC 18-1031) and B. irregulispora (MFLUCC 11-0437), which supports them to be different species (Jeewon and Hyde 2016).


Nigrogranaceae Jaklitsch & Voglmayr, Stud. Mycol. 85: 54 (2016)


Nigrograna Gruyter, Verkley & Crous, Stud. Mycol. 75: 31 (2012) [2013]

Saprobic on submerged wood, decaying twigs of shrubs and trees, old fructifications of pyrenomycetes, sometimes as human pathogenic. Sexual morph: Ascomata solitary to gregarious, immersed to superficial, somewhat globose, brown to black, ostiolate. Peridium comprising several layers, angular cells. Pseudoparaphyses numerous, trabeculate, filiform, septate, branching and anastomosing. Asci 8-spored, bitunicate, cylindrical to clavate, or broadly filiform, with short pedicellate and knob-like base. Ascospores overlapping, broadly fusiform to narrowly ellipsoid, inequilateral, septate, hyaline to brown, smooth or faintly verruculose (Jaklitsch and Voglmayr 2016; Hyde et al. 2017; Tibpromma et al. 2017). Asexual morph: Coelomycetous. Pycnidia similar to ascomata, solitary or rarely grerarious, superficial or submerged in agar, globose to subglobose or pyriform, olivaceous to olivaceous-black, with dark brown, septate mycelial outgrowths, with ostiolate papilla. Peridium comprising several layers of pseudoparenchymatous cells. Conidiophores filiform, simple to sparsely branched, with pegs along one or two sides and solitary terminal phialides, reduced in culture. Conidiogenous cells phialidic, discrete, ampulliform, lageniform or subcylindrical, hyaline. Conidia oblong, cylindrical or allantoid, sometimes ellipsoid, hyaline or subhyaline, brown in mass, aseptate, smooth (de Gruyter et al. 2013; Jaklitsch and Voglmayr 2016). Hyphomycetous. Synnemata superficial, effuse, gregarious, black. Mycelium mostly immersed, composed of septate, pale brown, smooth hyphae. Conidiophores macronematous, synnematous on substratum, septate, unbranched to branched, smooth, brown, compacted below half and flared at the tip. Conidiogenous cells polyblastic, terminal or intercalary, sympodial, smooth, swollen subcylindrical, ampulliform, denticulate, covered with several conidiogenous loci, hyaline to subhyaline. Conidia solitary, acrogenous, simple, dry, ellipsoidal, smooth, thin-walled, aseptate, hyaline.

Type species: Nigrograna mackinnonii (Borelli) Gruyter, Verkley & Crous

Notes: Most Nigrograna species were collected from terrestrial habitats and are morphologically very similar. Sexual and asexual morphs have been reported from natural substrates and characterized as ascomycetous and coelomycetous species (Jaklitsch and Voglmayr 2016; Tibpromma et al. 2017). Only Nigrograna cangshanensis was reported from freshwater habitats (China), and it differs from other speices by thicker peridium and molecular characters (Tibpromma et al. 2017). We collected a hyphomycetous species from submerged wood in Thailand, which expands the known morph of this genus.


List of freshwater Nigrograna species

*Nigrograna aquatica W. Dong, H. Zhang & K.D. Hyde, sp. nov.

Index Fungorum number: IF557907; Facesoffungi number: FoF09250; Fig. 37

Fig. 37
figure 37

Nigrograna aquatica (MFLU 17-1661, holotype). a, b Colonies on submerged wood. c–f Conidiophores with attached conidia. g, h Conidiogenous cells. i Conidia. j, k Colony on PDA (up-front, down-reverse). Scale bars: c, d = 20 μm, e = 40 μm, f, g = 10 μm, h = 3 μm, i = 5 μm

Full size image

Etymology: in reference to aquatic habitat of this fungus

Holotype: MFLU 17-1661

Saprobic on submerged wood in freshwater. Sexual morph: Undetermined. Asexual morph: Synnemata 110–125 × 8.5–14.5 μm (\( {\bar{\text{x}}} \) = 118 × 11.8 μm, n = 5), superficial, effuse, gregarious, black, upper part covered with a white or pale grey powdery mass of conidia, composed of paralleled conidiophores. Mycelium mostly immersed, composed of septate, pale brown, smooth hyphae. Conidiophores 100–120 × 1.5–2 μm (\( {\bar{\text{x}}} \) = 100.5 × 1.7 μm, n = 15), macronematous, synnematous on substratum, not compacted, become divergent after mounted in water, unbranched, septate, not constricted at the septa, smooth, dark brown at the base, gradually paler towards the apex, flared in upper part. Conidiogenous cells 4–7.5 × 1.7–2.5 μm (\( {\bar{\text{x}}} \) = 5.6 × 2.2 μm, n = 10), polyblastic, terminal or intercalary, integrated when terminal and discrete when intercalary, sympodial, smooth, swollen subcylindrical, flask-shaped, ampulliform, curved or straight, denticulate, covered with several conidiogenous loci at the upper part, hyaline to subhyaline. Conidia 6–7 × 2–2.5 μm (\( {\bar{\text{x}}} \) = 6.6 × 2.2 μm, n = 15), solitary, acrogenous, simple, dry, ellipsoidal or cylindrical, smooth, thin-walled, aseptate, hyaline.

Culture characteristics: On PDA, colony circular, reaching 25 mm diam. in 30 days at 25 °C, brown to grey from above, dark brown from below, surface rough, with dense mycelium, dry, raised, edge entire.

Material examined: THAILAND, Phayao Province, on submerged wood in a stream, 4 August 2017, G.N. Wang, G11 (MFLU 17-1661, holotype), ex-type living culture MFLUCC 17-2318.

Notes: Nigrograna aquatica is morphologically similar to Phaeoisaria species which are characterized by synnematous conidiophores with polyblastic, sympodial conidiogenous cells (Cai et al. 2006a). However, they cannot be congeneric because N. aquatica differs in having not compacted synnemata which easily become divergent after mounted in water. In contrast, the conidiophores of Phaeoisaria are compacted and parallel adpressed, and never become divergent in water (Liu et al. 2015; Hyde et al. 2018a; Luo et al. 2018). The ellipsoidal or cylindrical, hyaline, aseptate, small conidia of N. aquatica are similar to those coelomycetous species in Nigrograna (de Gruyter et al. 2013; Jaklitsch and Voglmayr 2016). However, phylogenetic analyses showed them as distinct species (Fig. 2). The phylogenetically closest species N. locuta-pollinis F. Liu & L. Cai was isolated from hive-stored pollen collected in Italian honey bee colonies, while N. aquatica was collected from submerged wood in freshwater habitats in Thailand. Morphology of N. locuta-pollinis was not given in protologue (Zhao et al. 2018). A comparison of their nucleotides shows that there are six and eight (including two gaps) nucleotide differences in LSU and ITS sequence data, respectively. We introduce N. aquatica sp. nov. based on the guideline of Jeewon and Hyde (2016) and this is the first hyphomycetous species in Nigrograna.


*Nigrograna cangshanensis Z.L. Luo, H.Y. Su & K.D. Hyde, Fungal Diversity 83: 52 (2017)

Freshwater distribution: China (Tibpromma et al. 2017)


Key to freshwater Nigrograna species

  1. 1.

    Sexual morph………N. cangshanensis

  2. 2.

    Asexual morph………N. aquatica


Corynesporascaceae Sivan., Mycol. Res. 100(7): 786 (1996)


Corynespora Güssow, Z. PflKrankh. 16: 13 (1906)

Pathogenic and saprobic on leaves or submerged wood. Sexual morph: Corynesporasca. Ascomata solitary to aggregated, superficial or immersed, uniloculate, globose, dark brown to black, lacking ostioles. Peridium comprising several layers of thin-walled, brown cells of textura globosa or angularis. Pseudoparaphyses cylindrical, branched, septate, apically free, disintegrate when asci mature. Asci 8-spored, bitunicate, obovoid, deliquescent. Ascospores oblong, pale to dark brown, usually asymmetric, 1-euseptate near the center, with or without indistinctly 1-distoseptate in the upper and lower cell, constricted at the central euseptum (Sivanesan 1996; Hyde et al. 2013). Asexual morph: Hyphomycetous. Colonies on the natural substrate effuse, hairy, amphigenous, brown. Mycelium superficial to immersed in the substrate, composed of branched, septate, thin-walled, smooth, subhyaline to brown hyphae. Conidiophores macronematous, mononematous, erect to procumbent, simple, none to few septate, smooth. Conidiogenous cells monotretic, integrated, terminal, determinate or percurrent proliferation. Conidia acrogenous, solitary, obclavate, wide at the truncate base, tapering towards the apex, pale brown to pale olivaceous brown, multi-distoseptate, smooth-walled, usually with a slightly melanized basal scar (Hyde et al. 2013).

Type species: Corynespora cassiicola (Berk. & M.A. Curtis) C.T. Wei

Notes: Corynespora is characterized by obclavate, brown, multi-distoseptate conidia with slightly melanized basal scar (Hyde et al. 2013). Sivanesan (1996) linked C. caryotae Sivan. as the sexual morph of Corynespora based on cultural method. Based on the extensive use of the name Corynespora for plant pathogenic fungi and its priority, Rossman et al. (2015) formally synonymized Corynesporasca Sivan. under Corynespora. Corynespora is therefore the only genus in Corynesporascaceae (Sivanesan 1996; Wijayawardene et al. 2018). Corynespora was revealed as polyphyletic (Schoch et al. 2009; Voglmayr and Jaklitsch 2017). Two freshwater species C. lignicola Z.L. Luo et al. and C. submersa Z.L. Luo et al. formed a well-supported clade with the type species C. cassiicola, which was represented by a strain CBS 100822 (Hyde et al. 2020a). Corynespora aquatica R.F. Castañeda et al. was reported from freshwater habitats, but not confirmed by molecular data (Castañeda-Ruiz et al. 2004).


List of freshwater Corynespora species

Corynespora aquatica R.F. Castañeda, Heredia & R.M. Arias, Mycotaxon 89: 298 (2004)

Freshwater distribution: Mexico (Castañeda-Ruiz et al. 2004)


*Corynespora lignicola Z.L. Luo, H.Y. Su & K.D. Hyde, Asian Journal of Mycology 3: 65 (2020)

Freshwater distribution: China (Hyde et al. 2020a)


*Corynespora submersa Z.L. Luo, H.Y. Su & K.D. Hyde, Asian Journal of Mycology 3: 63 (2020)