MycoKeys 88: 35-54 (2022) re, A peer-reviewed open-access journal doi: 10.3897/mycokeys.88.79346 RESEARCH ARTICLE . 03 MycoKkeys https://mycokeys.pensoft. net Launched to accelerate biodiversity research Three novel species of Distoseptispora (Distoseptisporaceae) isolated from bamboo in Jiangxi Province, China Zhi-Jun Zhai'”, Jun-Qing Yan'*, Wei-Wu Li', Yang Gao'’, Hai-Jing Hu!”, Jian-Ping Zhou'”, Hai-Yan Song'?, Dian-Ming Hu!” | Bioengineering and Technological Research Centre for Edible and Medicinal Fungi, College of Bioscience and Bioengineering, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China 2 Jiangxi Key Labora- tory for Conservation and Utilization of Fungal Resources, College of Bioscience and Bioengineering, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China 3 Key Laboratory of Crop Physiology, Ecology and Genetic Breeding, Ministry of Education of the P R. China, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China Corresponding author: Dian-Ming Hu (hudianming]@163.com) Academic editor: Cecile Gueidan | Received 16 December 2021 | Accepted 24 February 2022 | Published 22 March 2022 Citation: Zhai Z-J, Yan J-Q, Li W-W, Gao Y, Hu H-J, Zhou J-P, Song H-Y, Hu D-M (2022) Three novel species of Distoseptispora (Distoseptisporaceae) isolated from bamboo in Jiangxi Province, China. MycoKeys 88: 35-54. https:// doi.org/10.3897/mycokeys.88.79346 Abstract Decaying bamboo in freshwater is a unique eco-environment for fungi. Three new Distoseptispora (Distoseptisporaceae) species, D. meilingensis, D. yongxiuensis and D. yunjushanensis from submerged decaying bamboo culms in Jiangxi Province, China, were discovered, based on phylogenetic analyses and morphological characters. The combined data of ITS-LSU-SSU-Tef1 sequences were used to infer the phylogenetic relationship between D. meilingensis, D. yongxiuensis, D. yunjushanensis and related species. Both molecular analyses and morphological data supported D. meilingensis, D. yongxiuensis and D. yunjushanensis as three independent taxa. Keywords Hyphomycetes, phylogenetic analysis, Sordariomycetes, taxonomy, three new taxa Copyright ZhiJun Zhai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 36 Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) Introduction Distoseptispora was established by Su et al. (2016) as the single genus in Distoseptisporaceae. This genus morphologically resembles Ellisembia and Sporidesmium (Subramanian 1992; Shenoy et al. 2006; Yang et al. 2018), while they are not in sister clades in molecular phylogenetic trees (Su et al. 2016; Luo et al. 2019; Hyde et al. 2020, 2021). Multigene analysis showed that Distoseptispora formed a stable and well- supported clade within Distoseptisporales as a sister clade to Aquapteridospora (Luo et al. 2019; Hyde et al. 2020, 2021). Aquapteridospora has been raised as a new family Aquapteridosporaceae for the divergence time (110 million years ago (mya)) falling within the family-level range (50-130 mya) (Hyde et al. 2021). Aguapteridospora and Distoseptispora are similar in having macronematous, mononematous, unbranched conidiophores, mono- or polyblastic, holoblastic, conidiogenous cells and acrogenous, solitary conidia. Distoseptispora can easily be distinguished from Aquapteridospora by its short conidiophores and obclavate or cylindrical, rostrate, euseptate or distoseptate conidia. Additionally, Distoseptispora has terminal conidiogenous cells which lack circular scars (Hyde et al. 2021). Distoseptispora was regarded as saprobic lignicolous fungal genus, which has the ability to decompose lignocelluloses in wood (Wong et al. 1998; Hyde et al. 2016). In recent years, the number of new taxa in Distoseptispora is steadily increasing and currently comprises 35 species, which have been discovered mostly in freshwater and some in terrestrial habitats (Su et al. 2016; Dong et al. 2021; Hyde et al. 2021; Li et al. 2021). Except for the two species, D. adscendens and D. leonensis, which were found from Hungary and Malaysia, respectively (Shoemaker and White 1985; Mcken- zie 1995), 19 of the 33 species has been discovered in Thailand, while the remaining 14 species were introduced from China (Table 2). In China, Distoseptispora species are almost exclusively reported in Yunnan Province (Su et al. 2016; Luo et al. 2018; Hyde et al. 2019; Phookamsak et al. 2019; Li et al. 2021). Only three species, D. martinii, D. bambusae and D. suoluoensis, have been discovered from Guizhou Province (Xia et al. 2017; Yang et al. 2018; Sun et al. 2020). In this study, we introduce three new spe- cies of Distoseptispora, including D. meilingensis, D. yongxiuensis and D. yunjushanensis from Jiangxi Province in subtropical China. We describe the novel species, based on morphological illustrations and phylogenetic analyses. A synopsis of the morphologi- cal characters of Distoseptispora species is also provided. Materials and methods Samples collection, morphological observation and isolation Dead bamboo samples from different freshwater habitats in Jiangxi Province, China, were taken to the lab for detection of fungi using a Nikon SMZ-1270 microscope (Nikon Corporation, Japan). Micro-morphological characteristics were observed and Three novel spcies of Distoseptispora from bamboo af captured using a Nikon ECLIPSE Ni-U compound microscope (Nikon Corporation, Japan), equipped with a Nikon DS-Fi3 camera. All measurements were calculated using PhotoRuler Ver. 1.1 software (Ihe Genus Inocybe, Hyogo, Japan) and figures were processed using Adobe Photoshop CS6 Extended version 10.0 software (Adobe Systems, USA). Pure cultures of the fungi were obtained by the single spore isolation method (Chomnunti et al. 2014). The germinating conidia were transferred to potato dextrose agar (PDA) and incubated at 25 °C for two weeks. The fungal cultures were deposited in the Jiangxi Agricultural University Culture Collection (JAUCC) and the holotypic specimens with MycoBank numbers (842065, 842066, 842067) were de- posited in the Herbarium of Fungi, Jiangxi Agricultural University (HFJAUV) . DNA extraction, PCR amplification and sequencing Fungal genomes were extracted from fresh mycelium using a modified cetyltrimethyl- ammonium bromide (CTAB) method (Doyle and Doyle 1987). Four deoxyribonucle- ic acid (DNA) barcodes (ITS, LSU, SSU and Tef Ja) were chosen for polymerase chain reaction (PCR) using the primer pairs ITS1/ITS4 (White et al. 1990), LROR/LR7 (Hopple and Vilgalys 1999), NS1/NS4 (White et al. 1990) and EF983F/EF2218R (Orstadius et al. 2015), respectively. Amplification reactions were carried out in a volume of 25 ul, containing 12.5 pl 2 x Taq PCR MasterMix (Qingke, Changsha, China), 1 pl each forward and reverse primer (0.2 uM), 1 ul template DNA (circa 50-100 ng) and 9.5 ul ddH,O. Amplifications were conducted under the following conditions: 3 min at 98 °C, 35 cycles of 10s at 98 °C, 10s of annealing at 55 °C and extension at 72 °C for 10 s, with a final 2-min extension at 72 °C. Sequencing reactions were conducted with the corresponding forward and reverse primers commercially by QingKe Biotechnology Co. (Changsha, China). All sequences were edited with Sequencher v.4.14 (GeneCodes Corporation, USA) and have been deposited in the NCBI GenBank database (Table 1). Data analyses Reference sequences of 35 Distoseptispora species and three Aquapteridospora species, based on recent publications (Luo et al. 2019; Hyde et al. 2020; Monkai et al. 2020; Dong et al. 2021, Li et al. 2021) were downloaded from GenBank. Detailed informa- tion on fungal strains used in this paper are provided in Table 1. All obtained sequences were aligned using the online service of MAFFT (Madeira et al. 2019) and refined manually in MEGA vy.7.0 (Kumar et al. 2016). Maximum Likelihood (ML) analysis was conducted with RAxML 8.0 using a GTR-GAMMA model of evolution (Stamatakis 2014). Non-parametric bootstrap analysis was implemented using 1,000 replicates to estimate ML bootstrap (BS) values. Bayesian Inference (BI) analysis was carried out with MrBayes v.3.2 under partitioned models (Ronquist et al. 2012). The best-fit models of nucleotide substitutions were selected according to the Akaike Information Criterion (AIC) implemented in jModelTest2.1.1 38 Table |. Sequences used in this study. Taxa Aquapteridospora aquatica Aquapteridospora fusiformis Aquapteridospora lignicola Distoseptispora adscendens Distoseptispora appendiculata Distoseptispora aquatica Distoseptispora aquatica Distoseptispora aquatica Distoseptispora aquatica Distoseptispora aquatica Distoseptispora bambusae Distoseptispora bambusae Distoseptispora bambusae Distoseptispora cangshanensis Distoseptispora caricis Distoseptispora clematidis Distoseptispora clematidis Distoseptispora dehongensis Distoseptispora euseptata Distoseptispora euseptata Distoseptispora fasciculata Distoseptispora fluminicola Distoseptispora fluminicola Distoseptispora guttulata Distoseptispora hydei Distoseptispora leonensis Distoseptispora lignicola Distoseptispora longispora Distoseptispora martinii Distoseptispora meilingensis Distoseptispora meilingensis Distoseptispora multiseptata Distoseptispora multiseptata Distoseptispora neorostrata Distoseptispora obclavata Distoseptispora obpyriformis Distoseptispora palmarum Distoseptispora palmarum Distoseptispora phangngaensis Distoseptispora phangngaensis Distoseptispora rayongensis Distoseptispora rayongensis Distoseptispora rostrata Distoseptispora rostrata Distoseptispora rostrata Distoseptispora saprophytica Distoseptispora songkhlata Distoseptispora submersa Distoseptispora suoluoensis Distoseptispora suoluoensis Distoseptispora tectonae Distoseptispora tectonae Voucher MFLUCC 17-2371 MFLU 18-1601 MEFLU 15-1172 HKUCC 10820 MFLUCC 18-0259 GZCC 19-0452 MFLUCC 16-0904 MFLUCC 18-0646 MFLUCC 16-1357 S-965 MFLUCC 20-0091 MFLU 20-0261 MEFLU 17-1653 MFLUCC 16-0970 CPC 36498 MFLUCC 17-2145 KUN-HKAS 112708 KUMCC 18-0090 MFLUCC 20-0154 DLUCC 82024 KUMCC 19-0081 DLUCC 0391 DLUCC 0999 MELU 17-0852 MFLUCC 20-0481 HKUCC 10822 MFLUCC 18-0198 HFJAU 0705 CGMCC 318651 JAUCC 4727 JAUCC 4728 MFLUCC 15-0609 MEFLU 17-0856 MFLUCC 18-0376 MFLUCC 18-0329 DLUCC 0867 MFLUCC 18-1446 MELU 18-0588 MELU 17-0855 MFLUCC 16-0857 MFLUCC 18-0415 MELU 18-1045 MFLUCC 16-0969 DLUCC 0885 MELU 18-0479 MFLUCC 18-1238 MFLUCC 18-1234 MFLUCC 16-0946 MFLUCC 17-0224 MFLU 17-0854 MFLUCC 15-0981 MFLUCC 12-0291 LSU NG_075413 MK849798 KU221018 DQ408561 MN163023 MZ227216 MK849794 MK849793 MK849796 MK849792 NG_074430 MT232718 MT232717 MG979761 MN567632 MT214617 MW879523 MK079662 MW081544 MW081545 NG_075417 MG979762 MG979763 MF077554 MT742830 DQ408566 MK849797 MH555357 KX033566 OK562396 OK562397 KX710140 MFO077555 MN163017 MN163010 MG979765 MK079663 NG_067856 MFO077556 NG_073624 MH457137 MG979766 MG979767 NG_064513 NG_075419 MW287755 MG979768 NG_068552 MFO077558 MW287763 KX751713 Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) ITS NR_172447 MK828652 MN163009 MW 133908 MK828649 MK828648 MK828650 MK828647 NR_170068 MT232713 MT232712 MG979754 NR_166325 MT310661 MW723056 MK085061 MW081539 MW081540 NR_172452 MG979755 MG979756 MF077543 MT734661 MK828651 MH555359 KU999975 OK562390 OK562391 KX710145 MF077544 MN163008 MN163012 MG979757 MK085062 NR_165897 MF077545 NR_166230 NR_171938 MH457172 MG979758 MG979759 NR_157552 NR_172454 MW 286482 MG979760 NR_168764 MF077547 MW 286489 KX751711 SSU MW134689 MK828315 MK828317 MK828314 NG_070348 MT232716 MT226728 MW774580 MF077532 MK828318 MH555431 KX033537 OK562402 OK562403 NG_065693 MF077533 MKO079661 MF077534 NG_073504 MH457169 NG_070113 MF077536 Tef- 1x MN194056 MN174866 MN194051 MT232880 MG988419 MK087659 MW084994 MW396656 MG988420 MG988421 MF135651 OK562408 OK562409 MF135659 MG988423 MK087660 MK087660 MF135653 MG988424 MG988425 MW396651 MW396642 MG988426 MF135654 MW396641 KX751710 Three novel spcies of Distoseptispora from bamboo by, Taxa Voucher LSU ITS SSU Tef- 1x Distoseptispora tectonae S-2023 MW081543 MW081538 = — Distoseptispora tectonae GZ 25 MH555358 MH555361 — — Distoseptispora tectonigena MFLUCC 12-0292 KX751714 NR_154018 — — Distoseptispora thailandica MFLUCC 16-0270 MH260292 MH275060 MH260334 MH412767 Distoseptispora thysanolaenae KUN-HKAS 112710 MW879524 MW723057 — _ Distoseptispora thysanolaenae KUN-HKAS 102247 MK064091 MK045851 ah MK086031 Distoseptispora xishuangbannaensis ©KUMCC 17-0290 MH260293 MH275061 MH260335 MH412768 Distoseptispora yongxiuensis JAUCC 4725 OK562394 OK562388 OK562400 OK562406 Distoseptispora yongxiuensis JAUCC 4726 OK562395 OK562389 OK562401 OK562407 Distoseptispora yunjushanensis JAUCC 4723 OK562398 OK562392 OK562404 OK562410 Distoseptispora yunjushanensis JAUCC 4724 OK562399 OK562393 OK562405 OK562411 Distoseptispora yunnansis MFLUCC 20-0153 MW081546 MW081541 = MW084995 ic » . a —”, sequence is unavailable. (Darriba et al. 2012) on XSEDE in the CIPRES web portal (Miller et al. 2010). The models for ITS, LSU, SSU and ef Ja datasets used for phylogenetic analysis are GIT R+I+G model (-InL = 4965.1122), GTR+I+G model (-InL = 2716.7536), TIM2+G (-InL = 4344.2295) and TrN+I+G (-InL = 4479.4914), respectively. The datasets were run for 10,000,000 generations, with four chains and trees sampled every 1,000 generations. ‘The first 10% trees were discarded as burn-in. We used three Aquapteridospora species as outgroups. The Bayesian consensus tree with posterior probabilities (PP) was visualised with FigTree v.1.4.4 (Rambaut 2018) and was edited in Adobe Illustrator CS6. Our aligned matrices and trees can be obtained from TreeBASE (http://purl.org/phylo/treebase/phylows/study/TB2:S29465). Results Molecular phylogenetic results According to the results of BLAST analysis and sequence alignment, the ITS sequence of D. meilingensis has 11 different loci from those of D. yongxiuensis, the ITS sequence of which shares 99% similarity (five different loci) with that of D. suoluoensis. The ITS sequence of D. yunjushanensis is 97% similar (22 different loci) to that of D. obclavata. The aligned matrix for the combined analysis, ITS + LSU + SSU + Te 1a, had 4015 bp, including ITS 596 bp, LSU 799 bp, SSU 1715 bp and Tef- 1x 905 bp. The topologies of trees generated by ML and BI analyses are highly similar. The Bayesian tree with BS and PP is shown in Fig. 1. All species of Distoseptispora form a monophyletic group (BS/PP = 100/1.00). D. yongxiuensis groups together with D. suoluoensis (BS/PP = 60/0.99). These two species and collections of D. meilingensis form a strong-supported clade (BS/PP = 99/1.00), which is strongly linked with sequences of D. bambusae (BS/ PP = 100/1.00). Collections of D. yunjushanensis form a moderate-support clade (BS/ PP = 81/1.00) with the lineage consisting of D. obclavata and D. rayongensis. 40 Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) Distoseptispora tectonae MFLUCC 15-0981 Distoseptispora tectonae S-2023 Distoseptispora tectonae GZ 25 Distoseptispora tectonae MFLUCC 12-0291 Distoseptispora submersa MFLUCC 16-0946 83/1 Distoseptispora fasciculata KUMCC 19-0081 Distoseptispora A HITE KUMCC 17-0290 Distoseptispora fluminicola DLUCC 0391 ; —_- Distoseptispora fluminicola DLUCC 0999 Distoseptispora tectonigena MFLUCC 12-0292 Distoseptispora clematidis KUN HKAS 112708 Distoseptispora clematidis MFLUCC 17-2145 Distoseptispora adscendens HKUCC 10820 Distoseptispora aquatica MFLUCC 16-1357 Distoseptispora aquatica $-965 Distoseptispora aquatica MFLUCC 16-0904 Distoseptispora aquatica MFLUCC 18-0646 Distoseptispora longispora HFJAU 0705 Distoseptispora phangngaensis MFLUCC 16-0857 Distoseptispora phangngaensis MFLU 17-0855 Distoseptispora thailandica MFLUCC 16-0270 Distoseptispora multiseptata MFLUCC 15-0609 Distoseptispora multiseptata MFLU 17-0856 '—__ Djistoseptispora cangshanensis MFLUCC 16-0970 istoseptispora rostrata DLUCC-0885 Distoseptispora rostrata MFLU 18-0479 92/1 100/1 100 Distoseptispora rostrata MFLUCC 16-0969 4100/1 Distoseptispora hydei MFLUCC 20-0115 4100/1 Distoseptispora obpyriformis DLUCC 0867 [| 100/1, Distoseptispora thysanolaenae KUN HKAS 102247 | 100/1 ll 97/1 Ba 95/1 99/1 -10.99 99/1 95/1 100/1 100/1 99/1 100/104 99/ | | 1 100/- 2 004 100/1, Distoseptispora rayongensis 81/1 = Distoseptispora lignicola MFL 94/1 Aquapteridospora aquatica MFLUCC 17-2371 Aquaptendospora lignicola MFLU 15-1172 Aquapteridospora fusiformis MFLU 18-1601 Distoseptispora thysanolaenae KUN HKAS 112710 Distoseptispora dehongensis KUMCC 18-0090 FLUCC 18-0415 Distoseptispora rayongensis MFLU 18-1045 Distoseptispora obclavata MFLUCC 18-0329 Distoseptispora yunjushanensis JAUCC 4723 Distoseptispora yunjushanensis JAUCC 4724 Distoseptispora gutiulata MFLU 17-0852 Distoseptispora songkhlaensis MFLUCC 18-1234 Distoseptispora martinii CGMCC 318651 UCC 18-0198 * Distoseptispora caricis CPC 36498 Distoseptispora leonensis HKUCC 10822 Distoseptispora appendiculata MFLUCC 18-0259 Distoseptispora neorostrata MFLUCC 18-0376 Distoseptispora palmarum MFLUCC 18-1446 Distoseptispora palmarum MFLU 18-0588 Distoseptispora saprophytica MFLUCC 18-1238 Distoseptispora suoluoensis MFLUCC 17-0224 Distoseptispora suoluoensis MFLU 17-0854 Distoseptispora yongxiuensis JAUCC 4725 Distoseptispora yongxiuensis JAUCC 4726 Distoseptispora meilingensis JAUCC 4727 Distoseptispora meilingensis JAUCC 4728 Distoseptispora bambusae MFLUCC 20-0091 Distoseptispora bambusae MFLU 20-0261 Distoseptispora bambusae MFLU 17-1653 100/1,- Distoseptispora euseptata MFLUCC 20-0154 Distoseptispora euseptata DLUCC $ 2024 Distoseptispora yunnansis MFLUCC 20-0153 ] Aquapteridospora (outgroup) 100/1 Figure |. Phylogenetic tree of Distoseptispora, inferred from the combined regions (ITS-LSU-SSU- Tef 1a) using Bayesian Inference (BI) analysis. The Aquapteridospora clade was used as the outgroup. The lineages with new species were shown in bold. PP = 0.95 and BS = 75% were indicated around the branches. The new sequences generated in this study are given in bold. Taxonomy Distoseptispora meilingensis Z. J. Zhai & D. M. Hu, sp. nov. MycoBank No: 842067 Bigi2 Etymology. Referring to the collecting site of the Meiling Mountain in Jiangxi Province, China. Holotype. HFJAU 10009. Three novel spcies of Distoseptispora from bamboo Figure 2. Distoseptispora meilingensis (HFJAU10009, holotype) a, b colonies on bamboo culms c-e conidiophores with conidia f conidiogenous cells g, n conidiogenous cells with conidia h—m conidia © germinating conidium p culture on PDA from above and reverse. Scale bars: 100 pm (a, b), 20 um (c—e, 0), 5 um (f-n). 42 Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) Description. Saprobic on culms of bamboo. Sexual morph: Undetermined. Asexual morph: Uyphomycetous. Colonies effuse, brown to dark brown, hairy. Mycelium mostly immersed, composed of pale to dark brown, septate, branched, smooth, hyaline to subhyaline hyphae. Conidiophores 69-192 x 4—7 um (X = 120.6 x 5.5 um, n = 25), macronematous, mononematous, erect, cylindrical, straight or slightly flexuous, 5—12-septate, yellowish-brown or brown, robust at the base. Conidiogenous cells holoblastic, mono- to polyblastic, integrated, terminal, cylindrical, yellowish- brown or brown. Conidia 32-64.5 x (7-)9-12.5 um (x = 43.7 x 9.8 um, n = 30), acrogenous, solitary, straight or slightly curved, obclavate, 5—7-distoseptate, thick- walled, rounded at the apex, truncate at the base, tapering towards apex, bud scars disjunctors at base, mostly brown when mature. Cultural characteristics. Conidia germinating on PDA within 24 h and germ tubes produced from both ends. Colonies on PDA reaching 17—23 mm diam. at two weeks at 25 °C, in natural light, circular, with dense, light olivaceous mycelium on the surface with entire margin; reverse brown to dark brown. Material examine. Curna, Jiangxi Province, Nanchang City, Meiling Mountain, alt. 305 m, near 28.79°N, 115.72°E, on decaying bamboo culms submerged in a fresh- water stream, 16 Aug 2021, Z. J. Zhai, SLI-3 (HFJAU10009, holotype), ex-type liv- ing culture, JAUCC 4727 = JAUCC 4728. Notes. Distoseptispora meilingensis clusters with the clade including D. suoluoensis and D. yongxiuensis with high support in the phylogenetic tree (Fig. 1). Distoseptispora meilingensis is distinct from D. suoluoensis (Yang et al. 2018) and D. yongxiuensis by its conidial colour (mostly brown, yellowish-brown to dark olivaceous and yellowish- brown or brown, respectively). Furthermore, D. meilingensis has shorter conidia (32—64.5 um vs. (65—)80—-125(—145) um) than those of D. suoluoensis (Yang et al. 2018) and slightly shorter conidiophores (69-192 ym vs. 112—253 um) than those of D. yongxiuensis. Distoseptispora meilingensis resembles D. bambusae in similar habitats and polyblastic conidiogenous cells (Sun et al. 2020). However, D. meilingensis can be distinguished from D. bambusae in its longer conidiophores (69-192 um vs. 40— 96 um), slightly wider (up to 12.5 um vs. up to 9.5 pm) and brighter (light brown vs. brown) conidia (Sun et al. 2020). A comparison of morphological features of Distoseptispora species is provided in Table 2. Distoseptispora yongxiuensis Z. J. Zhai & D. M. Hu, sp. nov. MycoBank No: 842066 Fig. 3 Etymology. With reference to Yongxiu, from where the holotype was collected. Holotype. HFJAU10007 Description. Saprobic on decaying bamboo culms. Sexual morph: Undeter- mined. Asexual morph: Hyphomycetous. Colonies effuse, brown, hairy, glistening, often inconspicuous. Mycelium partly superficial, partly immersed in the substra- 43 Three novel spcies of Distoseptispora from bamboo poom snooovarjo “eidas 1 paiorsuo0s (IZOZ) JeIOIT ueuunZ ‘euryD possiourqns poynuepruy, soyeMmyYsery uayjo ‘ayeavpogo 01 uIOFTIAdGG aieydasno-/-4 6-8 x PS-LE G-¥ X 87-6 vywidasna ‘ snOdoRATTO poom ‘gepnSosa1 JO TeotrpurfAo peorq (6107) ‘Je opAEE ueUUNA ‘eUTYyD §podsioUgns poyNuepluA, sJoIeMYseLI ‘oyeaRpogo 0} wiso0pAdqca, areidasoisip-¢-¢ O1-S°Z x O€-ZI G- x 08-SP stsuasuogap ‘q JusUTYIeNe Jo ss ay} ye quasaid syojoun{sIp Jo sreds png (OZOZ) ‘Te rey Surry) SISUIUIYAIS SIIDULI]) 93UN Ud0I8 YIM UMOI ‘aIeISOI jo epyesmeyny g - :7 | - Figure 3. Distoseptispora yongxiuensis HFJAU 10007, holotype) a Colonies on bamboo culm b, d conidiophores with conidia € conidiogenous cell bearing conidium e conidiogenous cells with young conidia f-k conidia I germinating conidium m culture on PDA from above and reverse. Scale bars: 100 um (a), 20 um (b=e, I), 5 um (f-K). Three novel spcies of Distoseptispora from bamboo 47 tum, composed of hyaline to pale brown, septate, branched hyphae. Conidiophores 112-253 x 4-9 um (x = 198 x 6.9 um, n = 15), macronematous, mononematous, solitary or aggregated at the base, cylindrical, straight or slightly flexuous, 8—13-sep- tate, olivaceous to dark brown, sharply curving near the base, paler at the apical part, rounded at the apex. Conidiogenous cells integrated, terminal, monoblastic, rarely polyblastic, cylindrical, olivaceous to dark brown. Conidia 46—74(-86) x 10-13(-16) um (xX = 65.6 x 12.6 um, n = 30), acrogenous, solitary, obclavate or obspathulate, straight or flexuous, rostrate, 6—9-euseptate, olivaceous to yellowish-brown or brown, becoming paler or hyaline towards the apex, guttulate, 2.5—4 um wide at the base and 2.5—5 um wide at the apex, with a darkened scar at the base. Cultural characteristics. Conidia germinating on PDA within 24 h and germ tubes produced from both ends. Colonies on PDA reaching 24-32 mm diam. at two weeks at 25 °C, in natural light, circular, with dense, light olivaceous mycelium on the surface with entire margin; reverse dark brown to black. Material examined. Cuina, Jiangxi Province, Jiujiang City, Yongxiu County, alt. 680.5 m, 29.09°N, 115.62°E, on decaying bamboo culms submerged in a freshwater stream, 28 Apr 2020, Z. J. Zhai and W. W. Li, YJS-70 (HFJAU 10007, holotype), ex- type living culture, JAUCC 4725 = JAUCC 4726. Notes. In the multi-gene phylogenetic tree (Fig. 1), D. yongxiuensis clusters with D. suoluoensis. Nonetheless, D. yongxiuensis can be distinguished from D. suoluoensis by its shorter conidia (46—74(-86) um vs. (65—)80—125(—145) ym) and polyblastic conidiogenous cells (Yang et al. 2018). Additionally, D. swoluoensis has the percurrent proliferation of conidia, while it was not observed in D. yongxiuensis. Distoseptispora yongxiuensis is similar with D. bambusae (Sun et al. 2020), D. palmarum (Hyde et al. 2019) and D. meilingensis for the polyblastic conidiogenous cells, but D. yongxiuensis has wider conidia than those of D. bambusae (10—13(-16) um vs. 5.5—9.5 um) (Sun et al. 2020), shorter conidia than those of D. palmarum (46—74(-86) um vs. 35-180 pm) (Hyde et al. 2019) and paler (yellowish-brown or brown vs. bright brown) conidia than those of D. meilingensis. Distoseptispora yunjushanensis Z. J. Zhai & D. M. Hu, sp. nov. MycoBank No: 842065 Fig. 4 Etymology. The epithet refers to the collecting site from the Yunjushan Mountain in China. Holotype. HFJAU10005 Description. Saprobic on decaying bamboo culms submerged in freshwater habitats. Sexual morph: Undetermined. Asexual morph: Hyphomycetous. Colonies effuse, olivaceous or dark brown, hairy, velvety. Mycelium mostly immersed, consisting of branched, septate, smooth, subhyaline to pale brown hyphae. Conidiophores 100-175 um x 5.5-10 um (x = 129x7.1 um, n = 30), single or in groups of 2 or 3, macronematous, mononematous, erect, straight or slightly flexuous, 4—7-septate, Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) Figure 4. Distoseptispora yunjushanensis (HFJAU 10005, holotype) a, b colonies on bamboo culms c-f conidiophores with conidia g-i young conidia j-l mature conidia m conidium with proliferation N germinating conidium 0, p culture on PDA from above and reverse. Scale bars: 100 um (a, b), 20 um (c—f, m,n), 5 um (g-H). Three novel spcies of Distoseptispora from bamboo 49 unbranched, olivaceous to dark brown, smooth, cylindrical, rounded at the apex. Conidiogenous cells monoblastic, integrated, terminal, determinate, pale to dark brown, cylindrical. Conidia 39-67.5(-77) um x (7—)9.5-13.5(-16.5) um (x = 52 x 12 um, n = 30), acrogenous, solitary, obpyriform or obclavate, thick-walled, tapering towards the rounded apex, slightly curved, truncate at the base, 7—13-distoseptate, guttulate, smooth-walled, olivaceous, dark brown when mature, sometimes with the percurrent proliferation which forms another conidium from the conidial apex. Cultural characteristics. Conidia germinating on PDA within 24 h and germ tubes produced from both ends. Colonies on PDA reaching 12-18 mm diam. at 14 days at 25 °C, in natural light, with fluffy, dense, thin olivaceous mycelium in the cen- tre, becoming sparse and paler at the entire margin; reverse dark brown, pale brown at the smooth margin. Material examined. Cuina, Jiangxi Province, Jiujiang City, Yongxiu County, Yunjushan Mountain, alt. 672.5 m, 29.23°N, 115.59°E, on decaying bamboo culms submerged in a freshwater stream, 28 Apr 2020, Z. J. Zhai and W. W. Li, YJS-42 (HF- JAU 10005, holotype), ex-type living culture, JAUCC 4723 = JAUCC 4724. Notes. In the phylogenetic analysis, D. yunjushanensis clusters with D. obclavata and D. rayongensis with moderate support (BS/PP = 81/1.00). However, D. yunjushanensis is easily distinguished from D. obclavata by its comparatively wider (5.5—10 um vs. 5-7 um) conidiophores and conidia ((7—)9.5—13.5(—16.5) um vs. 9-11 um) (Luo et al. 2019). Moreover, the percurrent proliferation of conidia was not observed in D. obclavata (Luo et al. 2019). Distoseptispora yunjushanensis has shorter conidia (39-67.5(-77) um vs. (36—)60-106(-120) um) and wider conidiophores (5.5-10 um vs. 3.5—-5.5 um) than those of D. rayongensis (Hyde et al. 2020). The morphology of D. yunjushanensis is similar to D. guttulata and D. songkhlaensis in having the obclavate conidia, but differs in having wider (5.5—-10 um vs. 3.5—5.5 um and 4—5.5 um) conidiophores, shorter (39-67.5(-77) um vs. 75—130(—165) um and 44-125 pm) and proliferating conidia (Yang et al. 2018; Dong et al. 2021). Additionally, D. yunjushanensis can be distinguished from D. guttulata by its distoseptate conidia (Yang et al. 2018). Discussion Previous reports of Distoseptispora were mainly concentrated in tropical areas, such as Thailand (Chiang Rai, Phitsanulok, Phang Nga; Luo et al. 2019) and southwest Yun- nan, China (Su et al. 2016; Luo et al. 2018). Nonetheless, several new taxa were found sporadically in subtropical China, for example, Distoseptispora martinii (Xia et al. 2017), D. suoluoensis (Yang et al. 2018) and D. bambusae (Sun et al. 2020) in Guizhou Province and D. euseptata and D. yunnansis in northwest Yunnan (Li et al. 2021). The ongoing discovery of this taxa from other geographic regions in subtropical China will deepen our understanding of the species in this genus. In this study, we introduced another three new species of Distoseptispora from Jiangxi Province of subtropical China. It is in- teresting to note that all these species in subtropical China, except D. yunjushanensis and 50 Zhi-Jun Zhai et al. / MycoKeys 88: 35-54 (2022) D. martinii, formed a well-supported monophyletic clade in the phylogenetic tree and this clade was at the basal position (Fig. 1). Distoseptispora yunjushanensis and D.martinii were otherwise phylogenetically placed within other clades (Fig. 1) and, therefore, we suppose that other lineages might also comprise more Distoseptispora species distributed in subtropical China. Further discovery of Distoseptispora species in more extensive areas in subtropical and other regions of China are needed to be addressed if the phylogenetic position of species reflects their geographical and ecological distribution. Distoseptisporaceae is a holomorphic group of Sordariomycetes that are saprobic on decaying wood and plant debris in terrestrial and freshwater habitats (Su et al. 2016). The genus Distoseptispora seems not to have specific habitat preferences, as most species were reported from submerged wood in freshwater habitats, while some were intro- duced from terrestrial habitats (Table 2). So far, only five species of Distoseptispora have been found on bamboo, two of them (Distoseptispora bambusae and D. hydei, Table 2) from terrestrial habitats, the other three (this study) from freshwater. There may be more species in this genus existing on bamboo waiting to be discovered and further studies are needed to clarify if a specific species in Distoseptispora is specific to its host. Acknowledgements We are grateful to Deng-Mei Fan and Yi Yang (Agricultural college, Jiangxi Agricultural University) for the valuable advice in the context of this study. This study was supported by the National Natural Science Foundation of China (NSFC 32070023 and NSFC 32060014), the Natural Science Foundation of Jiangxi Province (20151BAB214002) and Science and Technology Plan Project of Jiangxi Province (GJJ160417). References Chomnunti P, Hongsanan S, Aguirre-Hudson B, Tian Q, Persoh D, Dhami MK, Alias AS, Xu J, Liu X, Stadler M, Hyde KD (2014) The sooty moulds. 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