Exploring ascomycete diversity in Yunnan II: Introducing three novel species in the suborder Massarineae (Dothideomycetes, Pleosporales) from fern and grasses Author Phookamsak, Rungtiwa https://orcid.org/0000-0002-6321-8416 Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand & Department of Economic Plants and Biotechnology, Yunnan Key Laboratory for Wild Plant Resources, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China & Honghe Center for Mountain Futures, Kunming Institute of Botany, Chinese Academy of Sciences, Honghe, 654400, Yunnan Province, China Author Hongsanan, Sinang https://orcid.org/0000-0003-0550-3152 Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand & Shenzhen Key Laboratory of Microbial Genetic Engineering, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen 518060, China Author Bhat, Darbhe Jayarama https://orcid.org/0000-0002-3800-5910 Department of Botany and Microbiology, College of Science, King Saud University, P. 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Box 2455, Riyadh 11451, Saudi Arabia & Vishnugupta Vishwavidyapeetam, Ashoke, Gokarna 581326, India Author Wanasinghe, Dhanushka N. https://orcid.org/0000-0003-1759-3933 Department of Economic Plants and Biotechnology, Yunnan Key Laboratory for Wild Plant Resources, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China & Honghe Center for Mountain Futures, Kunming Institute of Botany, Chinese Academy of Sciences, Honghe, 654400, Yunnan Province, China & CIFOR-ICRAF China Program, World Agroforestry (ICRAF), Kunming 650201, Yunnan Province, China & Center for Mountain Futures (CMF), Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China Author Promputtha, Itthayakorn https://orcid.org/0000-0003-3376-4376 Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand Author Suwannarach, Nakarin https://orcid.org/0000-0002-2653-1913 Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand Author Kumla, Jaturong https://orcid.org/0000-0002-3673-6541 Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand Author Xie, Ning https://orcid.org/0000-0002-5866-8535 Shenzhen Key Laboratory of Microbial Genetic Engineering, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen 518060, China Author Dawoud, Turki M. https://orcid.org/0000-0002-1444-4185 Department of Botany and Microbiology, College of Science, King Saud University, P. 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Box 2455, Riyadh 11451, Saudi Arabia Author Mortimer, Peter E. https://orcid.org/0000-0002-8507-7407 Department of Economic Plants and Biotechnology, Yunnan Key Laboratory for Wild Plant Resources, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China & Honghe Center for Mountain Futures, Kunming Institute of Botany, Chinese Academy of Sciences, Honghe, 654400, Yunnan Province, China Author Xu, Jianchu https://orcid.org/0000-0002-2485-2254 Department of Economic Plants and Biotechnology, Yunnan Key Laboratory for Wild Plant Resources, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China & Honghe Center for Mountain Futures, Kunming Institute of Botany, Chinese Academy of Sciences, Honghe, 654400, Yunnan Province, China & CIFOR-ICRAF China Program, World Agroforestry (ICRAF), Kunming 650201, Yunnan Province, China & Department of Economic Plants and Biotechnology, Yunnan Key Laboratory for Wild Plant Resources, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan Province, China jxu@mail.kib.ac.cn Author Lumyong, Saisamorn https://orcid.org/0000-0002-6485-414X Center of Excellence in Microbial Diversity and Sustainable Utilization, Chiang Mai University, Chiang Mai 50200, Thailand & Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand scboi009@gmail.com text MycoKeys 2024 2024-04-16 104 9 50 http://dx.doi.org/10.3897/mycokeys.104.112149 journal article http://dx.doi.org/10.3897/mycokeys.104.112149 1314-4049-104-9 BA6C35A352D8524293F9CFED6D649F88 Bambusicola hongheensis Phookamsak, Bhat & Hongsanan sp. nov. Fig. 4 Etymology. The specific epithet " hongheensis " refers to the locality, Honghe Hani and Yi Autonomous Prefecture (Yunnan, China), where the holotype was collected. Figure 4. Bambusicola hongheensis (KUN-HKAS 129042, holotype) A the appearance of ascomata on the host surface B vertical section of an ascoma C, D peridia E pseudoparaphyses F, G asci embedded in pseudoparaphyses H-K ascospores L, M ascospores stained in India Ink show a thin mucilaginous sheath surrounding ascospores. Scale bars: 100 μm ( B ); 20 μm ( C-G ); 10 μm ( H-M ). Holotype. KUN-HKAS 129042. Description. Saprobic on dead culm of bamboo in terrestrial habitats, visible as black, shiny, gnarled on the host surface. Sexual morph : Ascomata 225-350 μm high, 340-590 μm diam., scattered, sometimes forming stroma with a clustered 1-3 ascomata, gregarious, semi-immersed, raised, becoming superficial, dark brown, dome-shaped to subconical or subglobose, glabrous, coriaceous, ostiolate with inconspicuous papilla. Peridium 40-80(-130) μm wide at sides towards the apex, 10-25 μm wide at the base, composed of several layers of small, dark brown pseudoparenchymatous cells, outer layer fused with host cells, arranged in textura angularis to textura globulosa , inner layer composed of 1-3 strata of flattened cells, of textura globulosa to textura prismatica , with thick, palisade-like cells at the sides. Hamathecium composed of 1-3 μm wide, filiform, dense, septate, branched, pseudoparaphyses, anastomosed between and above the asci, embedded in a gelatinous matrix. Asci (58-)70-90(-105)(-119) x 12-15(-17) μm ( x̄ = 80.5 x 13.5 μm , SD = +/- 13.2 x 1.8, n = 25), 8-spored, bitunicate, fissitunicate, cylindrical-clavate, shortly pedicellate, apically rounded with well-developed ocular chamber. Ascospores 22-26(-30) x 4.5-7 μm ( x̄ = 24.6 x 5.4 μm , SD = +/- 2.3 x 0.5, n = 30), overlapping 1-3-seriate, hyaline, fusiform, slightly curved, 1-septate, occasionally 2-3-septate, slightly constricted at the septum, the upper cell slightly larger than the lower cell, smooth-walled, surrounded by a thin, indistinct, mucilaginous sheath. Asexual morph : Undetermined. Distribution. China (Yunnan). Specimen examined. China. Yunnan Province: Honghe Hani and Yi Autonomous Prefecture, Honghe County, rice terraces, on dead culm of bamboo, 26 Jan 2021, R. Phookamsak BN06 (KUN-HKAS 129042, holotype ). Notes : As the axenic culture is not active, the sequences of SSU and rpb2 were obtained from genomic DNA extracted from ascomata and dried culture. Notes. Based on the NCBI nucleotide BLAST search of ITS sequence, Bambusicola hongheensis (KUN-HKAS 129042) has the closest match with B. triseptatispora (MFLUCC 11-0166, ex-type strain) with 98.71% similarity (Identities = 535/542 with no gap) and is similar to B. loculata (MFLU 15-0056, ex-type strain) with 98.69% similarity (Identities = 528/535 with 1 gap) and B. splendida (MFLUCC 11-0611) with 98.25% similarity (Identities = 392/399 with no gap). The NCBI nucleotide BLAST search of LSU sequence indicated that B. hongheensis has the closest match with B. triseptatispora (MFLUCC 11-0166, ex-type strain) and B. didymospora (MFLUCC 10-0557, ex-type strain) with 100% similarity (Identities = 802/802 with no gap) and is similar to B. loculata (MFLU 15-0056, ex-type strain) with 99.75% similarity (Identities = 813/815 with 2 gaps) and B. nanensis (MFLUCC 21-0063, ex-type strain) with 99.49% similarity (Identities = 785/789 with no gap). The NCBI nucleotide BLAST search of rpb2 sequence indicated that B. hongheensis has the closest match with B. loculata (MFLU 15-0056, ex-type strain) with 99.90% similarity (Identities = 1042/1043 with no gaps) and is also similar to B. triseptatispora (MFLUCC 11-0166, ex-type strain) with 97.92% similarity (Identities = 990/1011 with no gap) and B. massarinia (voucher MFLU 11-0389) with 93.57% similarity (Identities = 975/1042 with 4 gaps). Phylogenetic analyses of a concatenated ITS, LSU, rpb2 , SSU and tef1-α sequence dataset demonstrated that Bambusicola hongheensis formed a separate branch (85% ML, 1.00 PP; Fig. 1 ), and clustered with B. loculata and B. triseptatispora with high support (100% ML, 1.00 PP; Fig. 1 ) and also clustered with the generic type of Bambusicola , B. massarinia with significant support (73% ML, 0.99 PP; Fig. 1 ). A nucleotide pairwise comparison of ITS sequence indicated that B. hongheensis differs from B. triseptatispora in 35/600 bp (5.83%), differs from B. loculata in 16/547 bp (2.92%) and differs from B. massarinia in 72/608 bp (11.84%). Whereas the nucleotide pairwise comparison of LSU sequence indicated that B. hongheensis is consistent with B. triseptatispora (0/802 bp) and B. loculata (1/816 bp), but differs from B. massarinia in 7/803 bp (0.87%). Furthermore, the nucleotide pairwise comparison of rpb2 sequence indicated B. hongheensis is not significantly different from B. loculata (1/1043 bp), but differs from B. triseptatispora in 21/1012 bp (2.07%) and differs from B. massarinia in 68/1042 bp (6.52%). Morphologically, Bambusicola hongheensis resembles B. loculata and B. triseptatispora in terms of the size range of ascomata, asci and ascospores. However, B. hongheensis has comparatively smaller ascomata (340-590 μm diam. of B. hongheensis vs. 350-600 μm diam. of B. loculata vs. 470-730 μm diam. of B. triseptatispora ), shorter and wider asci ((58-)70-90(-105)(-119) x 12-15(-17) μm vs. 80-105 x 8-13 μm vs. (78-)80-100(-110) x 10-12(-14) μm , respectively) and sharing the size range of ascospores (22-26(-30) x 4.5-7 μm vs. 22-26.5 x 5-6 μm vs. (25-)26-30(-31) x 4-6 μm , respectively). The ascospores of B. hongheensis are typically hyaline, 1-septate, whereas B. triseptatispora has hyaline to pale brown and 3-septate ascospores ( Dai et al. 2017 ). Distinguishing B. loculata from B. hongheensis , based on morphological characteristics alone is challenging, but B. loculata can be differentiated by its larger ascomata and asci ( Dai et al. 2015 ). However, a clear differentiation is achieved through phylogenetic evidence (Fig. 2 ) and nucleotide pairwise comparison of ITS gene region (2.92% difference).