﻿Three novel species of Distoseptispora (Distoseptisporaceae) isolated from bamboo in Jiangxi Province, China

﻿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.

Introduction 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 (The 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 deposited in the Herbarium of Fungi, Jiangxi Agricultural University (HFJAU) .

Data analyses
Reference sequences of 35 Distoseptispora species and three Aquapteridospora species, based on recent publications Monkai et al. 2020;Dong et al. 2021, Li et al. 2021 were downloaded from GenBank. Detailed information 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 v.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
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.

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 Yunnan, China 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 interesting to note that all these species in subtropical China, except D. yunjushanensis and 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 ). The genus Distoseptispora seems not to have specific habitat preferences, as most species were reported from submerged wood in freshwater habitats, while some were introduced 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.