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Research Article
Two new species of Neohelicosporium (Tubeufiaceae, Tubeufiales) from freshwater and terrestrial habitats in China
expand article infoXiao-Yan Ma‡, Yong-Zhong Lu‡, Lei He§, Dan-Dan Song|, Jian Ma‡
‡ Guizhou Institute of Technology, Guiyang, China
§ Guizhou Academy of Agricultural Sciences, Guiyang, China
| College of Brewing Engineering, Renhuai, China
¶ Guizhou Industry Polytechnic College, Guiyang, China

Corresponding author: Dan-Dan Song ( aisu927@163.com )

Corresponding author: Jian Ma ( yanmajian@163.com )

Academic editor: Samantha C. Karunarathna
© 2025 Xiao-Yan Ma, Yong-Zhong Lu, Lei He, Dan-Dan Song, Jian Ma.
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.
Citation: Ma X-Y, Lu Y-Z, He L, Song D-D, Ma J (2025) Two new species of Neohelicosporium (Tubeufiaceae, Tubeufiales) from freshwater and terrestrial habitats in China. MycoKeys 118: 1-17. https://doi.org/10.3897/mycokeys.118.151514
Open Access

Abstract

Neohelicosporium species are a group of helicosporous hyphomycetes with the potential to produce novel bioactive natural compounds. During our investigation on helicosporous hyphomycetes, six isolates were collected from freshwater and terrestrial habitats in the tropical and subtropical regions of southern China. Based on multi-gene phylogenetic analyses (LSU-ITS-tef1-α-rpb2) and morphological comparisons of Neohelicosporium taxa, two new species (N. guizhouense and N. wuzhishanense) are introduced. Detailed micro-morphological descriptions, illustrations, and DNA molecular data are provided for the newly introduced species to confirm their taxonomic placements.

Key words:

2 new species, asexual morph, Dothideomycetes, phylogeny, saprobic fungi, taxonomy

Introduction

Neohelicosporium was established by Lu et al. (2018a) based on morphological characteristics and phylogenetic evidence. In their study, five new species, Neohelicosporium aquaticum, N. guangxiense, N. hyalosporum, N. parvisporum (the type species), and N. thailandicum, were collected from submerged decaying wood in southern China and Thailand (Lu et al. 2018a). Jayasiri et al. (2017) reported the first asexual-sexual morph of Neohelicosporium fusisporum, collected from decaying fruit of a Malvaceae species in Thailand. Subsequently, Lu et al. (2018b) reassessed the order Tubeufiales based on DNA sequence data and morphological characteristics, introducing seven new species and nine new combinations within Neohelicosporium (Morgan 1892; Penzig 1897; Linder 1929; Moore 1954; Rao and Rao 1964; Matsushima 1971, 1980; Rossman 1979, 1987; Chouhan and Panwar 1980; Goos 1985, 1986, 1989; Ho et al. 2001; Tsui et al. 2001, 2006; Cai et al. 2006; Zhao et al. 2007; da Cruz et al. 2009; Promputtha and Miller 2010; Singh and Singh 2016; Jayasiri et al. 2017; Kuo and Goh 2018; Lu et al. 2018a). Dong et al. (2020) introduced another species, Neohelicosporium submersum, collected from a freshwater habitat in Thailand. Subsequent studies reporting novel Neohelicosporium species are listed chronologically: N. suae collected from submerged decaying wood in China (Li et al. 2022); N. bambusicola collected from dead culms of bamboo in Thailand (Tian et al. 2022); N. hainanensis collected from decaying wood in a terrestrial habitat in China (Lu et al. 2022); N. terrestris collected from a dead leaf petiole of Musa sp. in Thailand (Samarakoon et al. 2024); N. guineensis and N. xishuangbannaensis collected from rotting petioles of Elaeis guineensis in China (Xiong et al. 2024); and N. baihualingense, N. hainanense, N. jianfenglingense, and N. latisporum collected from freshwater and terrestrial habitats in China (Ma et al. 2024b).

To date, based on morphological and/or molecular data, Neohelicosporium contains 33 species, with four species (N. myrtacearum, N. nizamabadense, N. sympodiophorum, and N. vesiculiferum) lacking molecular data. Neohelicosporium species are widely distributed across freshwater and terrestrial habitats in Australia, Belgium, Brazil, Canada, China, Cuba, Germany, Honduras, India, Indonesia, Japan, New Zealand, Panama, Peru, the Solomon Islands, South Africa, Thailand, Venezuela, and the USA (Morgan 1892; Penzig 1897; Linder 1929; Moore 1954; Rao and Rao 1964; Matsushima 1971, 1980; Rossman 1979, 1987; Chouhan and Panwar 1980; Goos 1985, 1986, 1989; Ho et al. 2001; Tsui et al. 2001, 2006; Cai et al. 2006; Zhao et al. 2007; da Cruz et al. 2009; Promputtha and Miller 2010; Singh and Singh 2016; Jayasiri et al. 2017; Kuo and Goh 2018; Lu et al. 2018a, 2018b; Dong et al. 2020; Li et al. 2022; Tian et al. 2022; Yang et al. 2023; Zhang et al. 2023a, 2023b; Samarakoon et al. 2024; Ma et al. 2023, 2024b). Among them, 28 Neohelicosporium species exhibit a helicosporous asexual morph, 4 species display both asexual and sexual morphs, and one species (N. terrestris) produces only the sexual morph (Promputtha and Miller 2010; Jayasiri et al. 2017; Lu et al. 2018b; Samarakoon et al. 2024). The asexual morph of Neohelicosporium is characterized by white, light pink, or pale brown colonies; short and solitary conidiophores that are mostly branched and gregarious; mono- to polyblastic conidiogenous cells; and guttulate, hyaline, helicoid conidia (Lu et al. 2018a, 2018b; Dong et al. 2020; Li et al. 2022; Tian et al. 2022; Yang et al. 2023; Ma et al. 2024b). Its sexual morph is depicted by globose to subglobose, spherical or ellipsoidal-ovate, pale brown to yellow or dark brown to black ascomata, 8-spored or 2–4–6-spored, bitunicate asci, and fusiform, hyaline ascospores (Promputtha and Miller 2010; Jayasiri et al. 2017; Lu et al. 2018b; Samarakoon et al. 2024).

In this study, six helicosporous isolates representing two distinct species were collected from freshwater and terrestrial habitats in southern China. Based on comprehensive morphological descriptions and illustrations, as well as multi-gene phylogenetic analyses, two novel species, namely Neohelicosporium guizhouense and N. wuzhishanense, are introduced.

Materials and methods

Sample collection and specimen examination

Specimens were collected from freshwater and terrestrial habitats between December 2021 and April 2022 in Qiannan Buyi and Miao Autonomous Prefecture, Sandu Shui Autonomous County, Guizhou Province, and Ledong Li Autonomous County and Wuzhishan City, Hainan Province, China, and the important collection details were noted (Rathnayaka et al. 2024). Once the specimens were transported to the laboratory, the specimens from freshwater habitats were incubated at room temperature and maintained in a moist environment for 1–2 weeks. Fungal colonies on the surface of the natural substrates, including conidiophores, conidiogenous cells, and conidia, were examined using a stereomicroscope (SMZ-168, Nikon, Japan). Photographs were taken with an ECLIPSE Ni compound microscope (Nikon, Tokyo, Japan) equipped with a Canon 90D digital camera.

Isolation and material deposition

Single spore isolations were performed following the method described by Senanayake et al. (2020). The germinated coiled conidia were aseptically transferred onto fresh potato dextrose agar (PDA) plates as described by Senanayake et al. (2020). The fungal mycelia were cultured on PDA medium at a constant temperature of 25 °C for 33–41 days. During incubation, colony characteristics such as shape, color, size, margin, and elevation were systematically documented.

Dried fungal specimens were deposited in the Herbarium of Kunming Institute of Botany, Chinese Academy of Sciences (Herb. HKAS), Kunming, China, and the Herbarium of Guizhou Academy of Agriculture Sciences (Herb. GZAAS), Guiyang, China. Pure cultures were deposited at the Guizhou Culture Collection (GZCC), Guiyang, China. Descriptions of the new taxa were uploaded to the Faces of Fungi webpage, as per the guidelines provided by Jayasiri et al. (2015). The new species were registered in the MycoBank database (https://www.mycobank.org/), and MycoBank numbers were obtained.

DNA extraction, PCR amplification, and sequencing

Fresh mycelia were scraped using sterilized toothpicks. Genomic DNA was extracted using the Biospin Fungus Genomic DNA Extraction Kit (BioFlux, China), following the manufacturer’s protocol. Primer pairs LR0R/LR5 (Vilgalys and Hester 1990), ITS5/ITS4 (White et al. 1990), EF1-983F/EF1-2218R (Rehner and Buckley 2005), and fRPB2-5F/fRPB2-7cR (Liu et al. 1999) were used to amplify LSU, ITS, tef1-α, and rpb2 sequence fragments, respectively. The PCR amplification reactions were performed according to the protocol by Ma et al. (2024a). The PCR products were purified and sequenced by Qingke Biotechnology, Chongqing, China.

Phylogenetic analyses

Newly generated sequences were verified and assembled using BioEdit v. 7.0.5.3 (Hall 1999) and SeqMan v. 7.0.0 (Swindell and Plasterer 1997). Additional sequences used in this study were obtained from GenBank (Table 1; https://www.ncbi.nlm.nih.gov/). Alignments for the LSU, ITS, tef1-α, and rpb2 datasets were performed using MAFFT v. 7.473 (https://mafft.cbrc.jp/alignment/server/, Katoh et al. 2019), and were visually checked and converted to the nexus format using AliView v. 1.27 (Daniel et al. 2010; Larsson 2014). Subsequently, trimAl v. 1.2 software was used to trim each gene dataset (Capella-Gutiérrez et al. 2009). The LSU, ITS, tef1-α, and rpb2 datasets were then concatenated using SequenceMatrix v. 1.7.8 (Vaidya et al. 2011).

Table 1.
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List of taxa analyzed in this study, along with their corresponding GenBank accession numbers.

Taxon Strain GenBank Accession numbers
LSU ITS tef1–α rpb2
Acanthohelicospora aurea GZCC 16-0060 KY321326 KY321323 KY792600 MF589911
A. guianensis UAMH 1699 AY856891 AY916479 – –
Neohelicosporium abuense CBS 101688 – AY916470 – –
N. acrogenisporum MFLUCC 17-2019T MH558871 MH558746 MH550937 MH551069
N. aquaticum GZCC 23-0342 PP639461 PP626605 PP596362 PP596488
N. aquaticum MFLUCC 17-1519T MF467929 MF467916 MF535242 MF535272
N. astrictum MFLUCC 17-2004T MH558872 MH558747 MH550938 MH551070
N. aurantiellum GZCC 23-0135 PP639462 PP626606 – –
N. aurantiellum GZCC 23-0414 PP639463 PP626607 – –
N. aurantiellum ANM 718 GQ850485 GQ856140 – –
N. baihualingense CGMCC 3.25547T – PP626608 PP596363 PP596489
N. baihualingense GZCC 23-0235 PP639465 PP626609 PP596364 PP596490
N. bambusicola MFLUCC 21_0156T OL606146 OL606157 OL964517 OL964523
N. ellipsoideum GZCC 22-2072 PP639466 PP626610 PP596365 –
N. ellipsoideum MFLUCC 16-0229T MH558873 MH558748 MH550939 MH551071
N. fluviatile MFLUCC 15-0606T OP377957 – OP473050 OP473111
N. fluviatile HKUCC 10235 AY849942 – – –
N. fusisporum MFUCC 16-0642T MG017613 MG017612 MG017614 –
N. griseum CBS 961.69 AY856884 AY916474 – –
N. griseum CBS 113542 AY916088 AY916475 – –
N. griseum UAMH 1694 AY856902 AY916473 – –
N. griseum GZCC 23-0424 PP639467 PP626611 PP596366 –
N. griseum JCM 9265 AY856889 AY916476 – –
N. guangxiense GZCC 16-0042 MF467933 MF467920 MF535246 MF535276
N. guangxiense MFLUCC 17-0054 MH558875 MH558750 MH550941 MH551073
N. guangxiense GZCC 16-0067 MF467930 MF467917 MF535243 MF535273
N. guangxiense GZCC 16-0068 MH558874 MH558749 MH550940 MH551072
N. guangxiense GZCC 16-0077 MF467931 MF467918 MF535244 MF535274
N. guangxiense GZCC 16-0089 MF467932 MF467919 MF535245 MF535275
N. guangxiense MFLUCC 17-0050 MF467934 MF467921 MF535247 MF535277
N. guangxiense MFLUCC 17-1522T MF467935 MF467922 MF535248 MF535278
N. guangxiense CBS 257.59 AY916087 AY916471 – –
N. guangxiense GZCC 23-0520 PP639468 PP626612 PP596367 –
N. guineensis ZHKUCC 24-0113T PP860102 PP860090 PP858062 PP858074
N. guineensis ZHKUCC 24-0114 PP860103 PP860091 PP858063 PP858075
N. guizhouense GZCC 23-0023 PQ098524 PQ098487 PQ816252 PQ816247
N. guizhouense GZCC 23-0078T PQ098525 PQ098488 PQ816253 PQ816248
N. guizhouense GZCC 23-0545 PQ098526 PQ098489 PQ816254 PQ816249
N. hainanense CGMCC 3.25548T PP639469 PP626613 PP596368 PP596491
N. hyalosporum GZCC 16-0063 MH558876 MH558751 MH550942 MH551074
N. hyalosporum GZCC 16-0076T MF467936 MF467923 MF535249 MF535279
N. hyalosporum GZCC 22-2173 PP639470 PP626614 PP596369 PP596492
N. hyalosporum GZCC 22-2162 PP639471 PP626615 PP596370 PP596493
N. hyalosporum GZCC 23-0042 PP639472 – – –
N. hyalosporum GZCC 23-0226 PP639473 PP626617 PP596371 PP596494
N. hyalosporum GZCC 23-0138 PP639474 PP626618 PP596372 PP596495
N. irregulare MFLUCC 17-1796T MH558877 MH558752 MH550943 MH551075
N. irregulare MFLUCC 17-1808 MH558878 MH558753 MH550944 MH551076
N. jianfenglingense CGMCC 3.25566T PP639475 PP626619 PP596373 PP596496
N. krabiense MFLUCC 16-0224T MH558879 MH558754 MH550945 MH551077
N. latisporum CGMCC 3.25546T PP639476 PP626620 PP596374 PP596497
N. latisporum GZCC 22-2056 PP639477 PP626621 PP596375 –
N. latisporum GZCC 22-2089 PP639478 PP626622 PP596376 –
N. laxisporum GZCC 23-0224 PP639479 PP626623 – –
N. laxisporum MFLUCC 17-2027T MH558880 MH558755 MH550946 MH551078
N. morganii CBS 281.54 AY856876 AY916468 – –
N. morganii CBS 222.58 AY856880 AY916469 – –
N. ovoideum GZCC 16-0064T MH558881 MH558756 MH550947 MH551079
N. ovoideum GZCC 16-0066 MH558882 MH558757 MH550948 MH551080
N. parvisporum GZCC 16-0078 MF467937 MF467924 MF535250 MF535280
N. parvisporum GZCC 16-0100 MF467938 MF467925 MF535251 MF535281
N. parvisporum MFLUCC 16-0218 MF467940 MF467927 MH550955 MH551087
N. parvisporum MFLUCC 17-1523T MF467939 MF467926 MF535252 MF535282
N. parvisporum MFLUCC 17-1793 MH558884 MH558759 MH550950 MH551082
N. parvisporum MFLUCC 17-1804 MH558885 MH558760 MH550951 MH551083
N. parvisporum MFLUCC 17-1807 MH558886 MH558761 MH550952 MH551084
N. parvisporum MFLUCC 17-1995 MH558887 MH558762 MH550953 MH551085
N. parvisporum MFLUCC 17-2010 MH558888 MH558763 MH550954 MH551086
N. parvisporum GZCC 22-2158 PP639480 PP626624 PP596377 PP596498
N. parvisporum GZCC 23-0248 PP639481 PP626625 – –
N. parvisporum GZCC 23-0087 PP639482 PP626626 PP596378 PP596499
N. parvisporum MFLUCC 17-1521 MH558883 MH558758 MH550949 MH551081
N. suae CGMCC 3.23541T OP184068 OP184079 OP186052 –
N. submersum CBS 189.95 AY856882 AY916472 – –
N. submersum MFLUCC 17-2376T MN913738 MT627738 – –
N. submersum GZCC 23-0293 PP639483 PP626627 – –
N. taiwanense BCRC-FU30841T – LC316603 – –
N. terrestris MFLUCC 23-0234T PP800330 OR206384 OR206052 PP840926
N. thailandicum MFLUCC 16-0221T MF467941 MF467928 MF535253 MF535283
N. wuzhishanense GZCC 23-0278T PQ098523 PQ098486 PQ816251 –
N. wuzhishanense GZCC 23-0279 PQ098527 PQ098490 – –
N. wuzhishanense GZCC 23-0326 PQ098522 PQ098485 PQ816250 PQ816246
N. xishuangbannaensis ZHKUCC 24-0119T PP860104 PP860092 PP858064 PP858076
N. xishuangbannaensis ZHKUCC 24-0120 PP860105 PP860093 PP858065 PP858077

Note: Newly generated sequences are in bold black. A dash (“-”) indicates unavailable data in GenBank, and “T” represents ex-type strains.

The maximum likelihood (ML) tree was constructed using the IQ-Tree web server (http://iqtree.cibiv.univie.ac.at/, Nguyen et al. 2015). Bayesian inference (BI) was performed following the methods described by Ma et al. (2022). The best-fit substitution model of the LSU, ITS, tef1-α, and rpb2 datasets was determined using MrModeltest v. 2.3 under the Akaike Information Criterion (AIC) (Nylander et al. 2008).

Phylogenetic trees were visualized using FigTree v. 1.4.4 and edited with Adobe Illustrator CC 2019 v. 23.1.0 (Adobe Systems, USA). Photo-plates were prepared using Adobe Photoshop CC 2019 (Adobe Systems, USA), and measurements were made using the Tarosoft (R) Image Frame Work program v. 1.3.4.

Phylogenetic results

The phylogenetic placements of our six isolates were validated based on a multi-gene phylogenetic analysis incorporating ITS, LSU, tef1-α, and rpb2 sequence data. A total of 84 Tubeufiaceae strains, including isolates obtained in this study and two outgroups, Acanthohelicospora aurea (GZCC 16-0060) and A. guianensis (UAMH 1699), were analyzed. The concatenated sequence matrix contains 3,386 characters (LSU: 1–842, ITS: 843–1,429, tef1-α: 1,430–2,341, rpb2: 2,342–3,386). Fig. 1 illustrates the best-scoring RAxML tree, with a final likelihood value of -15233.387. Following the guidelines of Chethana et al. (2021), a biphasic approach incorporating both morphological and phylogenetic species concepts was employed to describe the new species.

Figure 1. 

Phylogenetic tree generated from the RAxML analysis based on the combined dataset of LSU, ITS, tef1-α, and rpb2 sequences. Bootstrap support values for ML (≥ 75%) and posterior probability values (PP) (≥ 0.95) are indicated near respective nodes. Acanthohelicospora aurea (GZCC 16-0060) and A. guianensis (UAMH 1699) were selected as outgroup taxa. Ex-type strains are indicated by “T”, and newly obtained isolates are in bold red font. A dash (“-”) indicates bootstrap support values below 75% for ML and PP values less than 0.95.

Our collections in our multi-gene phylogenetic tree (Fig. 1) represent two distinct novel Neohelicosporium species within Tubeufiaceae. Our isolates, GZCC 23–0078 and GZCC 23–0545, cluster together as a sister group to GZCC 23–0023, with 100% ML and 1.00 PP support. Additionally, GZCC 23–0278, GZCC 23–0279, and GZCC 23–0326 group together, forming a distinct lineage with N. guizhouense (GZCC 23–0023, GZCC 23–0078, and GZCC 23–0545), supported by 100% ML and 1.00 PP.

Taxonomy

Neohelicosporium guizhouense X.Y. Ma, J. Ma & Y.Z. Lu, sp. nov.

MycoBank No: 903462
Facesoffungi Number: FoF17243
Fig. 2

Etymology.

The epithet “guizhouense” refers to Guizhou Province, where the fungus was collected.

Holotype.

HKAS 128908

Description.

Saprobic on decaying wood in terrestrial habitats. Sexual morph Undetermined. Asexual morph Hyphomycetous, helicosporous. Colonies on natural substrate superficial, effuse, gregarious, white. Mycelium mostly immersed, partly superficial, composed of pale brown to brown, branched, septate, guttulate, smooth. Conidiophores 56.5–165 μm long, 4–6.5 μm wide (x̄ = 118.5 × 5.5 μm, n = 20), macronematous, mononematous, procumbent, aggregated, cylindrical, tapering towards the tip, straight or slightly flexuous, branched, septate, smooth- and thick-walled, brown at the base and hyaline towards the apex. Conidiogenous cells 9–16.5 μm long, 3.5–6 μm wide (x̄ = 12.5 × 4.5 μm, n = 30), holoblastic, mono- to poly-blastic, integrated, intercalary or terminal, hyaline, smooth-walled, cylindrical or subcylindrical, becoming truncate towards the apex after conidial secession with tiny tooth-like protrusions, mostly bearing one (rarely two) tiny conidiogenous loci. Conidia solitary, acropleurogenous, helicoid, tapering towards the ends, developing on tooth-like protrusions, 20.5–27.5 μm diam. and conidial filament 3.5–5 μm wide (x̄ = 23.5 × 4 μm, n = 30), 78–109.5 μm long (x̄ = 88 μm, n = 30), indistinctly multi-septate, slightly constricted at septa, tightly coiled up to 3 times, becoming loosely coiled or uncoiled in water, guttulate, hyaline, smooth-walled.

Figure 2. 

Neohelicosporium guizhouense (HKAS 128908, holotype) a, b Colonies on the natural substrate c–e Conidiophores and conidiogenous cells f–j Conidiogenous cells k–o Conidia p Germinated conidium q, r Surface and reverse view of colonies on PDA after 33 days of incubation at 25 °C. Scale bars: 50 μm (c); 40 μm (d, e); 10 μm (f–p).

Culture characteristics.

Conidia germinating on PDA within 13 h and germ tubes arising from the terminal ends and the middle of the conidium. Colonies reached 26 mm diam. after 33 days of incubation at 25 °C, irregular, with flat, brown to black mycelia on the surface, in reverse pale brown to black with undulate margin.

Material examined.

China • Guizhou Province, Qiannan Buyi and Miao Autonomous Prefecture, Libo County, Maolan Nature Reserve, on decaying wood in a terrestrial habitat, 10 April 2022, Jian Ma, MN4.1 (HKAS 128908, holotype), ex-type living culture GZCC 23-0078; China, Guizhou Province, Sandu Shui Autonomous County, Yao Renshan National Forest Park, on decaying wood in a terrestrial habitat, 10 April 2022, Jian Ma, YS17.4 (GZAAS 23-0550, paratype), living culture GZCC 23-0545; China, Hainan Province, Ledong Li Autonomous County, Jianfengling National Forest Park, on decaying wood in a forest, 28 December 2021, Jian Ma, JB2 (HKAS 128876, paratype), living culture GZCC 23-0023.

Notes.

In the phylogenetic tree (Fig. 1), three strains of Neohelicosporium guizhouense (GZCC 23-0078, GZCC 23-0023, and GZCC 23-0545) formed a distinct lineage and were sister to N. wuzhishanense (GZCC 23-0278, GZCC 23-0279, and GZCC 23-0326), supported by 100% ML and 1.00 PP. A comparison of LSU, ITS, tef1-α, and rpb2 sequences between the ex-type strain of N. guizhouense (GZCC 23-0078) and N. wuzhishanense (GZCC 23-0326) revealed 2/837 bp (0.2%, including one gap), 26/773 bp (3.4%, including one gap), 21/885 bp (2.4%, without a gap), and 35/914 bp (3.8%, including one gap) nucleotide base differences, respectively, which strongly support them as two distinct species. Morphologically, Neohelicosporium guizhouense (HKAS 128908) differs from N. wuzhishanense (HKAS 128903) by its distinct conidiogenous cells (tooth-like vs. tooth-like and/or bladder-like), smaller conidial filaments (3.5–5 μm vs. 4.5–6 μm), and different coiled states in water (loosely coiled vs. tightly coiled). Therefore, we designate the three isolates (GZCC 23-0023, GZCC 23-0078, and GZCC 23-0545) as a new species, Neohelicosporium guizhouense.

Neohelicosporium wuzhishanense X.Y. Ma, J. Ma & Y.Z. Lu, sp. nov.

MycoBank No: 903463
Facesoffungi Number: FoF17244
Fig. 3

Etymology.

The epithet “wuzhishanense” refers to Wuzhishan City, Hainan Province, where the fungus was collected.

Holotype.

HKAS 128903

Description.

Saprobic on submerged decaying wood in freshwater habitats. Sexual morph Undetermined. Asexual morph Hyphomycetous, helicosporous. Colonies on natural substrate superficial, effuse, solitary, scattered, or gregarious, white to pale brown. Mycelium mostly superficial, partly immersed, composed of pale brown to brown, branched, septate, guttulate, smooth, with mass glistening conidia. Conidiophores 75.5–203 μm long, 5.5–6.5 μm wide (x̄ = 134.5 × 6 μm, n = 25), macronematous, mononematous, erect, solitary, cylindrical, straight or slightly flexuous, occasionally branched, septate, smooth-walled, thick-walled, wider at the base and narrower towards the apex, and brown at the base, becoming hyaline to pale brown towards the apex. Conidiogenous cells 13–29 μm long, 3.5–5 μm wide (x̄ = 19 × 4.5 μm, n = 25), holoblastic, mono- to poly-blastic, integrated, intercalary or terminal, determinate, hyaline to brown, smooth-walled, cylindrical, truncate at the apex after conidial secession, with tiny tooth-like and/or bladder-like protrusions (7–16 μm long, 3–6 μm wide (x̄ = 10.5 × 4.5 μm, n = 15)). Conidia solitary, acropleurogenous, helicoid, tapering toward the ends, developing on tooth-like or bladder-like protrusions, 20.5–28.5 μm diam. and conidial filament 4.5–6 μm wide (x̄ = 25 × 5 μm, n = 30), 92.5–138 μm long (x̄ = 118 μm, n = 30), indistinctly multi-septate, slightly constricted at septa, tightly coiled 2½–3½ times, not becoming loose in water, guttulate, hyaline, smooth-walled.

Figure 3. 

Neohelicosporium wuzhishanense (HKAS 128903, holotype) a, b Colonies on the natural substrate c–e, k Conidiophores, conidiogenous cells, and attachment of conidia f–j Conidiogenous cells and attachment of conidia l–p Conidia q Germinated conidium r, s Surface and reverse view of colonies on PDA after 41 days of incubation at 25 °C. Scale bars: 50 μm (c); 40 μm (d, e); 10 μm (f–q).

Culture characteristics.

Conidia germinating on PDA within 18 h and germ tubes arising from the middle of the conidium. Colonies reached 37 mm diam. after 41 days of incubation at 25 °C, irregular, with raised, white to brown mycelia on the surface, in reverse brown to black-brown, with undulate margin.

Material examined.

China • Hainan Province, Wuzhishan City, Wuzhishan National Nature Reserve, on decaying wood in a freshwater stream, 28 December 2021, Jian Ma, WS68 (HKAS 128903, holotype), ex-type living culture GZCC 23-0326; • Ibid., WS19 (GZAAS 23-0282, paratype), living culture GZCC 23-0278; • Ibid., WS20 (GZAAS 23-0283), living culture GZCC 23-0279.

Notes.

Morphologically, Neohelicosporium wuzhishanense (HKAS 128903) closely resembles N. latisporum (HKAS 128955) in having macronematous, mononematous, erect, cylindrical, septate conidiophores; holoblastic, monoblastic, or polyblastic, integrated, cylindrical conidiogenous cells; and solitary, acrogenous, helicoid conidia (Ma et al. 2024b). However, based on multi-gene phylogenetic analyses, Neohelicosporium wuzhishanense formed a sister clade with N. guizhouense, which is phylogenetically distant from N. latisporum (Fig. 1). Morphologically, Neohelicosporium wuzhishanense (HKAS 128903) differs from N. latisporum (HKAS 128955) by its unique conidiogenous cells (tooth-like and/or bladder-like vs. tooth-like) and longer conidia (92.5–138 μm vs. 48.5–67.5) (Ma et al. 2024b). Based on morphological characteristics and DNA sequence data, we identified GZCC 23-0278, GZCC 23-0279, and GZCC 23-0326 as a new species, Neohelicosporium wuzhishanense.

Discussion

Including the newly described species (N. guizhouense and N. wuzhishanense), the genus Neohelicosporium comprises a total of 35 species (Samarakoon et al. 2024; Ma et al. 2024b). Among these, 15 species have been found in terrestrial habitats, 12 species in freshwater habitats, and eight species have been reported from both freshwater and terrestrial habitats (Morgan 1892; Penzig 1897; Linder 1929; Moore 1954; Rao and Rao 1964; Matsushima 1971, 1980; Rossman 1979, 1987; Chouhan and Panwar 1980; Goos 1985, 1986, 1989; Ho et al. 2001; Tsui et al. 2001, 2006; Cai et al. 2006; Zhao et al. 2007; da Cruz et al. 2009; Promputtha and Miller 2010; Singh and Singh 2016; Jayasiri et al. 2017; Kuo and Goh 2018; Lu et al. 2018a, 2018b; Dong et al. 2020; Li et al. 2022; Tian et al. 2022; Yang et al. 2023; Zhang et al. 2023a, 2023b; Samarakoon et al. 2024; Ma et al. 2023, 2024b). These species occur as saprobes on bamboo, Elaeis guineensis, Malvaceae sp., Musa sp., and decaying wood of unknown origin (Jayasiri et al. 2017; Lu et al. 2018a, b; Tian et al. 2022; Ma et al. 2024a; Samarakoon et al. 2024; Xiong et al. 2024).

Significant intra-species morphological variations have been observed within Neohelicosporium. For example, two collections (HKAS 128940 from submerged decaying wood in China and MFLU 17-1734 from submerged decaying wood in Thailand) represent the same species, Neohelicosporium submersum (Dong et al. 2020; Ma et al. 2024b). However, HKAS 128940 exhibits shorter conidiophores (38–77 µm) compared to MFLU 17-1734 (50–260 µm). Furthermore, the conidiogenous cells in HKAS 128940 are sympodial, a feature not observed in MFLU 17-1734 (Dong et al. 2020; Ma et al. 2024b). Similarly, Neohelicosporium laxisporum (MFLU 17-1107 and GZAAS 23-0228), collected from terrestrial habitats in China and Thailand, display notable morphological differences. GZAAS 23-0228 has wider conidiophores (4.5–8 μm vs. 3.5–5 μm) and wider conidiogenous cells (4.5–6 μm vs. 3–4 μm) than MFLU 17-1107 (Lu et al. 2018b; Ma et al. 2024b). Additionally, the conidiogenous cells of MFLU 17-1107 consist of tiny tooth-like protrusions, whereas GZAAS 23-0228 exhibits tiny tooth-like protrusions as well as bladder-like protrusions (Lu et al. 2018b; Ma et al. 2024b). These morphological variations are likely due to geographical differences.

The conidiophores and conidia of some Neohelicosporium species closely resemble those of Parahelicomyces and Tubeufia (Lu et al. 2018b; Li et al. 2022; Ma et al. 2024b). Neohelicosporium latisporum (HKAS 128955) resembles Parahelicomyces parvisporus (HKAS 128868) in having erect, solitary conidiophores and wider, tightly coiled, brown conidia (Jayasiri et al. 2017; Kuo and Goh 2018; Lu et al. 2018a, 2018b; Dong et al. 2020; Li et al. 2022; Tian et al. 2022; Yang et al. 2023; Samarakoon et al. 2024; Ma et al. 2023, 2024b). In addition, five Neohelicosporium species, namely N. ellipsoideum (GZAAS 22-2072), N. hainanense (HKAS 128921), N. jianfenglingense (HKAS 128914), N. suae (HKAS 124610), and N. submersum (HKAS 128940), morphologically resemble Tubeufia in having short, simple, erect, irregularly cylindrical conidiophores (Lu et al. 2017, 2018a, 2018b; Li et al. 2022; Ma et al. 2023, 2024b). These morphological similarities highlight the challenges of distinguishing genera based solely on morphology, emphasizing the importance of molecular and phylogenetic analyses for accurate identification and delineation.

Acknowledgments

We would like to thank Shaun Pennycook (Manaaki Whenua Landcare Research, New Zealand) for his valuable suggestions on the fungal nomenclature.

Additional information

Conflict of interest

The authors have declared that no competing interests exist.

Ethical statement

No ethical statement was reported.

Funding

This work was funded by the Guizhou Institute of Technology High-Level Talent Scientific Research Start-Up Fund, grant number 2023GCC066; the Moutai Institute High-Level Talent Scientific Research Start-Up Fund, grant number mygccrc[2022]013; and Guizhou Provincial Science and Technology Projects (Qiankehe Basic-ZK[2022] General 220).

Author contributions

Writing - original draft: JM, XYM, LH. Writing - review and editing: YZL, DDS.

Author ORCIDs

Yong-Zhong Lu https://orcid.org/0000-0002-1033-5782

Jian Ma https://orcid.org/0009-0008-1291-640X

Data availability

All of the data that support the findings of this study are available in the main text.

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