Nine Pezizomycotina strains were isolated from rotten dead branches and leaves collected from Guizhou Province. To obtain their accurate taxonomic placement, we provided the morphological characteristics of conidiophore cells and conidia. Phylogenetic relationships, based on ITS, rpb2, SSU, LSU and tub2 gene sequences, confirmed our strains represented three novel species, Peglionia falcata, Neoascochyta pseudofusiformis and Neomicrosphaeropsis cylindrica. Peglionia falcata produced falcate conidia and Neoa. pseudofusiformis generated fusiform conidia, while Neom. cylindrica possessed cylindrical conidia. The phylogenetic results also supported them as novel taxa. All the new species in the present study were found as saprophytic on forest litter with high rainfall, which suggest they may have a certain effect on nutrient decomposition and redistribution in forest ecosystems. Thus, it opened a way for further research on related ecological roles and their application production.

Ascomycota, morphology, phylogeny, taxonomy, three new taxa

Pezizomycotina is the largest subphylum of Ascomycota (Spatafora et al. 2006; Wijayawardene et al. 2022a), which is large and diverse (the 10^th edition of Ainsworth & Bisby’s Dictionary of the Fungi estimates close to 70,000 Pezizomycetes species) (Charley and David 2017). These taxa often exist as saprophytes fed by herbivore faeces or grow on woody and non-woody plant tissues. However, some can also be pathogenic to some plants and animals or symbiotic with some plants as endophytes (Money 2016). Fungal studies, related to ascomycetous taxa have been extensively carried out in China and many novel taxa are introduced annually (Hyde et al. 2016; Wijayawardene et al. 2022b).

The genus Peglionia (Goidànich 1934) is distinguished from Circinotrichum and Gyrothrix by the branching pattern of the setae (Hernández-Restrepo et al. 2022). According to Hernández-Restrepo et al., Gyrothrix verticiclada (CBS 127654 phenotype, CBS 140226 and CBS 148329) clustered with G. hughesii in the phylogenetic tree as a distinct branch, which were defined as a new genus (Peglionia) (Hernández-Restrepo et al. 2022). Peglionia verticiclada (type species) is characterised by the production of curved conidia and setae with verticillate and straight branches at the apex and never circinate as in Circinotrichum, Gyrothrix or other members of Xylariales. In terms of morphological characteristics, the conidiogenous cells of Peglionia vary from inverted clavate to lagenate, appearing hyaline or subhyaline. The conidia are attached to the tips of the conidiogenous cells, presenting a dry sickle shape, without septa and presenting hyaline (Hernández-Restrepo et al. 2022).

Neomicrosphaeropsis (Didymellaceae) was introduced by Thambugala et al. (2017) based on morphology and molecular data; meanwhile, Phoma tamaricicola were recollected and accommodated to Neomicrosphaeropsis. There are currently 11 epithets in the genus Neomicrosphaeropsis in Mycobank (www.mycobank.org), but Neom. cystisi, Neom. cystisicola, Neom. cytisina and Neom. minima have been transferred to Microsphaeropsis (Wijayawardene et al. 2022b). This genus contains some pathogens or endophytes (Wijayawardene et al. 2017). In terms of morphological features, the conidiophores of the genus Neomicrosphaeropsis appear pustulate, almost submerged in agar and slightly vesicular. The outer wall of the conidiophore ranges from light to dark brown. Conidiogenous cells are cylindrical, appear hyaline, have a smooth surface and are aggregated singly or in multiples. Conidia are hyaline to light brown, with a smooth outer wall and are ovate to ellipsoid (Thambugala et al. 2017).

Chen et al. (2015) introduced Neoascochyta to accommodate taxa that morphologically resemble to Ascochyta, but phylogenetically distinct. Neoascochyta belongs to the Didymellaceae family and species in this family are primarily parasitic on wood and dead herbaceous stems or leaves (Hyde et al. 2013). Currently, 20 Neoascochyta species have been listed in MycoBank database (2024). This genus is morphologically characterised by pseudothecial ascomata, cylindrical to subclavate asci, cylindrical to ovoid, hyaline 1-septate ascospores. The asexual morph is coelomycetous and is characterised by pycnidial conidiomata, pseudoparenchymatous wall, obpyriform or ampulliform to doliiform conidiogeneous cells and hyaline fusoid to cylindrical, obclavate-ovoid to ellipsoidal conidia (Chen et al. 2015).

The purpose of this study was to introduce three new Pezizomycotina taxa collected in Guizhou Province, viz. Peglionia falcata, Neoascochyta pseudofusiformis and Neomicrosphaeropsis cylindrica. The present study was of great significance to enrich the diversity of Pezizomycotina in southwest China on the basis of morphological description and phylogeny combined with ITS, LSU, tub2 and rpb2 sequence data analysis. Meanwhile, since all three new species identified are saprophytic fungi, which play an important role in the process of organic matter decomposition, they can be further studied for their ecological effects, which will provide an important theoretical and practical basis for relevant applied research and potential value exploration, based on their roles in natural ecosystems.

In the phylogenetic analyses, the MP, ML and Bayesian results obtained similar topologies, thus the ML topologies were edited and shown as Figs 1–3. For the Peglionia and related genera (Fig. 1), the combined data matrix of ITS–LSU–rpb2 consisted of 2366 characters (ITS: 761, LSU: 885 and rpb2: 720), amongst which 612 are parsimony informative characters. Maximum Parsimony analysis with the following parameters: TL = 2197; CI = 0.5294; HI = 0.4706; RI = 0.8182; and RC = 0.4331 indicated that Peglionia falcata strains (GUCC-0042, GUCC-0043 and GUCC-0044) without the DNA base differences in three loci formed an independent branch (ML = 100, MP = 100, PP = 1.00) and maintained a close relationship to P. verticiclada (CBS 127654, CBS 148329, CBS 101171 and CBS 140226) (ML = 100, MP = 99, PP = 1.00).

The combined data matrix of Neoascochyta (ITS–LSU–tub2) yielded 1784 characters (ITS: 489, LSU: 959 and tub2: 336). The MP analysis, based on 194 parsimony informative characters (1480 characters were constant and 110 variable characters), produced the phylogenetic tree with the following parameters: TL = 562; CI = 0.6975; HI = 0.3025; RI = 0.8932; and RC = 0.6230. The result (Fig. 2) displayed that Neoascochyta pseudofusiformis (GUCC23-0045, GUCC23-0046 and GUCC23-0047) formed an independent branch without the DNA base differences in three loci supported by strong statistic data (ML = 99, MP = 100, PP = 1.00) and were adjacent to the branch of Neoascochyta argentina CBS 112524 and N. tardicrescens (CBS 689.97) (ML = 75, MP = 87, PP = 0.65).

In Fig. 3, the combined data matrix of Neomicrosphaeropsis (ITS–LSU–SSU) including 2328 characters (ITS: 484, LSU: 833 and SSU: 1011), only had 17 parsimony informative characters. The MP analysis (TL = 51; CI = 0.8627; HI = 0.1373; RI = 0.9136; and RC = 0.7882) indicated the three strains of Neomicrosphaeropsis cylindrica (GUCC 23-0048, GUCC 23-0049 and GUCC 23-0050) as a branch without genetic distance (ML = 93, MP = 93.2, PP = 0.99) adjoining to the clade (ML = 86, MP = 60.3, PP = 0.99) including Neom. rossica, Neom. novorossica and Neom. italica. The phylogenetic placements of these novel taxa were also supported by DNA base-pair differences (Table 2).

Taxonomy

 Peglionia falcata S.M. Fu & Yong Wang bis, sp. nov.

MycoBank No: 854204

Facesoffungi Number: FoF15890

Fig. 4

Etymology

In reference to the fungus, which produced falcate conidia.

Diagnosis

Peglionia falcata is characterised by dry falcate meriform spores (24.1 × 2.9 μm; L/W = 8.005).

Figure 4. 

Peglionia falcata (GUCC23-0042) a, b appearance on host surface c, d culture characteristics on PDA (c above view d reverse view) e–m conidiophores, conidiogenous cells and conidia n conidia. Scale bars: 20 µm (e–n).

Type

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead branch, 19 July 2023, S.M. Fu, HGUP 23-0013 (holotype), ex-type culture GUCC23-0042.

Culture characteristics

Colonies on PDA, after 8 d, 20–25 mm diam., scarce aerial mycelium, dark brown, white to the periphery, margin entire, reverse dark brown. Occasionally, when a seta bears only two apical branches, one or both can be once forked. Conidiogenous cells polyblastic, obclavate to lageniform, hyaline to subhyaline. Conidia adherent in a continuous white layer on the conidiogenous cells, dry falcate, non-septate, hyaline. Chlamydospores (in culture) in chains, subglobose to irregularly-shaped, subhyaline to brown. Sexual morph not observed. Colonies hypophyllous, scattered, up to 1 mm wide, occasionally larger by confluence, velvety, black when sterile and whitish within when sporulating profusely. Conidiogenous cells obclavate to lageniform, hyaline to subhyaline, distally with a somewhat irregular contour, 5–16.5 × 4–7 µm (x̄ = 9.8 × 5.3 μm, n = 20). Conidia adherent in a continuous white layer on the conidiogenous cells, falcate, non-septate, hyaline,18–30 × 2.5–3.5 µm (x̄ = 24.1 × 2.9 μm, n = 30).

Habit

On rotten dead branches.

Distribution

China, Guizhou Province, Guiyang City

Other materials examined

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead branch, 19 July 2023, S.M. Fu, HGUP 23-0013, living culture GUCC23-0042, GUCC23-0043 and GUCC23-0044.

Notes

In morphology, Peglionia falcata differs to P. verticiclada by its larger conidiogenous cells (4–7 μm wide vs. 3–5 μm wide in P. verticiclada) and larger conidia (18–30 μm vs. 17–22 μm in P. verticiclada) (Hernández-Restrepo et al. 2022). The phylogenetic analyses and DNA base differences (Table 2) also supported P. falcata as a novel taxon was distinct from P. verticiclada.

 Neoascochyta pseudofusiformis S.M. Fu & Yong Wang bis, sp. nov.

MycoBank No: 854206

Facesoffungi Number: FoF15891

Fig. 5

Etymology

In reference to the fungus, which produced fusiform conidia morphologically similar to Neoascochyta fusiformis.

Figure 5. 

Neoascochyta pseudofusiformis (GUCC 23-0045) a, b appearance on host surface c colonies on host surface d–g culture characteristics on PDA, WA (d, f above view e, g reverse view) h colonies on WA i–k conidiophores, conidiogenous cells and conidia l conidia. Scale bars: 20 µm (i–l).

Diagnosis

Neoascochyta pseudofusiformis is characterised by oval to fusiform conidia (3.6 × 1.9; L/W = 1.896) with moderate growth rate.

Type

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead branch, 19 July 2023, S.M. Fu, HGUP 23-0014 (holotype), ex-type culture GUCC23-0045.

Culture characteristics

Colonies on PDA, 70–75 mm diam. after 7 d, margin regular, covered by floccose aerial mycelium, greyish-olivaceous, with flat and greenish-black flat mycelium near the margin; reverse black olivaceous. Mycelium is light to dark grey, separated, smooth, thin to thick wall. Acicular conidium, grey to dark grey, solitary or conjunctival, immersed in culture (WA), glabrous, subglobular, 100–250 × 90–130 μm, with a single pore neck; The angular textured cylindrical wall consists of 2 to 4 layers of flat polygonal cells 10–50 μm thick. The meristem cells are biparental, transparent, smooth-walled, pot or spherical, 3 × 5 μm wide. The conidia are 0–1 septum, transparent, smooth, thick-walled, mostly fusiform or slightly allantoic, 3.0–4.5 × 1.5–2.5 μm (x̄ = 3.6 × 1.9 μm, n = 30).

Habit

On rotten dead branches.

Distribution

China, Guizhou Province, Guiyang City

Other materials examined

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead branches, 19 July 2023, S.M. Fu, HGUP 23-0014, living culture GUCC23-0045, GUCC23-0046 and GUCC23-0047.

Notes

The present taxon differs morphologically to related species by conidial size range (3.0–4.5 × 1.5–2.5 μm vs. 16.5–27 × 5–8.5 μm in N. argentina and 2.5–3.5 × 1.0–1.5 μm in N. tardicrescens) (Chen et al. 2017; Valenzuela-Lopez et al. 2018). In phylogeny, our novel taxon maintained a close relationship to N. argentina CBS 112524 and N. tardicrescens CBS 689.97; however, DNA base differences (Table 2) supported that they belonged to different taxa.

 Neomicrosphaeropsis cylindrica S.M. Fu & Yong Wang bis, sp. nov.

MycoBank No: 854207

Facesoffungi Number: FoF15892

Fig. 6

Etymology

In reference to the fungus, which produced cylindrical conidia.

Diagnosis

Neomicrosphaeropsis cylindrica is characterised by broadly cylindrical conidia (15.4 × 3.4; L/W = 4.574) with moderate growth rate.

Figure 6. 

Neomicrosphaeropsis cylindrica (GUCC 23-0048) a appearance on host surface b–e culture characteristics on PDA, WA (b–d above view c–e reverse view) f–g colonies on PDA h–l conidiophores, Conidiogenous cells and Conidia m conidia. Scale bars: 100 µm (h); 50 µm (i–k); 20 µm (l–m).

Type

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead leaves, 19 July 2023, S.M. Fu, HGUP 23-0015 (holotype), ex-type culture GUCC23-0048.

Culture characteristics

Saprobic on dead leaves. Colony on PDA, 35–38 mm diameter, after 7 days, dense low-altitude hypha, light yellow, centre with abundant stigma; Turning light yellow to rose light yellow, the centre of concentric circles is darker; on MEA, after 7 days, 28–30 mm, the edge is intact, dense hypoxic mycelium, the edge is yellowish; reverse rose-yellowish to yellowish at margin with abundant scattered on stigma; Conidia cylindrical, spherical to kettle-shaped, 200–350 μm in diameter, tan to black, solitary, population centre abundant, banded, glabrous, without papillae; the cell wall is angular textured, light brown, bifid, cylindrical, thin-walled, transparent. Conidia occasionally septate, 12.5–18.5 × 2.4–4.0 μm (x̄ = 15.4 × 3.4 μm, n = 30), cylindrical, transparent, thin-walled.

Sexual stage

Not observed.

Habit

On rotten dead leaves.

Distribution

China, Guizhou Province, Guiyang City.

Other materials examined

China, Guizhou Province, Guiyang City, 26°57'N, 106°72'E, from rotten dead leaf, 19 July 2023, S.M. Fu, HGUP 23-0015, living culture GUCC23-0048, GUCC23-0049 and GUCC23-0050.

Notes

Neomicrosphaeropsis cylindrica (GUCC 23-0048) formed a clade with N. rossica (MFLUCC 14-0586) and N. alhagi-pseudalhagi (MFLUCC 17-0825) (Fig. 3). However, by morphological comparison, our species produced obviously longer conidia than N. rossica (12.5–18.5 × 2.4–4.0 μm vs. 4.4–5.7 × 2.9–3.9 μm) and smaller conidia than N. alhagi-pseudalhagi (12.5–18.5 × 2.4–4.0 μm vs. 30–45 × 18–22 μm) (Thambugala et al. 2017; Wanasinghe 2018).

According to Hernández-Restrepo et al. (2022), four strains of Peglionia verticiclada totally originated from decayed plant tissues in Europe and Australia. Our Peglionia species was also collected from rotten dead branches, which was the first discovery in China. In the database of Index Fungorum, Neoascochyta has 20 epithets and three of them were described by Chinese mycologists (Chen et al. 2017; Wang et al. 2024). Interestingly, the plant hosts of Neoascochyta spp. were mostly reported in the Gramineae family, such as Triticum sp., Hordeum sp. and Paspalum sp. (Chen et al. 2015, 2017; Hou et al. 2020). However, only two taxa (Neoascochyta soli and our N. pseudofusiformis) were both isolated from Guizhou, China and related to saprobic environments (Chen et al. 2017). In the test of inhibiting the germination of rust spores, the number of rust spots on leaves were significantly reduced after Neoascochyta treatment, which may provide a potential biological control method against rust diseases (Wilson et al. 2020). Neomicrosphaeropsis cylindrica was also the first species in Neomicrosphaeropsis to be discovered and described in China. This genus presented high correlations with alcohol and acids and was the highest contributors to the generation of volatile compounds, especially during alcohol production (Ma et al. 2021). Members of this genus were mostly obtained from Salicaceae as saprophytic fungi (Hyde et al. 2016; Thambugala et al. 2017). All our three Pezizomycotina taxa were isolated from rotten branches or leaves, which indicated that the diversity of this fungal group in Guizhou was relatively high. Thus, there was a need for a more comprehensive investigation.