Morphological and phylogenetic analyses reveal three new species of Apiospora in China

Abstract

Species of Apiospora are distributed worldwide as endophytes, pathogens and saprobes. In this study, we analysed Apiospora strains isolated from diseased leaves in Yunnan Province and dead culms in Shaanxi Province, China and we identified fungal species based on multi-locus phylogeny of ITS, LSU, tef1 and tub2 genes, along with the morphological characters, host and ecological distribution. Analyses revealed three new species, namely A.corylisp. nov., A.lophatherisp. nov. and A.oenotheraesp. nov. and one known species A.arundinis. Illustrations and descriptions of the four taxa are provided, along with comparisons with closely-related taxa in the genus.

Introduction

Species in Apiospora are distributed worldwide, primarily in temperate and tropical regions. These fungi can be found in various habitats, including soil, plant materials and insect exoskeletons (Pintos and Alvarado 2021). Many species of Apiospora are associated with plants as endophytic or saprophytic taxa and some can be important plant pathogens (Crous and Groenewald 2013; Wang et al. 2018; Kwon et al. 2021). In recent years, researchers have continuously discovered new Apiospora species in China (Wang et al. 2018; Senanayake et al. 2020, 2023; Feng et al. 2021; Liu et al. 2023).

Apiospora, the type genus of Apiosporaceae, was recognised and established by Saccardo (1875) with A.montagnei as the type species. For a long time, Apiospora was believed to be the sexual state of the genus Arthrinium (Ellis 1965; Samuels et al. 1981; Crous and Groenewald 2013). However, Ellis (1965) synonymised several other asexual genera with basauxic conidiogenesis under Arthrinium, such as Papularia, which was considered the asexual morph of Apiospora by von Höhnel (1919), Petrak (1925) and Hudson (1960, 1963). The asexual morph of Apiospora and Arthrinium are difficult to differentiate, based on morphology alone and the morphological relationships between Arthrinium and Apiospora have been hotly debated since Ellis (1965).

With the help of molecular phylogeny, Apiospora and Arthrinium were initially categorised in their own family Apiosporaceae (Hyde et al. 1998). Later, Crous and Groenewald (2013) considered that Apiospora was actually the sexual form of Arthrinium and both genera aligned to form a monophyletic clade. Following the principle of one fungi, one name policy (Hawksworth et al. 2011), the older name Arthrinium was recommended for use in unitary nomenclature (Réblová et al. 2016). However, due to several names with comparable sexual morphs to those of Arthrinium described as A.montagnei, the exact identity of A.montagnei remained uncertain (Hudson et al. 1976; Pintos et al. 2019; Pintos and Alvarado 2021). With the availability of sequence data of A.montagnei, Pintos and Alvarado (2022) revealed that Apiospora and Arthrinium are distinct genera. With most Apiospora species sharing similar morphologies, molecular phylogenetic information is necessary for accurate species identification (Pintos and Alvarado 2022).

The aim of the present study is to research new Apiospora samples found in western China, including one known species of A.arundinis and three new species and to describe them, based on morphological characters and phylogeny inferred from the combined ITS, LSU, tef1 and tub2 sequences dataset. To identify and compare these species with morphologically similar and phylogenetically related species, thorough analyses have been conducted.

Materials and methods

Sample collection and fungal isolation

Diseased leaves with dried dark brown spots of Oenotherabiennis and Lophatherumgracile, as well as diseased leaves with white round patches and black cracks of Brunfelsiabrasiliensis were collected from two locations in Yunnan Province: Lincang City (1547 m elevation; 23°52'12"N, 100°4'12"E) and Xishuangbanna City (763 m elevation; 22°1'48"N, 100°52'48"E). Dead plant culms of Corylusyunnanensis were collected in Ankang City (1683 m elevation; 33°26'37"N, 108°26'4"E), located in Shaanxi Province. All samples were placed in paper bags and transported to the laboratory for isolation. The samples were surface-sterilised by being exposed to 75% ethanol for one minute, followed by 1.25% sodium hypochlorite for three minutes, then another minute of exposure to 75% ethanol. The samples were then rinsed with distilled water for two minutes and dried on sterile filter paper. The affected portions of the leaves were excised into 0.5 × 0.5 cm fragments using a sterile razor blade. The fragments were then placed on to potato dextrose agar plates (PDA; containing 200 g potatoes, 20 g dextrose and 20 g agar per litre). The plates were incubated at a temperature of 25 °C to obtain pure cultures. All specimens were deposited at the Museum of Beijing Forestry University (BJFC) and all cultures were preserved at the China Forestry Culture Collection Center (CFCC).

Morphological observation

The morphology of the isolates was examined by analysing sporulating axenic cultures cultivated on PDA in darkness at 25 °C. After a 7-day incubation period, colony diameters were measured and colony characters were recorded. Slide mounts were prepared in lactic acid or water, obtained from colonies sporulating on PDA. Observations were conducted using a Leica DM 2500 dissecting microscope (Wetzlar, Germany) and a Nikon Eclipse 80i compound microscope, equipped with differential interference contrast (DIC) illumination. Images were captured with a Nis DS-Ri2 camera and processed using the Nikon Nis Elements F4.30.01 software. For measurement purposes, 50 conidiogenous cells and conidia were randomly selected. Conidial length was measured from the base of the basal cell to the base of the apical appendage, while conidial width was measured at its widest point. Taxonomic novelties were deposited in MycoBank (http://www.mycobank.org).

DNA extraction, PCR amplification and phylogenetic analyses

Genomic DNA was extracted from colonies grown on PDA using a cetyltrimethylammonium bromide (CTAB) method (Doyle and Doyle 1990). The extracted DNA products were stored at -20 °C until analysis. Four different loci were targeted for sequencing, including the nrDNA internal transcribed spacer regions 1 and 2 with the intervening 5.8S subunit (ITS), a partial sequence of the large subunit nrDNA subunit (LSU), a partial sequence of the translation elongation factor 1-alpha gene (tef1) and a partial sequence of the beta-tubulin gene (tub2). They were all amplified with the primer pairs and polymerase chain reaction (PCR) programme listed in Table 1.

Table 1.

Gene regions and respective primer pairs used in the study.

Locus	PCR primers	PCR: thermal cycles: (Annealing temperature in bold)	Reference
ITS	ITS1/ITS4	(94 °C: 30 s, 55 °C: 30 s, 72 °C: 45 s) × 35 cycles	White et al. 1990
LSU	LR0R/LR5	(94 °C: 30 s, 48 °C: 50 s, 72 °C: 1 min 30 s) × 35 cycles	Cubeta et al. 1991
tef1	EF1-728F/EF2	(95 °C: 30 s, 51 °C: 30 s, 72 °C: 1 min) × 35 cycles	O’Donnell et al. 1998; Carbone and Kohn 1999
tub2	Bt-2a/Bt-2b	(95 °C: 30 s, 56 °C: 30 s, 72 °C: 1 min) × 35 cycles	Glass and Donaldson 1995

The PCR products were assayed by electrophoresis in 2% agarose gels. Amplified PCR products were sent to a commercial sequencing provider (Tsingke Biotechnology Co. Ltd., Beijing, China). The quality of the chromatograms was verified and nucleotide sequences were assembled using SeqMan v.7.1.0. Reference sequences from related publications (Wang et al. 2018; Pintos and Alvarado 2021; Samarakoon et al. 2022; Liu et al. 2023) were retrieved from the National Center for Biotechnology Information (NCBI; https://www.ncbi.nlm.nih.gov). Sequences were aligned on the web server using MAFFT at the web server (http://mafft.cbrc.jp/alignment/server) (Katoh et al. 2019) and further corrected manually utilising MEGA 7.0.21 (Kumar et al. 2016).

The phylogenetic analyses of the combined loci were performed using Maximum Likelihood (ML) and Bayesian Inference (BI) methods. To implement ML, RAxMLHPC BlackBox 8.2.10 (Stamatakis 2014) was used on the CIPRES Science Gateway portal (https://www.phylo.org) employing a GTR GAMMA substitution model with a total of 1000 bootstrap replicates. The Bayesian posterior probabilities (BPP) were determined by Markov Chain Monte Carlo (MCMC) sampling in MrBayes v.3.2.6 (Ronquist et al. 2012). Six simultaneous Markov chains were run for 1 million generations starting from random trees, sampling trees every 100^th generation. To ensure accuracy, 25% of aging samples were discarded, running until the average standard deviation of the split frequencies dropped below 0.01. The phylogram was visualised in FigTree v.1.3.1 (http://tree.bio.ed.ac.uk/software) and edited using Adobe Illustrator CS5 (Adobe Systems Inc., USA). The newly-generated nucleotide sequences were deposited in GenBank (Table 2).

Table 2.

Isolates and GenBank accession numbers used in the phylogenetic analyses.

Species	Isolate/Strain	Host/ Substrate	Origin	GenBank accession numbers
				ITS	LSU	tef1	tub2
Apiosporaacutiapica	KUMCC 20-0210 (Type)	Bambusabambos	China	MT946343	MT946339	MT947360	MT947366
A.agari	KUC 21333 (Type)	Agarumcribrosum	Korea	MH498520	MH498440	MH544663	MH498478
A.aquatica	MFLU 18-1628 (Type)	Submerged wood	China	MK828608	MK835806	NA	NA
A.arctoscopi	KUC 21331 (Type)	Egg of Arctoscopusjaponicus	Korea	MH498529	MH498449	MN868918	MH498487
A.arundinis	CBS 10612	Unkown substrate	Germany	KF144883	KF144927	KF145015	KF144973
	LX 1918	Saccharumofficinarum	China	MW534386	NA	MW584370	MZ090019
	CFCC 58977	Brunfelsiabrasiliensis	China	OR125562	OR133584	OR139968	OR139976
	LS 107	Brunfelsiabrasiliensis	China	OR125563	OR133585	OR139969	OR139977
A.aurea	CBS 24483 (Type)	Air	Spain	AB220251	KF144935	KF145023	KF144981
A.balearica	CBS 145129 (Type)	Poaceae	Spain	MK014869	MK014836	MK017946	MK017975
A.bambusae	ICPM 6889 (Type)	Bamboo	China	MK014874	MK014841	MK017951	MK017980
A.bambusicola	MFLUCC 20-0144 (Type)	Schizostachyumbrachycladum	Thailand	MW173030	MW173087	MW183262
A.biserialis	CGMCC 320135 (Type)	Bamboo	China	MW481708	MW478885	MW522938	MW522955
A.camelliae-sinensis	LC 5007 (Type)	Camelliasinensis	China	KY494704	KY494780	KY705103	KY705173
A.chromolaenae	MFLUCC 17-1505 (Type)	Chromolaenaodorata	Thailand	MT214342	MT214436	MT235802	NA
A.chiangraiense	MFLUCC 21-0053 (Type)	Bamboo	Thailand	MZ542520	MZ542524	NA	MZ546409
A.cordylinae	GUCC 10027 (Type)	Cordylinefruticosa	China	MT040106	NA	MT040127	MT040148
A.coryli	CFCC 58978 (Type)	Corylusyunnanensis	China	OR125564	OR133586	OR139974	OR139978
	CFCC 58979	Corylusyunnanensis	China	OR125565	OR133587	OR139975	OR139979
A.cyclobalanopsidis	CGMCC 320136 (Type)	Cyclobalanopsidisglauca	China	MW481713	MW478892	MW522945	MW522962
A.descalsii	CBS 145130 (Type)	Ampelodesmosmauritanicus	Spain	MK014870	MK014837	MK017947	MK017976
A.dichotomanthi	LC 4950 (Type)	Dichotomanthustristaniaecarpa	China	KY494697	KY494773	KY705096	KY705167
A.dongyingensis	SAUCC 0302 (Type)	Bamboo	China	OP563375	OP572424	OP573264	OP573270
A.esporlensis	CBS 145136 (Type)	Phyllostachysaurea	Spain	MK014878	MK014845	MK017954	MK017983
A.euphorbiae	IMI 285638b	Bambusa	Bangladesh	AB220241	AB220335	NA	AB220288
A.fermenti	KUC21289 (Type)	Seaweed	Korea	MF615226	MF615213	MH544667	MF615231
A.gaoyouense	CFCC 52301 (Type)	Phragmitesaustralis	China	MH197124	NA	MH236793	MH236789
A.garethjonesii	JHB004 (Type)	Bamboo	China	KY356086	KY356091	NA	NA
A.gelatinosa	HKAS 111962 (Type)	Bamboo	China	MW481706	MW478888	MW522941	MW522958
A.guiyangensis	HKAS 102403 (Type)	Poaceae	China	MW240647	MW240577	MW759535	MW775604
A.guizhouensis	LC 5322 (Type)	Air in karst cave	China	KY494709	KY494785	KY705108	KY705178
A.hainanensis	SAUCC 1681 (Type)	Bamboo	China	OP563373	OP572422	OP573262	OP573268
A.hispanicum	IMI 326877 (Type)	Maritime sand	Spain	AB220242	AB220336	NA	AB220289
A.hydei	CBS 114990 (Type)	Bambusatuldoides	China	KF144890	KF144936	KF145024	KF144982
A.hyphopodii	MFLUCC 15-0003 (Type)	Bamboo	China	KR069110	NA	NA	NA
A.ibericum	AP 10118 (Type)	Arundodonax	Portugal	MK014879	MK014846	MK017955	MK017984
A.intestini	CBS 135835 (Type)	Gut of grasshopper	India	KR011352	MH877577	KR011351	KR011350
A.italicum	CBS 145138 (Type)	Arundodonax	Italy	MK014880	MK014847	MK017956	MK017985
A.jatrophae	CBS 134262 (Type)	Jatrophapodagrica	India	JQ246355	NA	NA	NA
A.jiangxiensis	LC 4577 (Type)	Maesa sp.	China	KY494693	KY494769	KY705092	KY705163
A.kogelbergensis	CBS 113333 (Type)	Restionaceae	South Africa	KF144892	KF144938	KF145026	KF144984
A.koreanum	KUC 21332 (Type)	Egg of Arctoscopusjaponicus	Korea	MH498524	MH498444	MH544664	MH498482
A.lageniformis	KUC 21686 (Type)	Phyllostachysnigra	Korea	ON764020	ON787759	ON806624	ON806634
A.locuta-pollinis	LC 11683 (Type)	Brassicacampestris	China	MF939595	NA	MF939616	MF939622
A.longistroma	MFLUCC 11-0481 (Type)	Bamboo	Thailand	KU940141	KU863129	NA	NA
A.lophatheri	CFCC 58975 (Type)	Lophatherumgracile	China	OR125566	OR133588	OR139970	OR139980
	CFCC 58976	Lophatherumgracile	China	OR125567	OR133589	OR139971	OR139981
A.malaysiana	CBS 102053 (Type)	Macarangahullettii stem colonised by ants	Malaysia	KF144896	KF144942	KF145030	KF144988
A.marianiae	AP18219 (Type)	Phleumpratense	Spain	ON692406	ON692422	ON677180	ON677186
A.marii	CBS 49790 (Type)	Atmosphere, pharmaceutical excipients, home dust and beach sands	Spain	MH873913	KF144947	KF145035	KF144993
A.marinum	KU 21328 (Type)	Seaweed	China	MH498538	MH498458	MH544669	MH498496
A.mediterranea	IMI 326875 (Type)	Air	Spain	AB220243	AB220337	NA	AB220290
A.minutisporum	17E-042 (Type)	Soil	Korea	LC517882	NA	LC518889	LC518888
A.montagnei	AP 301120 (Type)	Arundomicrantha	Spain	ON692408	ON692424	ON677182	ON67718
A.mori	MFLU 18-2514 (Type)	Morusaustralis	China	MW114313	MW114393	NA	NA
A.mukdahanensis	MFLUCC 22-0056 (Type)	Bambusoideae	Thailand	OP377735	OP377742	OP381089	NA
A.multiloculata	MFLUCC 21-0023 (Type)	Bambusae	Thailand	OL873137	OL873138	NA	OL874718
A.mytilomorpha	DAOM 214595 (Type)	Andropogon	India	KY494685	NA	NA	NA
A.neobambusae	LC 7106 (Type)	Bamboo	China	KY494718	KY494794	KY806204	KY705186
A.neochinensis	CFCC 53036 (Type)	Fargesiaqinlingensis	China	MK819291	NA	MK818545	MK818547
A.neogarethjonesii	HKAS 102408 (Type)	Bambusae	China	MK070897	MK070898	NA	NA
A.neosubglobosa	JHB007 (Type)	Bamboo	China	KY356090	KY356095	NA	NA
A.obovatum	LC4940 (Type)	Lithocarpus	China	KY494696	KY494772	KY705095	KY705166
A.oenotherae	CFCC 58972 (Type)	Oenotherabiennis	China	OR125568	OR133590	OR139972	OR139982
	LS 395	Oenotherabiennis	China	OR125569	OR133591	OR139973	OR139983
A.ovata	CBS 115042 (Type)	Arundinariahindsii	China	KF144903	KF144950	KF145037	KF144995
A.paraphaeosperma	MFLUCC13-0644 (Type)	Bambusa	Thailand	KX822128	KX822124	NA	NA
A.phragmitis	CBS 135458 (Type)	Phragmitesaustralis	Italy	KF144909	KF144956	KF145043	KF145001
A.phyllostachydis	MFLUCC 18-1101 (Type)	Phyllostachysheteroclada	China	MK351842	MH368077	MK340918	MK291949
A.piptatheri	CBS 145149 (Type)	Piptatherummiliaceum	Spain	MK014893	MK014860	MK017969	NA
A.pseudomarii	GUCC 10228 (Type)	Aristolochiadebilis	China	MT040124	NA	MT040145	MT040166
A.pseudohyphopodii	KUC 21680 (Type)	Phyllostachyspubescens	Korea	ON764026	ON787765	ON806630	ON806640
A.pseudoparenchymaticum	LC 7234 (Type)	Bamboo	China	KY494743	KY494819	KY705139	KY705211
A.pseudorasikravindrae	KUMCC 20-0208 (Type)	Bambusadolichoclada	China	MT946344	NA	MT947361	MT947367
A.pseudosinensis	CBS 135459 (Type)	Bamboo	Netherlands	KF144910	KF144957	KF145044	NA
A.pseudospegazzinii	CBS 102052 (Type)	Macarangahullettii	Malaysia	KF144911	KF144958	KF145045	KF145002
A.pterosperma	CPC 20193 (Type)	Lepidospermagladiatum	Australia	KF144913	KF144960	KF145046	KF145004
A.pusillisperma	KUC 21321 (Type)	Seaweed	Korea	MH498533	MH498453	MN868930	MH498491
A.qinlingense	CFCC 52303 (Type)	Fargesiaqinlingensis	China	MH197120	NA	MH236795	MH236791
A.rasikravindrae	NFCCI 2144 (Type)	Soil in karst cave	China	JF326454	NA	NA	NA
A.sacchari	CBS 21230	Phragmitesaustralis	Korea	KF144919	KF144965	KF145050	KF145008
A.saccharicola	CBS 19173	Air	Netherlands	KF144920	KF144966	KF145051	KF145009
A.sargassi	KUC21228 (Type)	Sargassumfulvellum	Korea	KT207746	KT207696	MH544677	KT207644
A.sasae	CBS 146808 (Type)	Sasaveitchii	Netherlands	MW883402	MW883797	MW890104	MW890120
A.septata	CGMCC 320134 (Type)	Bamboo	China	MW481711	MW478890	MW522943	MW522960
A.serenensis	IMI 326869 (Type)	Food, pharmaceutical excipients, atmosphere and home dust	Spain	AB220250	AB220344	NA	AB220297
A.setariae	CFCC 54041 (Type)	Setariaviridis	China	MT492004	NA	NA	NA
A.setostroma	KUMCC 19-0217 (Type)	Bambusoideae	China	MN528012	MN528011	MN527357	NA
A.sichuanensis	HKAS 107008 (Type)	Poaceae	China	MW240648	MW240578	MW759536	MW775605
A.sorghi	URM 93000 (Type)	Sorghumbicolor	Brazil	MK371706	NA	NA	MK348526
A.sphaerosperma	CBS114314 (Type)	Hordeumvulgare	Iran	KF144904	KF144951	KF145038	KF144996
A.stipae	CBS 146804 (Type)	Stipagigantea	Spain	MW883403	MW883798	MW890082	MW890121
A.subglobosa	MFLUCC 11-0397 (Type)	Bamboo	Thailand	KR069112	KR069113	NA	NA
A.subrosea	LC7292 (Type)	Bamboo	China	KY494752	KY494828	KY705148	KY705220
A.taeanensis	KUC21322 (Type)	Seaweed	Korea	MH498515	MH498435	MH544662	MH498473
A.thailandica	MFLUCC 15-0202 (Type)	Rotten wood	China	KU940145	KU863133	NA	NA
A.vietnamense	IMI 99670 (Type)	Citrussinensis	Vietnam	KX986096	KX986111	NA	KY019466
A.xenocordella	CBS 47886 (Type)	Soil from roadway	Zimbabwe	KF144925	KF144970	KF145055	KF145013
A.yunnana	MFLUCC 15-0002 (Type)	Bamboo	China	KU940147	KU863135	NA	NA
Arthriniumcrenatum	CBS 146353B (Type)	Grass	France	MW208931	MW208861	MW221917	MW221923

Notes: Strains in this study are marked in bold. NA = not available.

Results

Phylogeny

The combined ITS, LSU, tef1 and tub2 dataset comprised 99 strains, including eight newly-sequenced strains, with Arthriniumcrenatum (CBS 146353) as the outgroup taxon. Multi-locus sequences contain 2,709 characters including gaps with ITS (1–610), LSU (611–1399), tef1 (1400–1948) and tub2 (1949–2691). Of these characters, 1,635 were constant, 367 were variable and parsimony-uninformative and 707 were parsimony-informative. For ML analysis, the matrix had 1,192 distinct alignment patterns. Estimated base frequencies were A = 0.229212, C = 0.248907, G = 0.263837, T = 0.258044; substitution rates: AC = 1.129211, AG = 2.936388, AT = 0.925501, CG = 0.917970, CT = 4.199729, GT = 1.0; gamma distribution shape parameter: α = 0.250690; and likelihood value of ln: -22 496.696950.

The ML tree topology agreed with the BI analysis and, therefore, only the ML tree is presented (Fig. 1). The strains obtained in this study were categorised into four clades, representing one known species and three new species (Fig. 1). The known species is A.arundinis and three new species are now recognised as A.coryli, A.lophatheri and A.oenotherae.

Figure 1.

Phylogram of Apiospora, based on combined ITS, LSU, tef1 and tub2 genes. ML bootstrap support values (≥ 50%) and Bayesian posterior probability (≥ 0.90) are shown as first and second position above nodes, respectively. Strains from this study are shown in blue boxes, ex-type or ex-epitype cultures are indicated in bold face. Some branches were shortened according to the indicated mulipliers.

Taxonomy

. Apiospora arundinis

(Corda) Pintos & P. Alvarado, Fungal Systematics and Evolution 7: 205 (2021)

05626E62-5E3C-55EF-9D8A-04B58A926EA0

Fig. 2

Figure 2.

Apiosporaarundinis (CFCC 58977) A leaf of host plant B colony on PDA C conidiomata formed in culture D, E conidiogenous cells giving rise to conidia F conidia. Scale bars: 1000 µm (C); 10 µm (D–F).

Description.

Asexual morph: Mycelium consisting of smooth, hyaline, branched, septate, 1.1–5.9 µm diam. hyphae (n = 20). Conidiophores reduced to conidiogenous cells. Conidiogenous cells subglobose to ampulliform, erect, blastic, aggregated in clusters on hyphae, smooth, branched, 3.4–9.4 × 1.5–6.4 µm, mean (± SD): 6.8 (± 1.6) × 3.9 (± 1.3) µm (n = 50). Conidia globose, subglobose to lenticular, with a longitudinal germ slit, occasionally elongated to ellipsoidal, brown to dark brown, smooth to finely roughened, 6.4–10.4 × 5.2–8.3 µm, mean (± SD): 7.7 (± 0.6) × 6.8 (± 0.7) µm, L/W = 1.0–1.5 (n = 50). Sexual morph: Undetermined.

Culture characteristics.

On PDA, colonies thick and dense, margin undulate and irregular, pale yellow pigment diffused into medium, surface with patches of iron-grey aerial mycelia, reverse yellowish-brown, mycelia white to grey, sporulation on hyphae, reaching 9 cm in 7 days at 25 °C.

Specimens examined.

China, Yunnan Province: Xishuangbanna Botanical Garden, on diseased leaves of Brunfelsiabrasiliensis, 6 June 2022, S.J. Li, BJFC-S1918; living cultures CFCC 58977, LS 107).

Notes.

In this study, two isolates clustered together with the culture of A.arundinis with high-support values (ML/BI = 100/0.99)in the multi-locus phylogenetic tree (Fig. 1). Thus, these isolates were identified as A.arundinis and Brunfelsiabrasiliensis as a new host record for this species. Apiosporaarundinis was introduced from Phyllostachyspraecox, Castaneamollissima and Saccharumofficinarum in China (Chen et al. 2014; Jiang et al. 2021; Liao et al. 2022). Comparing with the description from Chen et al. (2014) (5–7 × 2–4 µm), Jiang et al. (2021) (3–4 µm) and Liao et al. (2022) (4.5–7.4 × 3.3–4.4 µm), the conidia in this study show larger sizes (6.4–10.4 × 5.2–8.3 µm). These differences may result from different host and habitat.

. Apiospora coryli

S.J. Li & C.M. Tian sp. nov.

C96D4D20-1AAE-5841-8C28-0671DF4611BB

849126

Fig. 3

Figure 3.

Apiosporacoryli (CFCC 58978, ex-holotype culture) A leaf of host plant B colony on PDA C conidiomata formed in culture D, E conidiogenous cells giving rise to conidia F conidia. Scale bars: 1000 µm (C); 10 µm (D–F).

Type.

China, Shanxi Province: Ankang City, Huoditang Forest Farm, on dead plant culms of Corylusyunnanensis, 16 July 2021, R. Yuan & S.J. Li, holotype BJFC-S1920, ex-type living cultures CFCC 58978, CFCC 58979.

Etymology.

Named after the host from which it was isolated.

Description.

Asexual morph: Derived from sporulated cultures on PDA, hyphae hyaline, branched, septate, 1.1–5.2 µm diam. Conidiophores reduced to conidiogenous cells. Conidiogenous cells erect, aggregated in clusters on hyphae, hyaline to pale brown, smooth, doliiform to clavate or lageniform, 2.6–10.6 × 2.1–5.8 µm, mean (± SD): 5.5 (± 2.4) × 3.4 (± 1.1) µm (n = 50). Conidia brown to dark brown, globose to subglobose, oval or irregular, smooth to finely roughened, guttulate, usually with a longitudinal germ slit, 7.4–18.4 × 6.2–12.5 µm, mean (± SD): 10.8 (± 1.7) × 9.4 (± 1.3) µm, L/W = 0.8–1.6 (n = 50). Sexual morph: Undetermined.

Culture characteristics.

On PDA, colonies circular, flat, entire margin, thick and cottony, concentrically spreading with aerial mycelium, margin regular, reddish-brown pigment diffused into medium, surface dark yellowish-brown, reverse dark reddish-brown to yellowish-brown from the centre, mycelia white to pale umber, sporulation on hyphae, reaching 9 cm in 7 days at 25 °C.

Notes.

Strains of A.coryli constitutes a distinct clade, but there is poor support value in concatenated gene trees (Fig. 1). The most prominent distinguishing characteristic is the production of reddish-brown pigments on the culture medium.

. Apiospora lophatheri

S.J. Li & C.M. Tian sp. nov.

2CD5427F-4DD4-56BE-BEC8-CEC89EE63775

849123

Fig. 4

Figure 4.

Apiosporalophatheri (CFCC 58975, ex-holotype culture) A leaf of host plant B colony on PDA C conidiomata formed in culture D conidiogenous cells giving rise to conidia E, F conidia. Scale bars: 1000 µm (C); 10 µm (D–F).

Type.

China, Yunnan Province, Xishuangbanna Primeval Forest Park, on diseased leaves of Lophatherumgracile, 4 June 2022, S.J. Li, holotype BJFC-S1917; ex-type living cultures CFCC 58975, CFCC 58976.

Etymology.

Named after the host from which it was isolated.

Description.

Asexual morph: Sporulated on PDA, mycelium consisting of hyaline, smooth, branched, septate hyphae 1.0–5.2 µm in diam. (n = 20). Conidiophores reduced to conidiogenous cells. Conidiogenous cells aggregated in clusters on hyphae, hyaline to pale brown, smooth, doliiform, clavate to ampulliform, 2.2–11.9 × 2.2–4.9 µm, mean (± SD): 6.4 (± 2.5) × 3.4 (± 0.6) µm (n = 50). Conidia globose, subglobose to lenticular, with a longitudinal germ slit, olive to dark brown, smooth to finely roughened and two or more conidia are produced on each conidiogenous cell, 5.1–8.9 × 4.6–7.7 µm, mean (± SD): 6.5 (± 0.8) × 5.9 (± 0.7) µm, L/W = 1.0–1.4 (n = 50). Sexual morph: Undetermined.

Culture characteristics.

On PDA, colonies flat, spreading, margin circular, thick, concentrically spreading with aerial mycelium, surface light greyish-brown, reverse tawny pigment diffused in media, mycelia white to grey and pale brown, sporulation on hyphae, reaching 9 cm in 7 days at 25 °C.

Notes.

Phylogenetic analysis indicated that Apiosporalophatheri is closely related to a clade comprising A.chromolaenae, A.euphorbiae, A.italicum, A.malaysiana, A.phyllostachydis, A.thailandica and A.vietnamense (Fig. 1). We compared the new species with phylogenetically similar taxa, based on morphological differences (Table 3) and base pair differences (Table 4). A.lophatheri can be differentiated from A.chromolaenae by its wider conidiogenous cells (2.2–11.9 × 2.2–4.9 µm vs. 6.5–12 × 1–2 µm) (from Euphorbia sp.; collected in Zambia; Ellis (1965)) and by 18 gene base pair differences (17/529 in ITS, 1/838 in LSU). A.lophatheri differs from A.euphorbiae by its larger olive to dark brown conidia (5.1–8.9 × 4.6–7.7 µm vs. 4–5.5 × 3–4 µm) (from Euphorbia sp.; collected in Zambia; Ellis (1965)), with nucleotide differences in ITS as 3/529, in LSU as 2/318, in tub2 as 22/801. A.italicum has smaller conidia (4–6 × 3–4 µm) (from Arundodonax; collected in Italy; Pintos et al. (2019)) and has 125 nucleotides differences (41/552 in ITS, 2/828 in LSU, 27/432 in tef1, 55/838 in tub2). Additionally, A.lophatheri is distinguished from A.malaysiana by having larger globose or subglobose conidia (5.1–8.9 × 4.6–7.7 µm vs. 5–6 × 3–4 µm) (from Macarangahullettii; collected in Malaysia; Crous and Groenewald (2013)), with 43 nucleotide differences (3/529 in ITS, 1/838 in LSU, 18/424 in tef1, 21/801 in tub2). A.lophatheri differs from A.phyllostachydis by its relatively shorter conidiogenous cells (2.2–11.9 × 2.2–4.9 µm vs. 20–55 × 1.5–2.5 µm) (from Phyllostachysheteroclada; collected in China; Yang et al. (2019)) and by 48 nucleotides differences (7/529 in ITS, 3/838 in LSU, 12/424 in tef1, 26/795 in tub2). A.lophatheri can be differentiated from A.thailandica by having shorter conidiogenous cells (2.2–11.9 × 2.2–4.9 µm vs. 11.5–39 × 2–3.5 µm) (from bamboo; collected in Thailand; Dai et al. (2017)) and by 12 nucleotides differences (9/529 in ITS, 3/828 in LSU). The conidia of A.lophatheri are significantly wider and paler-coloured than those of A.vietnamense (5.1–8.9 × 4.6–7.7 µm vs. 5–6 × 3–4 µm) (from Citrussinensis; collected in Vietnam; Wang et al. (2018)) and there are 7 nucleotides differences between the two species (2/526 in ITS, 2/803 in LSU, 3/315 in tub2). Therefore, A.lophatheri is described as a new species, based on phylogeny and morphological comparison.

Table 3.

Summary of morphology of new Apiospora species and phylogenetic related species.

Species	Isolation source	Country	Conidiogenous cells (µm)	Conidia in surface view		Conidia in side view		References
				Shape	Diam (μm)	Shape	Diam (μm)
A.gaoyouense	Phragmitesaustralis	China	1–2 × 2–3	globose to elongate ellipsoid	5–8	lenticular	4–8	Jiang et al. (2018)
A.hispanicum	Maritime sand	Spain	–	globose to ellipsoid	7.5–8.5 × 6–7.5	lenticular	6.5	Larrondo (1992)
A.locuta-pollinis	Brassicacampestris	China	3–7.5 × 3–6	globose to elongate ellipsoid	8–15× 5–9.5	–	–	Zhao et al. (2018)
A.longistroma	Bamboo	Thailand	–	asexual morph: Undetermined	–	–	–	Dai et al. (2017)
A.marii	Beach sand/ Poaceae	Spain	5–10 × 3–4.5	globose to elongate ellipsoid	8–10(−13)	lenticular	(5–)6(−8)	Crous and Groenewald (2013)
A.mediterranei	Airborn spore/ grass	Spain	–	lentiform	9–9.5 × 7.5–9	–	–	Larrondo (1992)
A.oenotherae	Oenotherabiennis	China	2.0–14.2 × 1.1–4.9	globose, subglobose to lenticular	6.6–13.9 × 5.5–10.1	–	–	This study
A.piptatheri	Piptatherummiliaceum	Spain	6–27 × 2–5	globose to elongate ellips oid	6–8 × 3–5	lenticular	4.5–6	Pintos et al. (2019)
A.pseudomarii	Aristolochiadebilis	China	8–13 × 2.5–5	subglobose to ellipsoid	6–9 × 4.5–6	–	–	Chen et al. (2021)
A.chromolaenae	Chromolaenaodorata	Thailand	6.5–12 × 1–2	elongated, broadly fliform to ampulliform	4–6×4.5–6.5	–	–	Mapook et al. (2020)
A.euphorbiae	Bambusa	Bangladesh	–	circular or nearly circular	(4–)4.7(–5.5)	lenticular	(3–)3.2(–4)	Sharma et al. (2014)
A.italicum	Arundodonax	Italy	(3–)4–7(–9) × (1.5–)2–3(–5)	globose	4–6×3–4	lenticular	–	Pintos et al. (2019)
A.lophatheri	Lophatherumgracile	China	2.2–11.9 × 2.2–4.9	globose, subglobose to lenticular	5.1–8.9 × 4.6–7.7	–	–	This study
A.malaysiana	Macarangahullettii	Malaysia	4–7 × 3–5	globose	5–6	lenticular	3–4	Crous and Groenewald (2013)
A.phyllostachydis	Phyllostachysheteroclada	China	20–55 × 1.5–2.5	globose to subglobose, oval or irregular	5–6 × 4–6	–	–	Yang et al. (2019)
A.thailandicum	Bamboo	Thailand	11.5–39 × 2–3.5	globose to subglobose, elongated to ellipsoidal	5–9 × 5–8	–	–	Dai et al. (2017)
A.vietnamense	Citrussinensis	Vietnam	4–7 × 3–5	globose	5–6	lenticular	3–4	Wang et al. (2017)

Table 4.

DNA base differences comparing Apiosporalophatheri sequences and sequences from related species.

Taxa	Loci	Nucleotides difference without gaps	Rates of base pair differences
A.chromolaenae	ITS	17/529 (40, 102, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122)	3.21%
	LSU	1/838 (426)	0.12%
A.euphorbiae	ITS	3/515 (26, 88, 89)	0.58%
	LSU	2/318 (146, 306)	0.63%
	tub2	22/801 (95, 96, 123, 151, 154, 163, 166, 182, 185, 193, 216, 237, 312, 347, 372, 429, 453, 454, 474, 559, 569, 574)	2.75%
A.italicum	ITS	41/552 (40, 82, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 132, 165, 177, 180, 205, 207, 213, 487, 529)	7.43%
	LSU	2/828 (406, 416)	0.24%
	tef1	27/432 (16, 18, 19, 20, 21, 22, 23, 24, 25, 27, 35, 46, 53, 60, 75, 80, 90, 102, 119, 123, 125, 172, 210, 211, 240, 248, 272)	6.25%
	tub2	55/838 (5, 29, 44, 45, 46, 92, 99, 119, 121, 122, 126, 155, 157, 171, 185, 188, 193, 194, 196, 198, 202, 297, 219, 229, 240, 265, 315, 338, 358, 363, 367, 368, 382, 384, 386, 390, 403, 407, 412, 430, 434, 454, 463, 465, 467, 480, 491, 499, 502, 556, 564, 580, 642, 756, 757)	6.56%
A.malaysiana	ITS	3/529 (40, 102, 103)	0.57%
	LSU	1/838 (426)	0.12%
	tef1	18/424 (15, 16, 19, 27, 29, 38, 52, 56, 82, 83, 91, 93, 95, 111, 115, 202, 203, 264)	4.25%
	tub2	21/801 (95, 96, 123, 151, 154, 163, 166, 182, 185, 193, 216, 237, 312, 347, 372, 429, 453, 474, 559, 569, 574)	2.62%
A.phyllostachydis	ITS	7/529 (40, 44, 85, 102, 106, 433, 500)	1.32%
	LSU	3/838 (7,8,9)	0.36%
	tef1	12/424 (16, 19, 26, 27, 51, 52, 53, 111, 197, 202, 203, 264)	2.83%
	tub2	26/795 (35, 52, 55, 84, 89, 112, 116, 147, 151, 175, 178, 186, 209, 211, 231, 329, 352, 354, 360, 462, 469, 489, 570, 572, 575, 608)	3.27%
A.thailandicum	ITS	9/529 (40, 82, 102, 107, 122, 175, 177, 183, 501)	1.70%
	LSU	3/828 (5, 416, 434)	0.36%
A.vietnamense	ITS	2/526 (37, 99)	0.38%
	LSU	2/803 (237, 391)	0.25%
	tub2	3/315 (72, 82, 87)	0.95%

. Apiospora oenotherae

S.J. Li & C.M. Tian sp. nov.

4E48C012-D878-50A9-B0E6-70877ECBD3A5

849125

Fig. 5

Figure 5.

Apiosporaoenotherae (CFCC 58972, ex-holotype culture) A leaf of host plant B colony on PDA C conidiomata formed in culture D, E conidiogenous cells giving rise to conidia F conidia. Scale bars: 1000 µm (C); 10 µm (D–F).

Type.

China, Yunnan Province, Lincang City Triangle Plum Garden, on diseased leaves of Oenotherabiennis, 26 April 2022, S.J. Li, holotype BJFC-S1919, ex-type living cultures CFCC 58972, LS 395.

Etymology.

Named after the host from which it was isolated.

Description.

Asexual morph: Hyphae hyaline, branched, septate, 1.2–4.8 µm in diam. (n = 20). Conidiophores reduced to conidiogenous cells. Conidiogenous cells smooth, ampulliform to doliiform, 2.0–14.2 × 1.1–4.9 µm, mean (± SD): 5.4 (± 2.9) × 3.1 (± 1.1) µm (n = 50). Conidia globose, subglobose to lenticular, with a longitudinal germ slit, occasionally elongated to ellipsoidal, colourless to dark brown, smooth to finely roughened, 6.6–13.9 × 5.5–10.1 µm, mean (± SD): 8.9 (± 1.2) × 7.8 (± 1.1) µm, L/W = 1.0–1.5 (n = 50). Sexual morph: Undetermined.

Culture characteristics.

On PDA, colonies thick, concentrically spreading with aerial mycelium, circular, margin irregular, yellow to pale green pigment diffused into medium, surface with aerial mycelia, the reverse lightly pigmented with a few dark yellow patches, mycelia white to grey, sporulation occurs after 10 days, reaching 9 cm in 7 days at 25 °C.

Notes.

Apiosporaoenotherae belongs to the large clade, where it shows a relationship with A.gaoyouense, A.hispanicum, A.locuta-pollinis, A.longistroma, A.marii, A.mediterranei, A.piptatheri and A.pseudomarii (Fig. 1), but differs in distinct morphological characters (Table 3) and nucleotide differences (Table 5). A.oenotherae differs from A.gaoyouense by its production of significantly conidiogenous cells (2.0–14.2 × 1.1–4.9 µm vs. 1–2 × 2–3 μm) (from Phragmitesaustralis; collected in China; Jiang et al. (2018)) and the presence of 30 distinct nucleotide positions (9/583 in ITS, 12/413 in tef1, 9/784 in tub2). A.oenotherae is distinct from A.hispanicum in producing larger conidial cells (6.6–13.9 × 5.5–10.1 µm vs. 7.5–8.5 × 6.2–7.6 µm) (from maritime sand; collected in Spain; Larrondo and Calvo (1992)) and in 30 nucleotides differences (1/539 in ITS, 1/320 in LSU, 28/796 in tub2). A.oenotherae differs from A.locuta-pollinis by its production of significantly conidiogenous cells (2.0–14.2 × 1.1–4.9 µm vs. 3–7.5 × 3–6 μm) (from hive-stored pollen; collected in China; Zhao et al. (2018)) and by the presence of 19 distinct nucleotide positions (1/539 in ITS, 7/416 in tef1, 11/485 in tub2). A.longistroma can be distinguished by growth rate, growing slowly on PDA, reaching 60 mm in 4 weeks (from bamboo; collected in Thailand; Dai et al. (2017)) and by the presence of 8 distinct nucleotide positions (6/572 in ITS, 2/840 in LSU). Moreover, A.mari produces elongated cells intermingled amongst conidia (from beach sand; collected in Spain; Crous and Groenewald (2013)), but A.oenotherae does not and can be distinguished by the presence of 23 distinct nucleotide positions (1/539 in ITS, 10/414 in tef1, 12/787 in tub2). Strains of A.mediterranei were isolated from pharmaceutical excipient, air-borne and on grass in Spain, while those of A.oenotherae collected from Oenotherabiennis in China. There are no discernible morphological characters distinguishing these species, but the elongated stem branches and the presence of 30 distinct nucleotide positions (1/539 in ITS, 1/320 in LSU, 28/796 in tub2) serve as clear indicators of their distinct and phylogenetically well-separated taxa. A.oenotherae differs from A.piptatheri because of its wider conidial cells (6.6–13.9 × 5.5–10.1 µm vs. 6–8 × 3–5 μm) (from Piptatherummiliaceum; collected in Spain; Pintos et al. (2019)) and the presence of 14 distinct nucleotide positions (10/528 in ITS, 4/827 in LSU). It also differentiates from A.pseudomarii through the production of notably wider conidial cells (6.6–13.9 × 5.5–10.1 µm vs. 6–9 × 4.5–6 µm) and through 12 unique nucleotide positions (5/556 in tef1, 7/416 in tub2) (from Aristolochiadebilis; collected in China; Chen et al. (2021)).

Table 5.

DNA base differences comparing Apiosporaoenotherae sequences and sequences from related species.

Taxa	Loci	Nucleotides difference without gaps	Rates of base pair differences
A.gaoyouense	ITS	9/583 (9, 10, 22, 36, 533, 535, 544, 555, 557)	1.54%
	tef1	12/413 (34, 48, 56, 57, 69, 90, 122, 129, 134, 170, 226, 228)	2.91%
	tub2	9/784 (538, 760, 766, 767, 768, 771, 775, 781, 782)	1.15%
A.hispanicum	ITS	1/539 (528)	0.19%
	LSU	1/320 (13)	0.31%
	tub2	28/796 (30, 186, 539, 761, 766, 767, 768, 769, 770, 771, 772, 773, 774, 775, 776, 777, 778, 779, 780, 781, 782, 783, 784, 785, 786, 787, 792, 794)	3.52%
A.locuta-pollinis	ITS	1/539 (528)	0.19%
	tef1	7/416 (33, 38, 94, 173, 177, 212, 258)	1.68%
	tub2	11/485 (237, 459, 465, 466, 467, 470, 474, 480, 481, 483, 485)	2.27%
A.longistroma	ITS	6/572 (20, 30, 38, 177, 213, 530)	1.05%
	LSU	2/840 (655, 825)	0.24%
A.marii	ITS	1/539 (528)	0.19%
	tef1	10/414 (35, 49, 57, 58, 91, 123, 135, 171, 227, 229)	2.42%
	tub2	12/787 (30, 186, 539, 761, 767, 768, 769, 772, 776, 782, 783, 785, 787)	1.52%
A.mediterranei	ITS	1/539 (528)	0.19%
	LSU	1/320 (13)	0.31%
	tub2	28/796 (30, 186, 539, 761, 766, 767, 768, 769, 770, 771, 772, 773, 774, 775, 776, 777, 778, 779, 780, 781, 782, 783, 784, 785, 786, 787, 792, 794)	3.52%
A.piptatheri	ITS	10/528 (30, 38, 142, 177, 182, 213, 420, 421, 430, 431)	1.89%
	LSU	4/827 (417, 431, 480, 632)	0.48%
A.pseudomarii	ITS	5/556 (425, 528, 541, 560, 561)	0.90%
	tef1	7/416 (33, 38, 94, 173, 177, 212, 258)	1.68%
	tub2	1/718 (520)	0.14%

Discussion

Apiospora has been revised using different approaches and its taxonomy and classification have changed several times since its introduction. The taxonomic classification of the genus in relation to Arthrinium has been a topic of debate (Crous and Groenewald 2013; Pintos and Alvarado 2021). Morphologically, Apiospora and Arthrinium share similarities in basauxic conidiogenesis. The conidia of Apiospora are typically lenticular or obovoid in the side view, with colours ranging from pale brown to brown. Conversely, the conidia of Arthrinium exhibit various shapes, such as angular, curved, fusiform, globose, navicular and polygonal (Kunze 1817; Hyde et al. 1998; Wang et al. 2018; Pintos and Alvarado 2021).

Recently, several revisions have been made in the course of unitary nomenclature resulting in the discovery of a plethora of new species, based on multigene phylogenies (Kwon et al. 2021; Pintos and Alvarado 2021, 2022; Liu et al. 2023). Currently there are 93 accepted species in Apiospora (Table 2), which are found on a wide range of materials.

In this study, A.arundinis and A.lophatheri were collected from the tropical region of Xishuangbanna City, while A.coryli was discovered in Ankang City and A.oenotherae was found in Lincang City, which are both subtropical regions. Consistent with previous studies, the majority of Apiospora species inhabit a diverse range of habitats primarily located in tropical and subtropical regions (Pintos and Alvarado 2021).

Specimens of Apiospora were collected from the Qinling Mountains in Ankang City and, in addition to A.coryli, Jiang et al. reported species found including A.qinlingense and A.neochinensis (Jiang et al. 2018; Jiang et al. 2020). Amongst these species, A.coryli was found to have longer conidiogenous cells (2.6–10.6 × 2.1–5.8 µm) compared to A.qinlingense (1–2 × 2–3 µm) and A.neochinensis (1.5–6.5 × 1–3.5 µm) and much larger spores than A.qinlingense (4–18.4 × 6.2–12.5 µm vs. 5–8 × 5–8 µm) (Table 6). These morphological differences suggest that A.coryli is distinct from A.qinlingense and A.neochinensis. This distinction is also supported by phylogenetic analysis shown in Fig. 1 which revealed that these species are phylogenetically distant from each other. Different species have been discovered in this region over several years, indicating that variation in species may be linked to the timing of collection, host plants, growth rates, developmental cycles and activity levels. These findings highlight the diversity of fungi within Apiospora genus in the subtropical region of the Qinling Mountains and suggest the existence of numerous undiscovered species with significant research potential. Further investigation is necessary to determine the value of specific regions for future research on fungi.

Table 6.

Synopsis of new Apiospora species and species collected from the Qinling Mountains in Apiospora.

Species	Conidiogenous cells (µm)	Conidia (µm)	Host	Date	References
Apiosporacoryli	2.6–10.6 × 2.1–5.8	4–18.4 × 6.2–12.5	Corylusyunnanensis	16 July 2021	Present study
A.qinlingense	1–2	5–8	Fargesiaqinlingensis	27 June 2017	Jiang et al. 2018
A.neochinensis	1.5–6.5 × 1–3.5	8–12 × 5.5–9	Fargesiaqinlingensis	16 July 2018	Jiang et al. 2020

* Newly described taxa are in bold.

This paper reports the initial discovery of A.lophatheri on Lophatherumgracile (Poaceae). While numerous Apiospora have been discovered on Poaceae plants worldwide, previous research has primarily focused on bamboo, with limited investigation into herbaceous plants, such as Lophatherum (Liu et al. 2023). However, prior to this study, Apiospora had not been previously found on Brunfelsia (Solanaceae) and Oenothera (Onagraceae). While Cercosporabrunfelsiicola has been reported on other host Brunfelsiauniflora within the genus and Pestalotiopsisoenotberae has been identified specifically on Oenotheralaciniata, the restricted cultivation of these plants along with insufficient research on their associated fungi have resulted in few related studies (Venkatasubbaiah et al. 1991; Hidayat and Meeboon 2014). This discovery highlights potential interactions between these plant species and their fungal counterparts, emphasising the importance of uncommon herbaceous plants for fungal taxonomy alongside Rosaceae and silvicultural species like Populus (Peng et al. 2022; Lin et al. 2022). Hence, collecting various specimens is crucial for studying and identifying the fungi of Apiospora, while also promoting fungal diversity.

Most Apiospora species exhibit round or lenticular conidia, as demonstrated in this study. Nevertheless, the sizes of these conidia often overlap amongst morphologically similar, but phylogenetically distinct species within the genus Apiospora. For example, the conidia of A.piptatheri (7.5–10 × 7–9 µm) and A.pseudosinense (8–10 × 7–10 µm) are similar, but the two species are comparable despite their distinct evolutionary lineages in Fig. 1 (Crous and Groenewald 2013; Pintos et al. 2019). Therefore, relying merely on morphology can pose challenges for accurate identification.

The monophyly of taxonomic classification units at every rank is crucially important. Morphology is frequently insufficient for phylogenetic classification and, thus, molecular evidence has become increasingly significant and indispensable for identifying and classifying fungal taxa. In recent years, there has been a steady growth in DNA sequencing data available for Apiospora species (Crous and Groenewald 2013; Wang et al. 2018; Pintos et al. 2019), leading to the recognition of 93 species of Apiospora. Sequence data are accessible for ITS in 93 species, LSU in 80, tef1 in 71 and tub2 in 73, facilitating accurate and swift identification (Wang et al. 2018; Pintos et al. 2019). However, using ITS alone has its limitations in identifying Apiospora species. Therefore, LSU, tef1, tub2 and multigene sequence data (ITS, LSU, tef1 and tub2) have been particularly useful in establishing phylogenetic relationships and increasing accuracy in Apiospora identification. Furthermore, this study yielded 32 sequence datasets for four gene regions (ITS, LSU, tef1 and tub2), enhancing our comprehension of the genus Apiospora. Novel species were identified by examining morphological and molecular characteristics, host associations and ecological distributions.

Citation

Li S, Peng C, Yuan R, Tian C (2023) Morphological and phylogenetic analyses reveal three new species of Apiospora in China. MycoKeys 99: 297–317. https://doi.org/10.3897/mycokeys.99.108384

Additional information

Conflict of interest

The authors have declared that no competing interests exist.

Ethical statement

No ethical statement was reported.

Funding

This study is financed by National Natural Science Foundation of China (Project No.: 31670647).

Author contributions

Conceptualization, Shuji Li and Chengming Tian; data curation, Shuji Li;funding acquisition, Chengming Tian; investigation, Shuji Li and Rong Yuan; project administration, Chengming Tian; resources, Shuji Li and Rong Yuan; supervision, Chengming Tian; writing-original draft, Shuji Li; writing-review and editing, Shuji Li, Cheng Peng, and Chengming Tian. All authors have read and agreed to the published version of the manuscript.

Author ORCIDs

Shuji Li https://orcid.org/0009-0006-4734-8399

Chengming Tian https://orcid.org/0000-0002-3352-7664

Data availability

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

Supplementary materials

Supplementary material 1

Isolates and GenBank accession numbers used in the phylogenetic analyses

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.

Shuji Li, Cheng Peng, Rong Yuan, Chengming Tian

Data type

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