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Research Article
Two new species in Capillidium (Ancylistaceae, Entomophthorales) from China, with a proposal for a new combination
expand article infoYong Nie§, Heng Zhao|, ZiMin Wang§, ZhengYu Zhou§, XiaoYong Liu, Bo Huang
‡ Anhui Agricultural University, Hefei, China
§ Anhui University of Technology, Ma'anshan, China
| Beijing Forestry University, Beijing, China
¶ Shandong Normal University, Jinan, China
Open Access

Abstract

A taxonomic revision of Conidiobolus s.l. (Ancylistaceae, Entomophthorales) delimited all members that form capilliconidia into the genus Capillidium. In this study, we report two new species of Capillidium that were isolated in China. Capillidium macrocapilliconidium sp. nov. is characterised by large capilliconidia. Capillidium jiangsuense sp. nov. is differentiated by large capilliconidia and long, slender secondary conidiophores. Phylogenetic analyses were performed using sequences from the nuclear large subunit of rDNA (nucLSU), the mitochondrial small subunit of rDNA (mtSSU) and elongation-factor-like (EFL). The analyses revealed sister relationships between Ca. macrocapilliconidium sp. nov. and Ca. globuliferus / Ca. pumilum and between Ca. jiangsuense sp. nov. and Ca. denaeosporum. Additionally, a new combination of Ca. rugosum (Drechsler) B. Huang & Y. Nie comb. nov. is proposed herein. An identification key is provided for the ten accepted Capillidium species.

Keywords

Ancylistaceae, Capilliconidia, morphology, new taxa, phylogeny

Introduction

The taxonomic name Capillidium was first introduced as a subgenus within the genus Conidiobolus (Ancylistaceae, Entomophthorales) (Ben-Ze’ev and Kenneth 1982). All its members were clustered into a monophyletic group in the family Ancylistacaceae, based on four molecular loci [i.e. small subunit of nuclear rDNA (nucSSU), large subunit of nuclear rDNA (nucLSU), small subunit of mitochondrial rDNA (mtSSU) and elongation-factor-like (EFL)] (Nie et al. 2020). Species in this genus are typically characterised by capilliconidia protruding from elongated, slender conidiophores (Nie et al. 2020). Based on this synapomorphy and a re-examination of the protologue for Conidiobolus s.l. species (Drechsler 1953a, b, 1954, 1955a, 1957; Srinivasan and Thirumalachar 1967, 1968; Callaghan et al. 2000), seven species so far have been recombined into the monophyletic genus Capillidium, including: Ca. adiaeretum (Drechsler) B. Huang & Y. Nie, Ca. bangalorense (Sriniv. & Thirum.) B. Huang & Y. Nie, Ca. denaeosporum (Drechsler) B. Huang & Y. Nie, Ca. heterosporum (Drechsler) B. Huang & Y. Nie, Ca. lobatum (Sriniv. & Thirum.) B. Huang & Y. Nie, Ca. pumilum (Drechsler) B. Huang & Y. Nie and Ca. rhysosporum (Drechsler) B. Huang & Y. Nie (Nie et al. 2020).

Although Capillidium is a small genus with only seven accepted species, it possesses high morphological diversity. For instance, primary conidia range from 18 μm (Ca. pumilum) to 46 μm (Ca. adiaeretum) in size (Drechsler 1953a, 1955a); resting spores are present in Ca. adiaretum, Ca. bangalorense and Ca. rhysosporum, but not in Ca. denaeosporum, Ca. heterosporum, Ca. lobatum and Ca. pumilum (Drechsler 1953b, 1955a, 1957); Ca. heterosporum has slender conidiophores that are branched at the base and end with 2–6 terminal capilliconidia each (Drechsler 1953b), whereas other members are unbranched and end with one capilliconidia (Nie et al. 2020); although nearly all Capillidium species only produce capilliconidia, Ca. adiaeretum also produces microconidia (Callaghan et al. 2000). These important diagnostic characteristics can help mycologists form a comprehensive understanding of this fungal group.

Two species Ca. adiaeretum and Ca. heterosporum have been identified in China (Wang et al. 2010; Nie et al. 2020). Continuing investigations into Chinese Conidiobolus s.l. led to the discovery of two new species in the genus Capillidium. We describe them herein, suggest a new combination for this genus and provide an updated identification key for the species of Capillidium.

Materials and methods

Isolates and morphology

Plant debris was collected from Wanfo Mountain, Shucheng County, Anhui Province, China and Laoshan National Forest Park and Tianwang Town, Jiangsu Province, China. Pre-sterilised plastic bags were used to pack these plant debris samples. Isolation procedures were the same as described by Drechsler (1952) and King (1976a). Plant debris samples were incubated on inverted Petri dishes containing PDA medium (potato 200 g, dextrose 20 g, agar 20 g, H2O, 1 litre) at 21 °C for 4 days. The incubated dishes were examined daily under a stereomicroscope (SMZ1500, Nikon Corporation, Japan). When an entomophthoroid fungus appeared, it was transferred to a clean PDA plate for purification and then sub-cultivated for morphological studies. Microscopic structure was observed under a light microscope (BX51, Olympus Corporation, Tokyo, Japan) and imaged using a microscope-camera system (DP25, Olympus Corporation, Tokyo, Japan). The size and shape of the primary conidia, primary conidiophores, secondary conidiophores, capilliconidia etc. were measured and described using the method by King (1976a) and the type of replicative conidia were observed on 2% water agar (agar 2 g, H2O, 1 litre). All isolates were deposited at the Research Center for Entomogenous Fungi at Anhui Agricultural University, Anhui Province, China (RCEF) and duplicated at the China General Microbiological Culture Collection Center, Beijing, China (CGMCC). A total of 13 ex-types of Conidiobolus s.l. were acquired from the American Type Culture Collection, Manassas, VA, USA (ATCC).

DNA extraction, PCR amplification and sequencing

Total cellular DNA was extracted using the method by Watanabe et al. (2010). For phylogenetic analyses, three loci were amplified using relevant primer pairs: LR0R (5’-ACC CGC TGA ACT TAA GC-3’) / LR5 (5’-TCC TGA GGG AAA CTT CG-3’) for nucLSU (Vilgalys and Hester 1990), mtSSU1 (5’-GCW GCA GTG RGG AAT NTT GGR CAA T-3’) / mtSSU2R (5’-GTR GAC TAM TSR GGT ATC TAA TC-3’) for mtSSU (Zoller et al. 1999) and EF983 (5’-GCY CCY GGH CAY CGT GAY TTY AT-3’) / EF1aZ-1R (5’-ACA TCW CCG ACA CCC TTG ATC TTG -3’) for EFL (Nie et al. 2012).

PCR amplification was carried out in a 50 µl mixture containing 1 μl dNTPs (200 μM), 1 μl MgCl2 (2.5 mM), 10 µl Phusion HF buffer (5×), 1 μl primers each (0.5 μM), 100 ng genomic DNA and 0.5 μl Taq polymerase (0.04 Unit/l, Super Pfx DNA Polymerase, Cowinbioscience Co. Ltd., Shanghai, China). PCR runs were conducted under the following conditions: an initial denaturation step at 94 °C for 3 min followed by 35 cycles of denaturation at 94 °C for 1 min, annealing at 55 / 54 / 57 °C (nucLSU / mtSSU / EFL), extension at 72 °C for 1 min; a final extension step at 72 °C for 7 min. DNA sequences were generated on both strands by performing dideoxy-nucleotide chain termination on an ABI 3700 automated sequencer at the Shanghai Genecore Biotechnologies Company (Shanghai, China). Sequences were processed with Geneious 9.0.2 (http://www.geneious.com, Kearse et al. 2012) and deposited in GenBank under the accession numbers listed in Table 1.

Table 1.

The species used in phylogenetic analyses.

Species Strains* GenBank accession numbers References
nucLSU EFL mtSSU
Azygosporus macropapillatus CGMCC 3.16068 (T) MZ542006 MZ555650 MZ542279 Cai et al. (2021)
A. parvus ATCC 14634 (T) KX752051 KY402207 MK301192 Cai et al. (2021)
Capillidium adiaeretum ARSEF 451 (T) KC461182 GenBank
Ca. adiaeretum CGMCC 3.15888 MN061284 MN061481 MN061287 Nie et al. (2020)
Ca. bangalorense ARSEF 449 (T) DQ364204 DQ364225 Chen and Huang (2018)
Ca. denaeosporum ATCC 12940 (T) JF816215 JF816228 MK301181 Nie et al. (2012, 2020)
Ca. globuliferum CBS 152.56 (T) MH869095 Vu et al. (2019)
Ca. heterosporum CBS 543.63 MH869973 Vu et al. (2019)
Ca. heterosporum RCEF 4430 JF816225 JF816239 MK301183 Nie et al. (2012, 2020)
Ca. lobatum ATCC 18153 (T) JF816218 JF816233 MK301187 Nie et al. (2012, 2020)
Ca. pumilum ARSEF 453 (T) EF392383 EF392496 GenBank
Ca. rhysosporum ATCC 12588 (T) JN131540 JN131546 MK301195 Nie et al. (2018, 2020)
Ca. rhysosporum CBS 141.57 MH869215 Vu et al. (2019)
Ca. rugosum CBS 158.56 (T) MH869097 Vu et al. (2019)
Ca. marcocapilliconidium CGMCC 3.16169 (T) OL830454 OL801337 OL830457 This article
Ca. marcocapilliconidium RCEF 6332 OL830455 OL801338 OL830458 This article
Ca. jiangsuense CGMCC 3.16168 (T) OL830456 OL801339 OL830459 This article
Conidiobolus coronatus NRRL 28638 AY546691 DQ275337 Lutzoni et al. (2004)
C. humicolus ATCC 28849 (T) JF816220 JF816231 MK301184 Nie et al. (2012, 2020)
C. khandalensis ATCC 15162 (T) KX686994 KY402204 MK301185 Nie et al. (2012, 2020)
C. lichenicolus ATCC 16200 (T) JF816216 JF816232 MK301186 Nie et al. (2012, 2020)
C. polytocus ATCC 12244 (T) JF816213 JF816227 MK301194 Nie et al. (2012, 2020)
Microconidiobolus nodosus ATCC 16577 (T) JF816217 JF816235 MK333388 Nie et al. (2012, 2020)
M. paulus ARSEF 450 (T) KC788409 Gryganskyi et al. (2013)
M. terrestris ATCC 16198 (T) KX752050 KY402208 MK301199 Nie et al. (2016, 2020)
Neoconidiobolus couchii ATCC 18152 (T) JN131538 JN131544 MK301179 Nie et al. (2016, 2020)
N. mirabilis CGMCC 3.17763 (T) MH282852 MH282853 MK333389 Nie et al. (2018, 2020)
N. pachyzygosporus CGMCC 3.17764 (T) KP218521 KP218524 MK333390 Nie et al. (2018, 2020)
N. stromoideus ATCC 15430 (T) JF816219 JF816229 MK301198 Nie et al. (2012, 2020)
N. thromboides ATCC 12587 (T) JF816214 JF816230 MK301200 Nie et al. (2012, 2020)

Phylogenetic analyses

The data for the three target loci (nucLSU, mtSSU and EFL) were produced during this study and during our previous study (Nie et al. 2020). Sequences were retrieved from GenBank and concatenated using SequenceMatrix 1.7.8 (Vaidya et al. 2011). For this analysis, fifteen species in four closely-related genera (Azygosporus, Conidiobolus s.s., Neoconidiobolus and Microconidiobolus) served as outgroups (Table 1). Local alignment was conducted with MUSCLE 3.8.31 (Edgar 2004) and manually refined with BioEdit v. 7.2.6 (Hall 1999). The aligned sequence matrix was deposited in TreeBase (https://treebase.org) under the submission ID S29102.

Phylogenetic analyses were performed using three different methods: Maximum Likelihood (ML), Maximum Parsimony (MP) and Bayesian Inference (BI). For ML and BI analyses, best-fit substitution models for each locus were estimated in Modeltest 3.7 using the Akaike Information Criterion (AIC) value (Posada and Crandall 1998). The ML phylogenetic analysis was statistically tested in RAxML 8.1.17 with 1000 bootstrap replicates (Stamatakis 2014). The BI analysis was carried out in MrBayes v.3.1.2 using Markov Chain Monte Carlo (MCMC) methods (Ronquist and Huelsenbeck 2003). Beginning with random starting trees, four MCMC chains ran simultaneously for 1 million generations. The trees were sampled once every 100 generations. These chains stopped when all convergences met and the standard deviation fell below 0.01. MP analyses were conducted using a heuristic search in PAUP* 4.0b10 (Swofford 2002). Bootstrap analyses were conducted with 1000 bootstrap replicates to determine the confidence levels of the nodes within the inferred tree topologies (Felsenstein 1985). Tree bisection-reconnection (TBR) was selected for branch swapping. Phylogenetic trees were checked with FigTree 1.4 (Rambaut 2012) and further modified with iTOL (https://itol.embl.de/).

Results

Phylogenetic analyses

The concatenated alignment included 30 strains, 15 of which were outgroups from Azygosporus, Conidiobolus s.s., Microconidiobolus and Neoconidiobolus (Table 1). The aligned three-locus datasets contained 1861 characters. Amongst these, 852 characters were constant, 159 were parsimony-uninformative and 850 were parsimony informative. The most parsimonious tree had a tree length (TL) consisting of 3445 steps, a consistency index (CI) of 0.5068, a homoplasy index (HI) of 0.4932, a retention index (RI) of 0.7145 and a rescaled consistency index (RC) of 0.3621. The ML and BI analyses were performed using the best models for nucLSU (TrNef+G), EFL (TIMef) and mtSSU (K81) partitioning. The final average standard deviation of the split frequencies was 0.0059 and the final likelihood value was -17189. The tree topology from ML analysis was identical to those obtained from MP and BI analyses. The final ML tree was generated with bootstrap support values from MP/ML analyses, as well as posterior probability values from BI analysis at each branch.

Figure 1. 

The phylogenetic tree of Capillidium constructed using Maximum Likelihood analyses on nucLSU, EFL and mtSSU sequences. Conidiobolus s.l. species were used as outgroups. New taxa are indicated by bold text. Maximum Parsimony bootstrap values (≥ 50%) / Maximum Likelihood bootstrap values (≥ 50%) / Bayesian posterior probabilities (≥ 0.50) of clades are provided alongside the branches. The scale bar at the lower left indicates substitutions per site.

The phylogeny revealed that three strains belong to the genus Capillidium. The strains CGMCC 3.16169 / RCEF 6332 and CGMCC 3016168 were grouped closely with Ca. pumilum / Ca. globuliferus (100/100/1.00) and Ca. denaeosporum (100/100/1.00), respectively.

Taxonomy

Capillidium macrocapilliconidium B. Huang & Y. Nie, sp. nov.

MycoBank No: 842227
Fig. 2

Etymology

macrocapilliconidium (Lat.), referring to the large size of its capilliconidia.

Known distribution

Jiangsu Province, China.

Typification

China, Jiangsu Province, Nanjing City, Laoshan National Forest Park, 32°5'52"N, 118°35'37"E, from plant debris, 1 Dec 2018, Y. Nie and Y. Gao, culture ex-holotype CGMCC 3.16169 (=RCEF 6553).

Additional specimens examined

China, Anhui Province, Shucheng County, Wanfo Mountain, 31°9'51"N, 116°57'86"E, from plant debris, 13 Mar 2016, X.X. Tang, culture RCEF 6332. GenBank: nrLSU = OL830455; EFL = OL801338; mtSSU = OL830458.

Description

Colonies on PDA at 21 °C after 3 d white, reaching ca. 28 mm in diameter, yellowish after 10 d. Mycelia hyaline, 5.5–10 μm wide, often branched. Primary conidiophores arising from hyphal segments, hyaline, 70–250 × 5–13 μm, unbranched and producing a single globose primary conidium, widening upwards near the tip. Primary conidia forcibly discharged, globose to subglobose, 25–34 × 20–28 μm, papillate or conical, 7–10 μm wide, 3–8 μm long. Secondary conidiophores short or long, arising from primary conidia, bearing a single replicative conidium similar to, but smaller than those primary ones and forcibly discharged, producing another kind of replicative conidia called capillidiconidia from slender secondary conidiophores on the 2% water agar. Capillidiconidia colourless, elongate ellipsoidal, 25–37 μm long, 14–17 μm wide. Slender secondary conidiophores unbranched, 85–130 μm long, 4–6 μm wide at the base, tapering gradually to a width of 1–2 μm at the tip. Zygospores usually formed between adjacent segments of the same hypha after 10 d, yellowish, mostly boldly wrinkled, sometimes smooth, globose, elongate ellipsoidal or irregular, 18–35 μm long, 17–28 μm wide, with a wall 1–2 μm thick.

Figure 2. 

Capillidium macrocapilliconidium a colony on PDA after 3 d at 21 °C b colony on PDA after 10 d at 21 °C c Mycelia d Mycelia unbranched at the edge of the colony e, f primary conidiophores bearing primary conidia g, h, i primary conidia j, k primary conidia bearing a single secondary conidium i, m, n a primary conidium bearing a single capilliconidium o, p, q, r Capilliconidia s zygospores that were formed on adjacent segments of the same hypha t immature zygospores u, v mature zygospores. Scale bars: 10 mm (a–b); 100 μm (c–d); 20 μm (e–v).

Notes

Capillidium macrocapilliconidium is characterised by having larger capilliconidia compared to other Capillidium species. It produces yellowish and wrinkled zygospores like Ca. rhysosporum (Drechsler 1954). However, Ca. macrocapilliconidium has larger capilliconidia than Ca. rhysosporum (25–37 × 14–17 μm in Ca. macrocapilliconidium vs. 12–32 × 6.5–16 μm in Ca. rhysosporum). Ca. macrocapilliconidium is phylogenetically distant from Ca. rhysosporum (Fig. 1) and most closely related to Ca. pumilum. It is distinguished from Ca. pumilum by larger primary conidia (25–34 × 20–28 μm in Ca. macrocapilliconidium vs. 9–18 × 7.3–14 μm in Ca. pumilum) and capilliconidia (25–37 × 14–17 μm in Ca. macrocapilliconidium vs. 8.8–12 × 5–7.5 μm in Ca. pumilum) (Drechsler 1955b).

Capillidium jiangsuense B. Huang & Y. Nie, sp. nov.

MycoBank No: 842228
Fig. 3

Etymology

jiangsuense (Lat.), referring to the region where the fungus was isolated.

Known distribution

Jiangsu Province, China.

Typification

China, Jiangsu Province, Jurong City, Tianwang Town, 31°6'94"N, 119°26'91"E, from plant debris, 25 Mar 2018, Y. Nie, culture ex-holotype CGMCC 3.16168 (=RCEF 6545).

Description

Colonies on PDA at 21 °C after 3 d white, reaching ca. 21 mm in diameter. Mycelia haline, often unbranched, vegetative hyphae filamentous, 5–10 μm wide. Primary conidiophores unbranched, producing a single primary conidium, widening upwards near the tip, 50–240 × 6–10 μm. Primary conidia forcibly discharged, subglobose to turbinate, 21–31 × 12–29 μm. Papilla 4–10 μm wide, 2–4 μm long. Replicative conidia two kinds on 2% water agar, arising from primary conidia, one similar and smaller to the primary conidia, the other elongate and passively detached, 17–32 × 10–15 μm. Slender secondary conidiophores unbranched, 65–120 μm long, 2.5–3 μm wide at the base, tapering gradually to a width of 1 μm at the tip. Resting spore not observed.

Figure 3. 

Capillidium jiangsuense a colony on PDA after 3 d at 21 °C b Mycelia c Mycelia unbranched at the edge of the colony d, e, f primary conidiophores arising from mycelia segments g, h, i, j primary conidia k, i secondary conidia arising from primary conidia m, n primary conidia bearing a single capilliconidium o, p, q, r Capilliconidia. Scale bars: 10 mm (a); 100 μm (b, c); 20 μm (d–r).

Notes

Morphologically, the present isolate resembles Ca. denaeosporum because of the size of its primary conidia (13–32 × 6–21 μm in Ca. denaeosporum vs. 21–31 × 12–29 μm in Ca. jiangsuense) (Drechsler 1957). However, Ca. denaeosporum has larger capilliconidia (10–18 × 6–10 μm in Ca. denaeosporum vs. 17–32 × 10–15 μm in Ca. jiangsuense) and longer, more slender secondary conidiophores (35–65 μm in Ca. denaeosporum vs. 65–120 μm in Ca. jiangsuense) (Drechsler 1957). Although they grouped together with relatively little divergence on the phylogram, DNA similarity levels between the two species are only around 97.9% (nucLSU) (Nie et al. 2012). This evidence supports the present isolate being a new species, which we have named Capillidium jiangsuense sp. nov.

Capillidium rugosum (Drechsler) B. Huang & Y. Nie, comb. nov.

MycoBank No: 842229

Basionym

Conidiobolus rugosus Drechsler, Am. J. Bot. 42: 437 (1955).

Description

Refer to Drechsler (1955a).

Notes

The ex-type living culture is ATCC 12586 (United States, New Jersey, Moorestown, 25 February 1954, Drechsler). Historically, Conidiobolus rugosus was synonymised with Co. heterosporus (King 1976b). However, we have re-established its taxonomic status at the species level, based on the phylogeny herein and the morphological traits of the capilliconidia.

Discussion

From the 1950s-1970s, a total of eight Conidiobolus species have been reported to produce capilliconidia, including Conidiobolus denaeosporus, Co. globuliferus, Co. heterosporus, Co. inordinatus, Co. lobatus, Co. pumilus, Co. rhysosporus and Co. rugosus (Drechsler 1953a, b, 1954, 1955a, b, 1956, 1957; Srinivasan and Thirumalachar 1968). Based on the numerical taxonomy of Conidiobolus (King 1976a, b, 1977), four species were rejected. Co. rugosus was considered synonymous with C. heterosporus. On the other hand, Conidiobolus denaeosporus, Co. globuliferus and Co. inordinatus were considered synonymous with Co. pumilus. Consequently, only four species forming capilliconidia were accepted into this genus. Based on this synapomorphy, the subgenus Capillidium was erected in the latter taxonomic study of Conidiobolus (Ben-Ze’ev and Kenneth 1982; Humber 1989). Interestingly, it appears that Co. adiaeretus and Co. bangalorensis develop both microconidia and capilliconidia (Callaghan et al. 2000). Unfortunately, there was no molecular evidence at the time to support these morphological results. Recently, we summarised molecular data from available Conidiobolus s.l. ex-types and identified a monophyletic lineage of Capillidium producing capilliconidia. Since then, some taxonomic revisions have been conducted. For example, Co. denaeosporus was separated from Co. pumilus and recombined into Capillidium. Co. adiaeretus and Co. bangalorensis were also recombined into Capillidium. In total, Capillidium now has seven accepted species.

Conidiobolus heterosporus (= Capillidium heterosporum) and Co. rugosus share distinct morphological characteristics. For instance, Co. heterosposus bears no resting spores and has conidiophores that are often branched at the base and bear 2–6 terminal capilliconidia (Drechsler 1953a). The conidiophores of Co. rugosus, though, have yellowish zygospores with wrinkled or smooth surfaces, are unbranched and bear a single capilllicondiuma (Drechsler 1955b).

Based on a phylogenetic analysis of three gene regions (nucLSU, mtSSU and EFL), the ex-type of Co. rugosus (Strain No: CBS 158.56) and Co. heterosposus diverged into two distinct lineages. Consequently, we identified Co. rugosus as an independent species and recombined it into Capillidium as a new combination: Capillidium rugosum (Drechsler) B. Huang & Y. Nie comb. nov. On a side note, while researchers previously considered Co. denaeosporus (= Ca. denaeosporum), Co. globuliferus and Co. inordinatus to be synonymous with Co. pumilus (= Ca. pumilum) (King 1976b), the present phylogeny confirmed that Co. denaeosporus (= Ca. denaeosporum) is an independent species and Co. globuliferus is synonymous with Co. pumilus (= Ca. pumilum). More molecular evidence is needed to clarify the taxonomic status of Co. inordinatus.

Capillidium bangalorense may be another Capillidium species that forms microspores, based on its close phylogenetic relationship with Ca. adiaeretum. Besides microspores, these two species possess another morphological characteristic that is distinctive compared with the other members of Capillidium, that being the width between the primary conidiophores and the hyphae (Drechsler 1955a; Srinivasan and Thirumalachar 1967). This could explain why Ca. adiaeretum and Ca. bangalorense are grouped into a single clade in the phylogenetic tree (Fig. 1). However, Ca. bangalorense should be re-examined and more evidence should be supplied to confirm that this clade is in a separate taxon.

With the current description of Azygosporus, most members of Conidiobolus s.l. have now received suitable taxonomic placements. Yet, there are still many other taxonomic challenges to be resolved in the future, such as replacing the missing ex-type Co. utriculosis and assigning Co. coronatus as the epitype of Conidiobolus s.s., isolating lost ex-types to confirm their taxonomic placements etc. (Nie et al. 2018, 2020, 2021; Cai et al. 2021). For the first time, this study used partial sequence data from nucLSU, mtSSU and EFL genes to identify two new species of Capillidium from China, increasing the total number of species in the genus to ten. A key to the species of Capillidium is provided below.

Key to the Species of Capillidium

1 Capilliconidia and microconidia produced, the width of primary conidiophores offers a pronounced dimensional contrast with the mycelial filaments 2
Only capilliconidia produced, the width of primary conidiophores offers a similar dimensional contrast with the mycelial 3
2 Primary conidia larger, up to 46 μm, chlamydospores produced Ca. adiaeretum
Primary conidia smaller, less than 25 μm, zygospores produced Ca. bangalorense
3 Slender conidiophores branched at the base, bearing 2–6 terminal capilliconidia Ca. heterosporum
Slender conidiophores unbranched at the base, bearing a single capilliconidia 4
4 Resting spores of zygospores produced, yellowish, mostly wrinkled, sometimes smooth 5
Resting spores not observed 6
5 Primary conidia and zygospores larger, more than 30 μm 7
Primary conidia and zygospores smaller, less than 25 μm Ca. rugosum comb. nov.
6 Primary conidia larger, more than 30 μm 8
Primary conidia smaller, less than 26 μm 9
7 Capilliconidia larger, up to 37 μm Ca. macrocapilliconidium sp. nov.
Capilliconidia smaller, less than 32 μm Ca. rhysosporum
8 Capilliconidia larger, 17–32 × 10–15 μm, primary conidiophores longer, 50–240 μm Ca. jiangsuense sp. nov.
Capilliconidia smaller, 10–18 × 6–10 μm, primary conidiophores shorter, 15–50 μm Ca. denaeosporum
9 Primary conidia larger, 21–26 × 20–24 μm, capilliconidia larger, 18–25 × 8–10 μm Ca. lobatum
Primary conidia smaller, 9–18 × 7.3–14 μm, capilliconidia smaller, 8.8–12 × 5–7.5 μm Ca. pumilum

Acknowledgements

We thank Dr. Yang Gao (Jiangxi Agricultural University) for helping with sample collection. We also thank Dr. Jeffery Hannahn (Michigan State University) for his assistance with English language and grammatical editing. This study was supported by the National Natural Science Foundation of China (Nos. 31900008, 30770008 31670019 and 31970009).

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