Abstract
Akanthomyces species have only been reported from Guizhou and Qinghai Province, with few reports from other regions in China. In this research, the species diversity of Akanthomyces in the Jinyun Mountains, Chongqing was investigated. Fourteen infected spider specimens were collected and two new species (A.bashanensis and A.beibeiensis) and a known species (A.tiankengensis) were established and described according to a multi-locus phylogenetic analysis and the morphological characteristics. Our results reveal abundant Akanthomyces specimens and three species were found at Jinyun Mountain. Due to its being an important kind of entomopathogenic fungi, further attention needs to be paid to the diversity of other entomopathogenic fungi in Chongqing, China.
Key words: entomopathogenic fungi, morphology, phylogenetic analysis, Sordariomycetes, spider-pathogenic fungi
Introduction
Akanthomyces Lebert was established by Lebert for the species, A.aculeatus Lebert in 1858. Mains (1950) reported three additional species: A.ampullifer (Petch) Mains, A.angustisporus Mains and A.aranearum (Petch) Mains. Subsequently, several other species have been reported (Samson and Evans 1974; Koval 1977; Vincent et al. 1988; Hywel-Jones 1996; Hsieh et al. 1997; Huang et al. 2000; Mongkolsamrit et al. 2018; Chen et al. 2018, 2019c, 2020b, 2020c, 2022a; Shrestha et al. 2019; Aini et al. 2020; Wang et al. 2023).
Based mainly on phylogenetic analyses, several Akanthomyces species (A.arachnophilus (Petch) Samson & H.C. Evans, A.cinereus Hywel-Jones, A.koratensis Hywel-Jones, A.longisporus B. Huang et al., A.novoguineensis Samson & B.L. Brady, A.ovalongatus L.S. Hsieh et al. and A.websteri Hywel-Jones) were transferred to the new genus Hevansia Luangsa-ard et al. (Kepler et al. 2017). In addition, Lecancilliumattenuatum Zare & W. Gams, L.lecanii (Zimm.) Zare & W. Gams, L.longisporum (Petch) Zare & W. Gams, L.muscarium (Petch) Zare & W. Gams, L.pissodis Kope & I. Leal and L.sabanense Chir.-Salom., S. Restrepo & T.I. Sanjuan were transferred to Akanthomyces. As a result, the genus Akanthomyces currently consists of 34 species.
Akanthomyces is an important genus in entomopathogenic fungi and its diverse bioactive substances have attracted widespread attention (Lee et al. 2005; Madla et al. 2005; Kuephadungphan et al. 2014; Putri et al. 2014; Kinoshita et al. 2017). However, Akanthomyces species have only been reported from Guizhou and Qinghai Province and there have been few reports from other regions in China (Chen et al. 2018, 2019c, 2020b, 2020c, 2022a; Wang et al. 2023). In this research, the species diversity of Akanthomyces in the Jinyun Mountains, Chongqing was investigated. Several spider-associated specimens were found and a few new Akanthomyces strains were isolated and purified. The goal of this research was to identify these new strains by multigene phylogeny as well as by morphological characteristics.
Materials and methods
Specimen collection and identification
Fourteen infected spider specimens were collected from Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E), Beibei District, Chongqing, in May 2021. The surface of each spider body was rinsed with sterile water, followed by surface sterilisation with 75% ethanol for 3–5 s and rinsing 3 times with sterilised water. After drying on sterilised filter paper, the mycelium or a part of the sclerotium was removed from the specimen and inoculated on potato dextrose agar (PDA) and improved potato dextrose agar (PDA, 1% w/v peptone) plates (Chen et al. 2019b). Fungal colonies emerging from the specimens were isolated and cultured at 25 °C for 14 days under 12 h light/12 h dark conditions following protocols described by Zou et al. (2010). The specimens and axenic cultures were deposited at the Institute of Fungus Resources, Guizhou University (formally Herbarium of Guizhou Agricultural College; code, GZAC), Guiyang City, Guizhou, China.
Macroscopic characterisation was determined from PDA cultures incubated at 25 °C for 14 days, including the growth rate of the colony, the presence of octahedral crystals, and the colours of the colony (surface and reverse) were observed. To investigate the microscopic characteristics, a small amount of the colony was removed and mounted in lactophenol cotton blue or 20% lactate acid solution and observed using an optical microscope (OM, DM4 B, Leica, Germany).
DNA extraction, polymerase chain reaction amplification and nucleotide sequencing
DNA extraction was carried out using a fungal genomic DNA extraction kit (DP2033, BioTeke Corporation) according to Liang et al. (2011). The extracted DNA was stored at −20 °C. Polymerase chain reaction (PCR) was used to amplify genetic markers using the following primer pairs: ITS4/ITS5 for the internal transcribed spacer (ITS) region (White et al. 1990), LR0R/LR5 for 28s large subunit ribosomal (LSU) (Vilgalys and Hester 1990), fRPB2-7cR/fRPB2-5F for RNA polymerase II second largest subunit (RPB2) (Liu et al. 1999) and 2218R/983F for translation elongation factor 1 alpha (TEF) (Castlebury et al. 2004). The thermal cycle of PCR amplification for these phylogenetic markers was set up following the procedure described by Chen et al. (2021). PCR products were purified and sequenced at Sangon Biotech (Shanghai) Co. The resulting sequences were submitted to GenBank (Table 1).
Table 1.
List of strains and GenBank accession numbers of sequences used in this study.
Sequence alignment and phylogenetic analyses
DNASTAR Lasergene (version 6.0) was used to edit the DNA sequences. The ITS, LSU, RPB2 and TEF sequences were downloaded from GenBank, based on Sung et al. (2007), Kepler et al. (2017), Mongkolsamrit et al. (2018), Chen et al. (2018, 2019c, 2020b, 2020c, 2022a), Aini et al. (2020) and Wang et al. (2023) and others selected on the basis of BLAST searches in GenBank. ITS sequences and other loci were aligned and edited by MAFFT online service (Katoh et al. 2019) and MEGA6 (Tamura et al. 2013). Combined sequences of ITS, LSU, RPB2 and TEF were obtained using SequenceMatrix v.1.7.8 (Vaidya et al. 2011). The model was selected for Bayesian analysis by ModelFinder (Kalyaanamoorthy et al. 2017) in PhyloSuite software (Zhang et al. 2020).
The combined loci were analysed using Bayesian inference (BI) and maximum likelihood (ML) methods. For BI, a Markov chain Monte Carlo (MCMC) algorithm was used to generate phylogenetic trees with Bayesian probabilities using MrBayes v.3.2 (Ronquist et al. 2012) for the combined sequence datasets. The Bayesian analysis resulted in 20,001 trees after 10,000,000 generations. The first 4,000 trees, representing the burn-in phase of the analysis, were discarded, while the remaining 16,001 trees were used to calculate posterior probabilities in the majority rule consensus tree. After the analysis was finished, each run was examined using the programme Tracer v.1.5 (Drummond and Rambaut 2007) to determine burn-in and confirm that both runs had converged. ML analyses were performed with IQ-TREE (Trifinopoulos et al. 2016), using an automatic selection of the model. The final alignment and the original phylogenetic tree are available from TreeBASE under submission ID 30378.
Genealogical Concordance Phylogenetic Species Recognition (GCPSR) analysis
The Genealogical Concordance Phylogenetic Species Recognition model was applied to analyse the related species. The pairwise homoplasy index (PHI) (Bruen et al. 2006) is a model test based on the fact that multiple gene phylogenies will be concordant between species and discordant due to recombination and mutations within a species. The test was performed in SplitsTree4 (Huson and Bryant 2006) as described by Quaedvlieg et al. (2014) to determine the recombination level within phylogenetically closely-related species using a four-locus concatenated dataset. The new species and their closely-related species were analysed using this model. The relationships between closely-related species were visualised by constructing a split graph, using both the LogDet transformation and splits decomposition options.
Result
Phylogenetic analyses
In the phylogenetic tree, Samsoniellaaurantia Mongkols., et al. (TBRC 7271 and TBRC 7272) was used as the outgroup. The concatenated sequences (ITS, LSU, RPB2 and TEF) included 23 species (49 strains) and consisted of 2,620 characters with gaps (ITS, 478; LSU, 745; RPB2, 717; and TEF, 680).
The final value of the highest scoring tree was –11,790.345, which was obtained from the ML analysis of the dataset (ITS+LSU+RPB2+TEF). The parameters of the GTR model used to analyse the dataset were estimated, based on the following frequencies: A = 0.236, C = 0.283, G = 0.272, T = 0.209; substitution rates AC = 1.00000, AG = 2.12340, AT = 1.00000, CG = 1.00000, CT = 5.43884 and GT = 1.00000, as well as the gamma distribution shape parameter α = 0.557. The selected model for BI analysis was GTR+F+I+G4 (ITS+LSU+TEF) and K2P+G4 (RPB2). The phylogenetic trees (Fig. 1) constructed using ML and BI analyses were largely congruent and strongly supported in most branches. Phylogenetic analyses demonstrated that eight new strains formed a subclade with Akanthomycestiankengensis (KY11571 and KY11572) with high statistical support in ML analysis (92% ML). Strains CQ05171, CQ05172, CQ05811 and CQ05812 clustered with A.tiankengensis into a subclade, while the new species A.beibeiensis (CQ05921 and CQ05922) and A.bashanensis (CQ05621 and CQ05622) clustered in a subclade with high statistical support (96% ML/0.98 PP; Fig. 1).
Figure 1.
Phylogenetic relationships amongst the new strains and their allies based on multigene dataset (ITS, LSU, RPB2 and TEF). Statistical support values (≥70%/0.95) are shown at the nodes for ML bootstrap support/BI posterior probabilities.
GCPSR analysis
A four-locus concatenated dataset (ITS, LSU, RPB2 and TEF) was used to determine the recombination level within Akanthomycesbashanensis (CQ05621), A.beibeiensis (CQ05921) and A.tiankengensis (KY11571, CQ05171, CQ05811). Chaiwan et al. (2022) noted that, if the PHI is below the 0.05 threshold (Φw < 0.05), it indicates that there is significant recombination in the dataset, meaning that related species in a group and recombination level are not different. If the PHI is above the 0.05 threshold (Φw > 0.05), it indicates that it is not significant, which means the related species in a group level are different. The result of the pairwise homoplasy index (PHI) test of A.bashanensis, A.beibeiensis and A.tiankengensis was 0.333 and revealed that the three species were different (Fig. 2).
Figure 2.
Results of the pairwise homoplasy index (PHI) test of closely-related species using both LogDet transformation and splits decomposition. PHI test results (Φw) < 0.05 indicate significant recombination within the dataset. The new taxon is in bold type.
Taxonomy
. Akanthomyces bashanensis
W.H. Chen, Y.F. Han & J.D. Liang sp. nov.
07F0239E-420E-5EC7-A10F-54FD0F07399D
847339
Figure 3.
AkanthomycesbashanensisA infected spider (Araneae) B, CPDA culture plate showing top (B) and reverse (C) sides of the colony D–H phialides and conidia. Scale bars: 10 mm (B, C); 10 μm (D–H).
Type.
China, Chongqing, Beibei District, Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E). On a dead spider (Araneae), 1 May 2021, Wanhao Chen, GZAC CQ0562 (holotype), ex-type living culture, CQ05621.
Description.
Spider host completely covered by white mycelium. Conidiophores mononematous, arising from the lateral hyphae. Colonies on PDA, attaining a diameter of 26–27 mm after 14 days at 25 °C, white, consisting of a basal felt, floccose hyphal overgrowth; reverse yellowish. Hyphae septate, hyaline, smooth-walled, 1.5–1.9 μm wide. Conidiophores mononematous, hyaline, smooth-walled, with single phialide or whorls of 2–4 phialides or verticillium-like from hyphae directly, 12.1–20.5 × 1.5–2.1 μm. Phialides consisting of a cylindrical, somewhat inflated base, 11.8–12.9 × 1.3–1.6 μm, tapering to a thin neck. Conidia hyaline, smooth-walled, fusiform to ellipsoidal, 1.7–2.6 × 1.6–1.8 μm, forming divergent and basipetal chains. Sexual state not observed.
Etymology.
Referring to its location in Jinyun Mountain, which was formerly known as Bashan.
Additional strain examined.
China, Chongqing, Beibei District, Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E). On a dead spider (Araneae), 1 May 2021, Wanhao Chen, CQ05622.
Remarks.
Akanthomycesbashanensis was easily identified as Akanthomyces, based on the BLASTn result in NCBI and the phylogenetic analysis of combined datasets (ITS, LSU, RPB2, TEF) (Fig. 1) and it has a close relationship with another new species, A.beibeiensis. A.bashanensis was easily distinguished from A.beibeiensis by its longer phialides and smaller conidia. Jeewon and Hyde (2016) recommended that a minimum of > 1.5% nucleotide differences in the ITS regions and protein coding genes may be indicative of a new species. The pairwise dissimilarities of ITS, LSU, RPB2 and TEF sequences show 11 bp differences within 569 bp (1.93%), 19 bp differences within 881 bp (2.15%), 13 bp differences within 1070 bp (1.21%) and 4 bp differences within 973 bp (0.41%) between A.bashanensis and A.beibeiensis, respectively. The pairwise dissimilarities of ITS, LSU, RPB2 and TEF sequences show 10 bp differences within 569 bp (1.75%), 20 bp differences within 881 bp (2.27%), 19 bp differences within 1070 bp (1.77%) and 4 bp differences within 973 bp (0.41%) between A.bashanensis and A.tiankengensis, respectively. Furthermore, A.aranearum (Petch) Mains and A.ryukyuensis (Kobayasi & Shimizu) Mongkols., Noisrip., Thanakitp., Spatafora & Luangsa-ard were both absent from the available sequence in NCBI and having a spider host. Comparing with the typical characteristics (Table 2), A.bashanensis was easily distinguished from A.aranearum by its cylindrical phialide, smaller fusiform to ellipsoidal conidia and absence of synnemata and distinguished from A.ryukyuensis by absence of teleomorphs. Thus, the morphological characteristics and molecular phylogenetic results support A.bashanensis as a new species.
Table 2.
Morphological comparison of two new species with other related Akanthomyces species.
| Species | Synnemata (mm)/ Perithecia (μm) | Conidiophores (μm) | Phialides (μm)/ Asci (μm) | Conidia (μm)/ Part-spores (μm) | Reference |
|---|---|---|---|---|---|
| Akanthomycesaranearum | Cylindrical to clavate, 0.8–10 × 0.1–0.2, brown | Obovoid or ellipsoid 6–12 × 4–8 | Narrowly obclavate, 8–14 × 1.5–3 | Mains (1950) | |
| Akanthomycesbashanensis | Synnemata not observed | Mononematous, 12.1–20.5 × 1.5–2.1. | cylindrical, somewhat inflated base, 11.8–12.9 × 1.3–1.6 | fusiform to ellipsoidal, 1.7–2.6 × 1.6–1.8 | This study |
| Akanthomycesbeibeiensis | Synnemata not observed | Mononematous, 14.2–19.4 × 1.0–2.1 | cylindrical, somewhat inflated base, 7.0–9.2 × 2.1–2.5 | fusiform to ellipsoidal, 2.0–3.3 × 2.0–2.6 | This study |
| Akanthomycesryukyuensis | Synnemata not observed/ Pyriformia, 570–630 × 170–250 | Conidiophores not observed | Phialides not observed/5 wide, cap 3 wide | Conidia not observed/ 1 × 1–4 | Kobayasi and Shimizu (1982) |
| Akanthomycestiankengensis | Synnemata not observed | Erect, usually arising from the aerial hyphae | 13.9–17.1 × 1.1–1.6 with a cylindrical basal portion | Fusiform, 2.3–3.0 × 1.5–2.3 | Chen et al. (2022a) |
. Akanthomyces beibeiensis
W.H. Chen, Y.F. Han & J.D. Liang sp. nov.
5C544799-C9EC-5E4C-AF0E-CBA47AA982FF
847340
Figure 4.
AkanthomycesbeibeiensisA infected spider (Araneae) B, CPDA culture plate showing top (B) and reverse (C) sides of the colony D–J phialides and conidia. Scale bars: 10 mm (B, C); 10 μm (D–J).
Type.
China, Chongqing, Beibei District, Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E). On a dead spider (Araneae), 1 May 2021, Wanhao Chen, GZAC CQ0592 (holotype), ex-type living cultures, CQ05921.
Description.
Spider host completely covered by white mycelium. Colonies on PDA, attaining a diameter of 34–37 mm after 14 days at 25 °C, white, consisting of a basal felt, floccose hyphal overgrowth; reverse yellowish. Hyphae septate, hyaline, smooth-walled, 1.4–1.9 μm wide. Conidiophores mononematous, hyaline, smooth-walled, with single phialide or whorls of 2–6 phialides or verticillium-like from hyphae directly, 14.2–19.4 × 1.0–2.1 μm. Phialides consisting of a cylindrical, somewhat inflated base, 7.0–9.2 × 2.1–2.5 μm, tapering to a thin neck. Conidia hyaline, smooth-walled, fusiform to ellipsoidal, 2.0–3.3 × 2.0–2.6 μm, forming divergent and basipetal chains. Sexual state not observed.
Etymology.
Referring to its location in Beibei District.
Additional strain examined.
China, Chongqing, Beibei District, Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E). On a dead spider (Araneae), 1 May 2021, Wanhao Chen, CQ05692.
Remarks.
Akanthomycesbeibeiensis was easily identified as Akanthomyces according to the blast result in NCBI and the phylogenetic analysis of combined datasets (ITS, LSU, RPB2, TEF) (Fig. 1) and it has a close relationship with another new species, A.bashanensis. A.beibeiensis is easily distinguished from A.bashanensis by its shorter phialide and larger conidia. The pairwise dissimilarities of ITS, LSU, RPB2 and TEF sequences show 4 bp differences within 569 bp (0.7%), 8 bp differences within 881 bp (0.9%), 20 bp differences within 1070 bp (1.86%) and 2 bp differences within 973 bp (0.2%) between A.beibeiensis and A.tiankengensis, respectively. Furthermore, A.aranearum and A.ryukyuensis were both absent from the available sequences in NCBI and had spider hosts. Comparing with the typical characteristics (Table 2), A.beibeiensis was easily distinguished from A.aranearum by its cylindrical phialide, smaller fusiform to ellipsoidal conidia and absence of synnemata, and distinguished from A.ryukyuensis by absence of teleomorphs. Thus, the morphological characteristics and molecular phylogenetic results support A.beibeiensis as a new species.
. Akanthomyces tiankengensis
W.H. Chen, Y.F. Han, J.D. Liang & Z.Q. Liang, Microbiology Spectrum 10(5): e01975-22
D507F260-E51B-5C2F-97F1-1338749100CF
Figure 5.
AkanthomycestiankengensisA infected spider (Araneae) B, CPDA culture plate showing reverse (B) and top (C) sides of the colony D–J phialides and conidia. Scale bars: 10 mm (B, C); 10 μm (D–J).
Description.
Spider host completely covered by white mycelium. Colonies on PDA, attaining a diameter of 27–28 mm after 14 days at 25 °C, white, consisting of a basal felt, floccose hyphal overgrowth; reverse yellowish. Hyphae septate, hyaline, smooth-walled, 2.4–2.6 μm wide. Conidiophores mononematous, hyaline, smooth-walled, with single phialide or whorls of 2 phialides. Phialides consisting of a cylindrical, somewhat inflated base, 16.2–25.3 × 2.1–2.9 μm, tapering to a thin neck. Conidia hyaline, smooth-walled, subglobose to ellipsoidal, 2.4–3.8 × 2.1–3.0 μm, forming divergent and basipetal chains. Sexual state not observed.
Strains and specimen examined.
China, Chongqing, Beibei District, Jinyun Mountain (29°50'22.14959"N, 106°23'18.0744"E). On a dead spider (Araneae), 1 May 2021, Wanhao Chen, GZAC CQ0581, living cultures, CQ05811 and CQ05812; GZAC CQ0517, living cultures, CQ05171 and CQ05172; GZAC CQ0501, CQ0509, CQ0510, CQ0512, CQ0516, CQ0558, CQ0582, CQ0586, CQ0598, CQ0590.
Remarks.
Strains CQ05811 and CQ05812 were identified as belonging to Akanthomyces, based on the phylogenetic analyses (Fig. 1) and clustered with A.tiankengensis in a subclade. The characteristics of CQ05811 and CQ05812 were very closely linked with A.tiankengensis, which had fusiform conidia (2.3–3.0 × 1.5–2.3 μm) and shorter phialide (13.9–17.1 × 1.1–1.6 μm). Furthermore, the pairwise dissimilarities of ITS sequences show 3 bp difference within 569 bp between CQ05811 and A.tiankengensis (0.52%) and no difference within 569 bp between CQ05171, CQ05172 and A.tiankengensis. Thus, molecular phylogenetic results and morphologically based conclusions supported the idea that strains CQ05811, CQ05812, CQ05171 and CQ05172 were A.tiankengensis.
Discussion
Akanthomyces species are widely distributed and commonly isolated from soil, insects and spiders (Chen et al. 2018, 2019c; Shrestha et al. 2019). Amongst the 33 species, A.aranearum (Petch) Mains, A.araneicola W.H. Chen, et al., A.araneogenus Z.Q. Liang, W.H. Chen & Y.F. Han, A.araneosus W.H. Chen, Y.F. Han, J.D. Liang & Z.Q. Liang, A.coccidioperitheciatus (Kobayasi & Shimizu) Spatafora, Kepler & B. Shrestha, A.kanyawimiae Mongkols., Noisrip., Thanakitp., Spatafora & Luangsa-ard, A.neoaraneogenus (W.H. Chen et al.) W.H. Chen et al., A.lecanii (Zimm.) Spatafora, Kepler & B. Shrestha, A.ryukyuensis (Kobayasi & Shimizu) Mongkols., Noisrip., Thanakitp., Spatafora & Luangsa-ard, A.sulphureus Mongkols., Noisrip., Thanakitp., Spatafora & Luangsa-ard, A.thailandicus Mongkols., Spatafora & Luangsa-ard, A.tiankengensis W.H. Chen, et al. and A.waltergamsii Mongkols., Noisrip., Thanakitp., Spatafora & Luangsa-ard have been reported to be spider-associated fungi.
In the present study, the new strains differed from other spider-pathogenic species and had a close relationship with Akanthomycestiankengensis, based on the phylogenetic analysis. Two new species were established by combining phylogenetic analysis and morphological characteristics. Interestingly, A.tiankengensis was located at Monkey-Ear Tiankeng and found in November, indicating that it had adapted to the cold environment. Whether these new species can adapt to their environment and have special metabolic processes is worthy of further research.
The hosts of Akanthomyces species cover Hemiptera, Coleoptera, Lepidoptera, Orthoptera and Araneae (Hodge 2003; Mongkolsamrit et al. 2018; Chen et al. 2020b, 2022c). Chen et al. (2022b) noted that a host jump may be common in Simplicillium species, the spider-associated species may have originated from insects and then jumped to a spider host. An abundant diversity in insects and spiders has been discovered at Jinyun Mountain (Huang and Zhang 1991; Li et al. 2009a, 2009b; Wang et al. 2012; Huang et al. 2021; Yan et al. 2021). Whether the new species originally came from an insect host or other substrates and then jumped to a spider host, is also worthy of further research.
Mains (1950) and Vincent et al. (1988) surmised that cylindrical synnemata covered by a hymenium-like layer of phialides producing one-celled catenulate conidia were the typical characteristics of Akanthomyces. However, Chen et al. (2020b, 2020c) reported two new Akanthomyces species with mononematous conidiophores. In the present study, the two new species had mononematous conidiophores. Akanthomyces species with mononematous conidiophores are often present on the surface of moss or in open places, from which their conidia can easily be spread by airflow diffusion or other methods. Those Akanthomyces species with synnematous conidiophores often appear in the shrubbery of the original forest, litter layer or shallow soil (Hywel-Jones 1996), where air flow under the forest canopy is slow and humidity is high and where dispersal of conidia through airflow diffusion is difficult. Therefore, the presence of synnematous conidiophores may be the result of convergent evolution, which could help them to fit in their niche (Abbott 2002). Thus, the Akanthomyces species may change their type of conidiophores to increase their adaptability to different environmental conditions.
The taxonomic delimitation of Akanthomyces was originally based on morphological characteristics. Kepler et al. (2017) proposed the rejection of Torrubiella Boud. and Lecanicillium W. Gams & Zare in favour of Akanthomyces and transferred Torrubiella and Lecanicillium species into Akanthomyces, which has resulted in a combined analysis of morphological characteristics and phylogenetic analysis for the taxonomy of Akanthomyces. In this research, a PHI test and base difference rate were added, which could solve the taxonomic delimitation of cryptic species. Amongst the four loci (ITS, LSU, RPB2 and TEF), the locus TEF could not be used to distinguish A.bashanensis, A.beibeiensis and A.tiankengensis. However, any two of the three loci could easily be used to distinguish these three species. Thus, we recommend that at least two loci should be provided for the cryptic Akanthomyces species and analysis of the cryptic species with its related species should be done using the multiple methods. Furthermore, the genomics data, phylogenetic networks and haplotype analysis should be applied to cryptic species and the taxonomy of Akanthomyces made it closer to the natural taxonomy system.
Currently, the diversity of entomopathogenic fungi in some Natural Reserves and Forest Parks in different regions of China has shown that the abundant diversity of entomopathogenic fungi is present in the study areas, and there is a high species diversity in specific areas (Chen et al. 2019a, 2020a; Fan et al. 2020; Zhao et al. 2020, 2021; Zhang et al. 2021). Jiyun Mountain is located in Chongqing City and has varied altitudes, abundant plant and animal resources, which have nurtured abundant fungal resources (Zhou et al. 2012a, b). In this research, abundant Akanthomyces specimens were found at Jinyun Mountain and further attention needs to be paid to the diversity of other entomopathogenic fungi in Chongqing, China.
Supplementary Material
Citation
Chen W-H, Liang J-D, Ren X-X, Zhao J-H, Han Y-F (2023) Study on species diversity of Akanthomyces (Cordycipitaceae, Hypocreales) in the Jinyun Mountains, Chongqing, China. MycoKeys 98: 299–315. https://doi.org/10.3897/mycokeys.98.106415
Funding Statement
This work was funded by National Natural Science Foundation of China (31860002, 81960692), “Hundred” Talent Projects of Guizhou Province (Qian Ke He [2020] 6005), Science and Technology Foundation of Guizhou Province (Qiankehejichu [2020]1Y060), Program of Innovative Scientific and technological Talent Team of Guizhou Province (2020-5010), Construction Program of Guizhou Engineering Research Center (Qian Fa Gai Gao Ji 2020-896).
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 National Natural Science Foundation of China (31860002, 81960692), “Hundred” Talent Projects of Guizhou Province (Qian Ke He [2020] 6005), Science and Technology Foundation of Guizhou Province (Qiankehejichu [2020]1Y060), Program of Innovative Scientific and technological Talent Team of Guizhou Province (2020-5010) and the Construction Program of Guizhou Engineering Research Center (Qian Fa Gai Gao Ji 2020-896).
Author contributions
W.H. Chen: Writing-original draft, Data curation, Formal analysis, Funding acquisition, Resources; Jiandong Liang: Writing-review and editing, Funding acquisition, Resources; Xiuxiu Ren: Writing-review and editing, Resources; Jiehong Zhao: Writing-review and editing, Funding acquisition; Yanfeng Han: Writing-review and editing, Funding acquisition.
Author ORCIDs
Wan-Hao Chen https://orcid.org/0000-0001-7240-6841
Jian-Dong Liang https://orcid.org/0000-0002-3939-3900
Yan-Feng Han https://orcid.org/0000-0002-8646-3975
Data availability
All of the data that support the findings of this study are available in the main text.
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All of the data that support the findings of this study are available in the main text.