Spider-pathogenic fungi serve as critical regulators of spider populations in natural systems, playing irreplaceable roles in maintaining ecological balance and also serving as reservoirs of bioactive compounds. Despite recent taxonomic refinements at the generic level, their broader phylogenetic diversity remains significantly underrepresented compared to entomopathogenic fungi. In this study, we collected several novel spider-pathogenic fungi from various provinces in China and conducted comprehensive taxonomic and phylogenetic analyses. Based on integrated morphological characterization and multigene phylogenetic analyses of five loci (ITS, nrLSU, TEF1, RPB1, and RPB2), eight novel species are described and illustrated: Arachnidicola (1), Gamszarella (1), Gibellula (2), Hevansia (1), and Liangia (1) within Cordycipitaceae; Husseyia (1) within Clavicipitaceae; and Purpureocillium (1) within Ophiocordycipitaceae. Additionally, six new combinations are proposed, one Chinese new record is reported, the type specimen of one known species is revised, and five potential cryptic species are identified. Our phylogenetic analyses provide robust evidence for the taxonomic placement of Chlorocillium and Husseyia within Clavicipitaceae. Molecular clock analysis, utilizing a dataset of five loci from 648 fungal strains, estimated the stem and crown ages of Hypocreales and indicated that spider-pathogenic fungi likely emerged during the mid-Cretaceous and subsequently diversified. Multiple lineages displayed marked trends toward host specialization, suggesting that these fungi developed highly efficient parasitic strategies to exploit constrained ecological niches. This research substantially expands the documented diversity of araneopathogenic fungi, providing a robust phylogenetic framework for elucidating their evolutionary origins and diversification patterns, while offering valuable biological resources for future biotechnological and ecological applications.

Cordyceps chanhua is a traditional Chinese medicinal fungus renowned for producing a variety of bioactive compounds, including beauvericin (BEA). BEA has garnered significant attention due to its therapeutic potential and associated food safety concerns. In this study, we identified an ATP-binding cassette (ABC) transporter-encoding gene, CcT1, located within the BEA synthesis gene cluster of C. chanhua. Disruption of CcT1 resulted in a substantial decrease in BEA production. RT-qPCR analysis demonstrated that the loss of CcT1 significantly downregulated the expression of several BEA synthesis-related genes, including pyruvate kinase, branched-chain amino acid aminotransferase, and ketoisovalerate reductase. Beyond its role in BEA biosynthesis, CcT1 was found to influence hyphal growth, conidiation, conidial germination, and the oxidative stress response in C. chanhua. Additionally, the CcT1 knockout strain exhibited a reduced ability to penetrate host cuticles, highlighting the gene's role in fungal pathogenicity. These findings offer a comprehensive understanding of the multifaceted roles of the ABC transporter CcT1 in hyphal development, conidiation, BEA biosynthesis, and stress resistance in C. chanhua. Moreover, targeting CcT1 presents a promising strategy for reducing BEA content through molecular breeding, thereby enhancing the safety and efficacy of C. chanhua as a medicinal agent. IMPORTANCE:Beauvericin (BEA) is one of the bioactive components in Cordyceps chanhua, a significant medicinal fungus with widespread use in Asia and beyond. BEA also possesses mycotoxin properties, with certain cytotoxicity and potential in vivo toxicity. However, few studies report the regulation of BEA anabolism. ABC transporters are a superfamily of membrane proteins and have multiple functions such as regulating fungal metabolism. Here, we report an ABC transporter CcT1 involved in BEA synthesis. The disruption of its encoding gene CcT1 led to a 64.22% reduction in BEA content compared to the wild-type by regulating the expression levels of several BEA synthesis-related genes. It also affected hyphal growth, conidiation, spore germination, penetration, and oxidative stress resistance of the fungus. The findings in this study enrich the understanding of the function of ABC transporter in fungal metabolism and growth and development.

Ophiocordyceps, a species-rich genus in Ophiocordycipitaceae, is a holomorphic genus in which most of the species are reported with hirsutella-like anamorphs. In this study, we introduce two new species of hirsutella-like anamorphs from lepidopteran larvae (viz., Ophiocordycepstielingensis sp. nov. and Ophiocordycepskeqinii sp. nov.). Ophiocordycepsradiata (syn. Hirsutellaradiata), a new combination, exhibits a pathogenic association with a fly, and it is reported as a new geographic record from China, based on integrated morphological and molecular analyses. We provide a checklist of Ophiocordyceps species with hirsutella-like anamorphs and comprehensively review their characteristics of anamorphs and teleomorphs. These definitive findings establish a foundation for the classification and diversity of Ophiocordyceps species with hirsutella-like anamorphs.

A novel genus, Paradingleyomyces was introduced to accommodate Pa. lepidopterorumsp. nov., based on a multigene phylogenetic analysis and its distinct morphological characteristics. Maximum likelihood (ML) and Bayesian inference analyses (BI) of ITS, SSU, LSU, tef-1α, rpb1, and rpb2 sequence data shown that Pa. lepidopterorum formed an independent lineage nested between Perennicordyceps and Dingleyomyces. Morphologically, Paradingleyomyces is distinguished from Perennicordyceps by the presence of a white subiculum on the stromata of Ophiocordyceps cf. cochlidiicola. Perithecia are produced sporadically from the base to the apex of the stromata, and the secondary ascospores exhibit a notable length-to-width ratio. These characteristics distinguish Paradingleyomyces from Perennicordyceps which exhibits tortuous, branched, clavate to cylindrical stromata with rhizomorphs, parasitism of coleopteran and hemipteran larvae, and colonizes a broader range of fungal hosts. Additionally, perithecia in Perennicordyceps typically arise from the middle to the upper regions of the stromata, with secondary ascospores displaying a comparatively lower length-to-width ratio. Paradingleyomyces is morphologically identical to Dingleyomyces in its direct production of superficial perithecia on the stromata of Ophiocordyceps species. However, the phylogenetic analysis indicates that Paradingleyomyces and Dingleyomyces are not congeneric. Moreover, this study introduces another novel species, Polycephalomyces tengchongensis, and a novel sexual morph of Pleurocordyceps yunnanensis. Dimorphic phialides and conidia of Pleurocordyceps parvicapitata were observed and described for the first time based on a fresh collection from Tengchong County, Yunnan Province, China.

The species of Purpureocillium are cosmopolitan and multitrophic fungi that can infect a wide range of invertebrate hosts. This study reports the mitogenome of P. atypicola, a specialized spider pathogenic fungus. The 112,465 bp mitogenome encoded genes typically found in fungal mitogenomes, and a total of 52 introns inserted into seven genes. A comparison with three other Purpureocillium species revealed significant differences in length and intron number, primarily due to intron variation; however, there was no dynamic variation in the introns of the cox1 gene within the same species of the Purpureocillium genus. Different mitochondrial protein-coding genes showed variable degrees of genetic differentiation among these species, but they were all under purifying selection. Additionally, frequent intron loss or gain events were detected to have occurred during the evolution of the Ophiocordycipitaceae mitogenomes, yet the gene arrangement remains conserved. A phylogenetic analysis of the combined mitochondrial gene set gave identical and well-supported tree topologies. The estimated age of the crown of Ophiocordycipitaceae and Purpureocillium were around the Early Cretaceous period (127 Mya) and Late Cretaceous period (83 Mya), respectively. The results of this study advance our understanding of the genomics, evolution, and taxonomy of this important fungal group.

Cordyceps chanhua, an important cordycipitoid medical mushroom with wide use in Asia, has gained attention for its bioactive component beauvericin (BEA), which is of medicinal value as a drug lead, but also of food safety risk. Recent observations by our group revealed a significant decrease of BEA content in C. chanhua when exposed to light, but the underlying regulatory mechanisms remain elusive. In this study, a comprehensive approach combining metabolomics and transcriptomics was employed to investigate the effects of white light on the secondary metabolism of C. chanhua for elucidation of the influence of light on BEA biosynthesis in this fungus. The result showed that the genes and metabolites involved in the synthesis of D-hydroxyisovaleric acid, a precursor of BEA synthesis, were down-regulated under light exposure, while those associated with the synthesis of phenylalanine, another precursor of BEA synthesis, were up-regulated leading to elevated phenylalanine levels. It suggested that the suppressive effect of light on BEA synthesis in C. chanhua occurred primarily through the inhibition of D-hydroxyisovaleric acid synthesis, while the enhanced phenylalanine biosynthesis likely directed towards other metabolic pathway such as pigment synthesis. These results contributed to a better understanding on how light modulates the secondary metabolism of C. chanhua and provided valuable guidance for optimizing BEA production in cultivation practices.

Macro- and microscopic morphological studies and multilocus phylogenetic analysis were made on larval specimens of a ghost moth collected from a pigeon pea plantation in Huánuco, Peru. DNA sequences from the cadaver and the fungal isolates obtained represented a monophyletic clade based on the phylogeny. All morphological characters and molecular data showed that the pathogenic fungus infecting the ghost moth larvae was an unknown cordycipitoid species, herein described as, Harposporium incensis sp. nov. based on morphological features and multilocus phylogenetic analysis on the cadaver and fungus isolated from the same specimen. The far-related and ecologically different hosts of teleomorph and anamorph of this new species display a peculiar inter-phylum host jumping between the insect Trichophassus giganteus of the phylum Arthropoda and the nematode Caenorhabditis elegans of the phylum Nematoda and have biological control potential.

The genus Samsoniella was erected based on orange cylindrical to clavate stromata, superficial perithecia and conidiophores with Isaria-like phialides and to segregate them from the Akanthomyces group. In this study, based on morphological features and multigene (SSU, LSU, TEF, RPB1 and RPB2) phylogenetic analysis six Samsoniella species parasitizing spiders were collected in China. Three of them belong to known species S. alpina, S. erucae and S. hepiali. Three new species S. anhuiensis sp. nov., S. aranea sp. nov. and S. fusiformispora sp. nov. are illustrated and described. They are clearly distinct from other species in Samsoniella occurring in independent subclades. Furthermore, among the four insect-pathogenic fungi specimens collected from similar sites, three of them were identified as the new species described below. Our study significantly broadens the host range of Samsoniella from Insecta to Arachnida, marking a noteworthy expansion in understanding the ecological associations of these fungi. Additionally, the identification of both mononematous and synnematous conidiophores in our study not only expands the knowledge of Samsoniella species but also provides a basis for future research by comparing the ecological significance between these conidiophore types. In conclusion, our study enhances the understanding of Samsoniella diversity, presenting a refined phylogenetic framework and shedding light on the ecological roles of these fungi in spider parasitism.

Cordyceps chanhua on artificial media has been approved as a medicine food homology product. However, the metabolites have not been extensively studied. HPLC-HRMS analysis showed that there were 11 main metabolites in the EtOAc extract including 4 probable unknown compounds. Fumosoroseain A with anti-aging effects is the most abundant known compound and was identified from C. chanhua for the first time. The second abundant compound is N6-(2-Hydroxyethyl) adenosine, a typical metabolite of C. chanhua. All the known compounds have consistent health function with that of the fungus. HRMS, 1D and 2D NMR analyses revealed that compounds 2, 3, and 4 are new nucleosides named as chanhuanosides A, B, and C. Compound 1 is the known compound cordyrrole B isolated from C. chanhua for the first time whose structure is firstly confirmed by single crystal X-ray analysis. Bioactivity analysis revealed that 1-4 significantly inhibited pancreatic lipase activity, and strongly promoted the proliferation of RAW264.7 and 293T cells, suggesting that they might have ant-obesity, immunoregulation, and renal protection functions. Structure-bioactivity analysis revealed that the esterification on ribose can increase their bioactivity. Present metabolites study suggests that C. chanhua cultured on the artificial medium is a promising health food.

根据考古和史料证据推断,人们对虫草的认识有着5 000多年的历史,涉及文化、艺术、历史、哲学、宗教、习俗、医药及生物科学等方面.根据各阶段的特点可分为崇拜时代、本草时代、描述时代和分子时代.(1)崇拜时代(红山文化-神农本草经问世):中国考古发掘出众多玉蚕和玉琀蝉,反映在远古时期对虫草的蒙昧认识以原始的轮回崇拜为特点.至今中外仍遗留着一些古代虫草崇拜的习俗.(2)本草时代(《神农本草经》问世-1818年虫草属建立):古代苏美尔人可能使用过一种虫草入药;而中国人2 000年来一直使用白僵蚕、蝉花和冬虫夏草等入药.此时期物化观念对东西方都有较大影响,对于虫草东西方均有"动物化植物""竹根化蝉""虫草互变"等认知,反映了物化论的深刻影响.(3)描述时代(1818年-20世纪末):1818年Fries建立虫草属,揭开虫草菌科学发展300年的篇章.物化论逐渐被近代科学取代;庞大的虫草家族逐渐形成.欧洲和日本流行虫草绘画,加快了虫草知识的普及.19世纪后期兴起利用虫草菌防治害虫,并在20世纪中期形成规模较大的真菌杀虫剂产业.中国将虫草用于食品和药品的古老传统得以传承并发展,在20世纪后期形成较大产业;但欧美各国发展较慢.(4)分子时代(21世纪初以来):在现代生物科学和技术的带动和支撑下,虫草研究从上世纪末开始进入分子时代.生物药物素研究缓慢进展,而虫草的多基因系统发育、虫草侵染昆虫的分子致病机制以及以基因组学引领的组学研究则快速进展.国际和国内学术交流活跃,对于弘扬虫草文化起到有力的促进作用.

Cordyceps chanhua is an important cordycipitoid mushroom widely used in Asia and beyond. Beauvericin (BEA), one of the bioactive compounds of C. chanhua, has attracted much attention because of its medicinal value and food safety risk. In order to clear up the relationship between oxidative stress and BEA synthesis, we investigated the impact of H2O2-induced oxidative stress on the secondary metabolism of C. chanhua using untargeted metabolomics and a transcript profiling approach. Metabolic profiling of C. chanhua mycelia found that in total, 73 differential metabolites were identified, including organic acids, phospholipids, and non-ribosomal peptides (NRPs), especially the content of BEA, increasing 13-fold under oxidative stress treatment. Combining transcriptomic and metabolomic analyses, we found that the genes and metabolites associated with the NRP metabolism, especially the BEA biosynthesis, were highly significantly enriched under H2O2-induced stress, which indicated that the BEA metabolism might be positive in the resistance of C. chanhua to oxidative stress. These results not only aid in better understanding of the resistance mechanisms of C. chanhua against oxidative stress but also might be helpful for molecular breeding of C. chanhua with low BEA content.

Cicada flower, Isaria cicadae Miq., has been a traditional Chinese medicine for approximately 1600 years. Many works on its identification, bioactivities, and clinical use against some disorders have been published, but some inaccuracies and inconsistencies need to be further clarified. In combination with our > 20 years of research and application of cicada flower and examination of the literature and patents published in recent years, this article summarizes and reviews the life cycle and taxonomy, genome size and mating type, molecular systematic classification and cultivation, active ingredients, and pharmacological functions of I. cicadae.

报道了1种新的昆虫-昆虫病原菌-重寄生菌三级营养关系,其中蝉若虫被昆虫病原真菌蝉花Cordyceps chanhua感染,而蝉花又被重寄生真菌寄生.经形态学观察和分子生物学鉴定该重寄生真菌为寄生凹孢Syspastospora parasitica.柯赫氏法则验证结果表明:该重寄生真菌可以重寄生于蝉花Cordyceps chan-hua、细脚棒束孢Isaria tenuipes以及球孢白僵菌Beauveria bassiana等虫草科真菌,寄主范围广泛.

我国重要药用真菌蝉花的分类地位和学名在国内外长期混乱,因其有性型未被发现,长期以来误作为根据巴西标本命名的Isaria(Paecilomyces)cicadae.作者最近在井冈山发现了其有性型,研究了标本及分离物的形态特征及多位点系统发育特征,并与GenBank中相关种类的序列进行了对比,证明l.cicadae是物种复合群,确定蝉花是虫草科虫草属中的新种,使用传统药用真菌蝉花的古老名称将其命名为Cordyceps chanhua.蝉花子座淡橙色到淡桔黄色,子囊壳半埋生,475-602×222-319μm,子囊孢子246-360×1.5-1.8μm,分孢子圆筒形,6.4-13.8×2.1-3.1μm.孢梗束淡黄色至淡黄褐色,分生孢子长椭圆形或圆筒形,3.5-10.5×1.5-4.5μm.

Entomopathogenic fungi are one of the key regulators of insect populations in nature. Some species such as Beauveria bassiana with a wide host range have been developed as promising alternatives to chemical insecticides for the biocontrol of insect pests. However, the long-term persistence of the released strains, the effect on non-target hosts and local fungal populations remains elusive, but they are considerable concerns with respect to environmental safety. Here we report the temporal features of the Beauveria population genomics and evolution over 20 years after releasing exotic strains to control pine caterpillar pests. We found that the isolates within the biocontrol site were mostly of clonal origins. The released strains could persist in the environment for a long time but with low recovery rates. Similar to the reoccurrence of host jumping by local isolates, the infection of non-target insects by the released strains was evident to endemically occur in association with host seasonality. No obvious dilution effect on local population structure was evident by the releases. However, the population was largely replaced by genetically divergent isolates once per decade but evolved with a pattern of balancing selection and towards expansion through adaptation, non-random outcrossing and isolate migration. This study not only unveils the real-time features of entomopathogenic fungal population genomics and evolution but also provides added values to alleviate the concerns of environmental safety regarding the biocontrol application of mycoinsecticides.

蝉花是我国重要药用真菌,但长期以来其名称特别是学名的使用一直混乱.本文从古代医药典籍记载确认,"蝉花"是历史最久、使用最广的中名,对于其他中名具有优先权.其学名Isaria cicadae系Miquel(1838)根据巴西标本命名,但模式标本己失,原描述文字及插图均极其简单,与蝉花形态差异较大.其有性型迄今未发现,曾先后被认为是小蝉草Ophiocordyceps sobolifera和大蝉草Tolypocladium dujiaolongae(=Cordyceps cicadae Shing).本文根据文献考证澄清了国内外关于小蝉草和大蝉草的误用,分析了长期以来大量日本文献中的命名混乱对我国认知蝉花的影响,并质疑I.cicadae在中国及其他地区的分布,提出它是一复合种,因此蝉花的分类地位及学名需要进一步研究.

Isaria cicadae is one of the fungi used in traditional Chinese medicine with the longest tradition. It is used not only as a herbal medicine but also as a health food in Asia, together with cultured cordyceps and mycelia of the fungus used as substitute. However, the differences in their metabolite are unknown. Using a high-performance liquid chromatography-mass spectrometry (HPLC-MS)-based metabolomic method, we found that the fungus varies in its metabolism during growth on wild insects, artificially raised insects and artificial medium. There were 109 discriminatory metabolites detected in the samples by orthogonal projection to latent structure discriminant analysis and one-way ANOVA. High level of nonribosomal peptides (NRPs) only existed in the insect portions of the wild cordyceps (WI) and cultured cordyceps (CI), revealing that immunostimulation of the host insects enhanced the synthesis of NRPs in the fungus. The finding of a significantly higher level of sphingolipids in both the insect portions (WI, CI) and the coremia of the wild cordyceps (WC) and cultured cordyceps (CC) but not in cultured mycelia (CM) of I. cicadae implies that the immunostimulation of the live insects can induce the fungus to produce more sphingolipids, and this enhanced ability is probably heritable. Apart from NRPs and sphingolipids, the insect portions also contained higher levels of bioactive compounds such as lateritin, anisomycin, streptimidone and ustiloxins. In contrast, the coremium groups (WC, CC) and CM contained 10-fold less NRP but much higher levels of sanative metabolites such as tocotrienol, 3'-deoxy-hanasanagin, γ-aminobutyric acid and phospholipids than the insect portions. The significantly higher content of antioxidants in WC, CC and CM than in WI and CI suggests that environmental oxygen has a significant effect on the metabolites. The temperature stress which the wild cordyceps encounters during growth is responsible for the relatively high content of trehalose. These findings indicate that the immunity of the host insect and growth environment have a strong impact on the metabolomic variation in Isaria cicadae. The variation in metabolites suggests differential utilization value for the insect portions, coremia and mycelia of the fungus.

A common cicada pathogen occurring in bamboo forests of southern China is described as a new species, Tolypocladium dujiaolongae. The previously unknown anamorphic state was obtained from discharged ascospores. Based on its morphological characteristics and a molecular analysis of the internal transcribed spacer rDNA sequence, we found that the asexual features of T. dujiaolongae are similar to those of Tolypocladium species. The typical characteristics of the fungus are described. The fruiting bodies formed on artificial culture media were identical to those of natural specimens, strongly supporting the idea that the isolated strain is the asexual stage of T. dujiaolongae. Our results indicate that there is no relationship between Tolypocladium dujiaolongae and the famous traditional Chinese medicine Isaria cicadae.

Cordycipitoid fungi, a large group of Cordyceps sensu lato species, contain more than 1000 species reported worldwide, some of them with a great value and application potential. New taxa are contin...

Background Ascomycete Cordyceps species have been using as valued traditional Chinese medicines. Particularly, the fruiting bodies of Cordyceps cicadae (syn. Isaria cicadae ) have long been utilized for the treatment of chronic kidney disease. However, the genetics and bioactive chemicals in this fungus have been largely unexplored. Results In this study, we performed comprehensive omics analyses of C. cicadae , and found that, in contrast to other Cordyceps fungi, C. cicadae produces asexual fruiting bodies with the production of conidial spores instead of the meiotic ascospores. Genome sequencing and comparative genomic analysis indicate that the protein families encoded by C. cicadae are typical of entomopathogenic fungi, including the expansion of proteases and chitinases for targeting insect hosts. Interestingly, we found that the MAT1-2 mating-type locus of the sequenced strain contains an abnormally truncated MAT1-1-1 gene. Gene deletions revealed that asexual fruiting of C. cicadae is independent of the MAT locus control. RNA-seq transcriptome data also indicate that, compared to growth in a liquid culture, the putative genes involved in mating and meiosis processes were not up-regulated during fungal fruiting, further supporting asexual reproduction in this fungus. The genome of C. cicadae encodes an array of conservative and divergent gene clusters for secondary metabolisms. Based on our analysis, the production of known carcinogenic metabolites by this fungus could be potentially precluded. However, the confirmed production of oosporein raises health concerns about the frequent consumption of fungal fruiting bodies. Conclusions The results of this study expand our knowledge of fungal genetics that asexual fruiting can occur independent of the MAT locus control. The obtained genomic and metabolomic data will benefit future investigations of this fungus for medicinal uses.