Studies On Entomogenous Fungi In The Anhui Province And Antimicrobial Activities Of The Strain RCEF6202

Resources investigation of Cordyceps and its allies is a very significant task for mycologists. In 2012 and 2013, collection of Cordyceps specimen from the Guniujiang Nature Preserve of Qimen and Yaoluoping Natural Reserve of Yuexi in the Anhui Province was conducted several times. The discovery of new species riching Anhui resource library of entomogenous fungi, exploring the biological characteristics of Cordyceps and solving problems pazzled the academics are the theoretical value of this study. Screening the species with medical value and their exploitation and utilization have significant economic value and social value.Cordyceps sensu lato was divided into six genera encompassing Cordyceps sensu stricto, Metacordyceps, Ophiocordyceps, Tyrannicordyceps, Polycephalomyces and Tolypocladium. Referring to original documents and MycoBank and GenBank authority database, Cordyceps and its allies were accurately identified, based on traditional morphological characteristics and a molecular analysis of the internal transcribed spacer rDNA sequence. To reflect changes in Article 59 of the Melbourne Code of the International Code of Nomenclature for algae, fungi, and plants (ICN), we classified and named these fungi by the new taxonomic system. Additionally, some Tolypocladium strains, including T. inflatum, T. geodes, T. terricola, T. sinense and T. cylindrosporum, were known to produce Cyclosporin A, the well-known immunosuppressant drug. Oxford cup assay was applied to test antimicrobial metabolites of T. cicadae. The main results and conclusions are reported as follows:Ophiocordyceps nutans is the dominant population in Yaoluoping Natural Reserve, followed by Cordyceps militaris. And other species in this region mainly include Metacordyceps martialis, Isaria farinosa, I. tenuipes and Beauveria bassiana.The main species of entomopathogenic fungi collected from Guniujiang Nature Preserve are Isaria cicadae, Tolypocladium cicadae, Polycephalomyces kanzashianus, Ophiocordyceps gracilioides and a new species Ophiocordyceps sp.Over the past 100 years, the famous traditional Chinese medicine Isaria cicadae was regarded as the anamorph of Cordyceps cicadae. Its anamorphic isolate was obtained from ascospores discharged for the first time and shares 100% homo logy with the stroma of C. cicadae specimen. Meanwhile, 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 anamorphic stage of C. cicadae. Morphological characteristics and molecular data supported the transfer of C. cicadae to the genus Tolypocladium and the new combination Tolypocladium cicadae (S.Z. Shing) Y.P. Cao & C.R. Li comb. nov. was proposed. Our results indicate that there is no relationship between Tolypocladium cicadae and Isaria cicadae. The molecular phylogenetic analysis of T. cicadae revealed a new clade and suggested that T. cicadae was a independent taxon in the genus Tolypocladium. The teleomorph and anamorph of T. cicadae were clustered into one group and strongly supported by bootstrap proportions (BP=100%). A sister-group relationship between the clades of T. paradoxum and T. cicadae was also strongly supported (BP=100%).In addition, the survey found that hyperparasitism in the genus Cordyceps s.1. was widespread. The common mycoparasite Polycephalomyces sp. of T. cicadae and P. kanzashianus as well as the mycoparasite of O. nutans, P. ramosus and Polycephalomyces sp.1 were reported. P. ramosus was a new record species in Anhui Province, and the other two were two new ones. The molecular phylogenetic study within Polycephalomyces provided a high support that the Polycephalomyces strains RCEF6203 and RCEF6207 were placed in one group and revealed that the two mycoparasitic strains were the one species. P. ramosus and Polycephalomyces sp.1 were independent in this analysis and the latter was clustered into another group with P. yunnanensis (Unpublished species). The genetic relationship between them were very close. Some ITS sequences and their Latin name may be inconsistent in Genbank database.At the concentration of 10 mg/mL the ethyl acetate extracts from fermentation broth of the strain RCEF6202 showed a high inhibition to the fungi Aspergillus niger and Candida albicans. And high performance liquid chromatography (HPLC) method was used to rule out the possibility of these extracts containing cyclosporin. The ethyl acetate extracts from mycelia and fruiting body had no antimicrobial activities.