Isolation,Identification And Bioactivity Of Entomogenous Fungi In Soils Of Sichuan Province,China

Entomogenous fungi(EFs)are important economic fungal resources which play a key role in the population control of agricultural and forestry pests,as well as an important source of traditional Chinese medicine.There are rich EFs in soil,therefore,it is of great significance to collect and separate entomogenous fungi from soil,for the development and utilization of biopesticides.In order to understand the biodiversity of EFs in soil of China,this research is intended to investigate the distribution and biological activity of entomogenous fungi in the soils of Sichuan Province which is an area with high altitude fluctuation,changeable geographical conditions,complex vegetation types and rich biodiversity.The soil samples of different environmental conditions in representative areas were collected,and the EFs strains were isolated and purified by selective medium.The morphological,molecular biological and phylogenetic tree methods were used to identify the strains.The EFs biodiversity in Sichuan were assessed,and the biological activities of representative strains against Spodoptera litura and Bemisia tabaci were determined.The main results are as follows:(1)A total of 132 soil samples were collected,and 571 fungal strains were isolated from 127 soil samples,the isolation rate was 96.2%,and the average number of strains per soil sample was 4.3.Vegetation types affected the fungal distributions in soil.Among them,38 strains of fungi were isolated from 100%vegetable soil samples,with an average of 4.2strains in each soil sample;12 strains were isolated from 75%oil crop soil samples,with an average of 3.0 strains;192 strains were isolated from 93.3%soil samples of food crops,with an average of 4.3 strains;227 strains were isolated from 98.0%weed soil samples,with an average of 4.4 strains;40 strains were isolated from 100%orchard soil samples,with an average of 4.4 trees;62 strains were isolated from 100%soil samples of other trees,with an average of 4.8 trees.Therefore,the diversity of fungi in the soil samples of trees and weeds is good.(2)According to the colony morphological characteristics,conidia and sporulation structures,ITS sequence and phylogenetic tree analysises,571 strains were identified,which belong to 35 genera and 64 species.There are 11 genera and 17 species of known EFs including 86 strains of Pur.lilacinum,46 strains of M.anisopliae,40 strains of Pen.citrinum,37 strains of M.marquandii,22 strains of A.terreus,18 strains of Clo.rosea,13strains of Pen.chrysogenum,9 strains of I.javanica,7 strains of I.fumosorosea,7 strains of B.bassiana,4 strains of M.carneum,3 strains of Poc.chlamydosporia.2 strains of L.coprophilum,2 strains of Pen.simplicissimum,2 strains of R.oryzae,1 strains of A.fumigatus,1 strains of Cla.sphaerospermum。The results showed that Pur.lilacinum,M.anisopliae,Pen.citrinum and M.marquandii are the dominant EF species.(3)Under latoratory conditions,the bioactivity of 34 unknown EFs strains and 7 EFs strains against the second-instar nymph of B.tabaci and second-instar larvae of S.litura were evaluated by impregnation method.The results indicated that,when at the treatment concentration of 1×10~8spores/m L,the four unknown EFs strains,P.humicola Ph SC57A06,M.cirrus Mc SC50a01 and T.trachyspermus Tt SC34B04 were pathogenic to B.tabaci with the corrected mortality of 20-30%,while the known EFs strains,I.fumosorosea If SC62A03,I.javanica IjSC44B11 and R.oryzae Ro SC49B03,respectively had the corrected mortality of 57.78%,29.63%and 28.15%.For the S.litura,the unknown EFs strains,Cla.halotolerans Ch SC55C04 and Pen.janthinellum PjSC31A03 had certain pathogenicity with the corrected mortality of 36.84%and 26.31%,respectively,while the strains I.fumosorosea If SC62A03 and I.javanica IjSC44B11 caused diseases of 63.16%and 78.95%to the insect.(4)Scanning electron microscopy(SEM)showed that conidia of I.javanica IjSC44B11 were mainly attached to the antennae,foot joints,chest,chest foot and internodal membrane after treatment with IjSC44B11 spore suspension.The most dense attachment site was the foot end of the chest foot,followed by the foot joint.After 12 hours of treatment,conidia began to germinate and grow dental tube and appressorium;24 hours later,a large number of appressorium formed;48 hours later,mycelia began to penetrate from the thinnest internode membrane of the body surface;48 hours to 72 hours,mycelia continued to grow and gradually covered the surface of S.litura.