Screening,Identification And Characterization Of Viruses That Infected In Rhizoctonia Solani,a Pathogen Of Potato Black Scurf Disease

Potato nevus is a soil-borne fungal disease of potatoes.It is harmful to the growth of potatoes at all stages.It harms the young buds and roots and makes them rotten or unable to germinate.The black-brown particles are produced when the tubers of the infected plants become mature,making the appearance of the products poor.The pathogen of potato nevus is Rhizoctonia solani,which confers serious harm in potato producing areas all over the world,and its control is difficult.It has been found that there are a lot of virus resources in the fungus,and they can affect the pathogenicity of R.solani to a certain extent.In this experiment,we collected some sterile fungi with declining pathogenicity from potato producing areas in Inner Mongolia Province of China.The viruses carried by Rhizoctonia strains were studied biologically in order to find viruses that could be used as biocontrol materials for potato nevus disease.The contents and results of this study are as follows:1.Purification and preservation of Rhizoctonia solani strains collected from potato black nevus disease field.A total of 23 strains were collected,and the dominant group was AG3 fusion group.The strains could not fuse with each other.2.Screening of Rhizoctonia solani infecting viruses.After preliminary dsRNA screening of 23 fungal strains,8 strains of virus-carrying strains were obtained,and these strains may be co-infected with multiple viruses.Four viruses fromsix fungal isolates were identified by high-throughput sequencing using mixed RNA of eight strains.Through sequence alignment and phylogenetic analysis,the four viruses were identified as Cucumber mosaic virus,Betaendornavirus and Alphaendornavirus,and mitovirus.The sequence of CMV-Rs and RsMV-328 have been uploaded to NCBI with accession numbers of MG025947.1,MG025948.1,MG025949.1 and NC₀40563.1.3.RT-PCR detection of virus in R.solani strains.According to the dsRNA profile,Rs25 was found to be the least complex infection strain.The specific primers designed for RsbEV-3740 and RsMV-328 of the two known viruses detected the viruses in 10 fungal strains.RT-PCR amplification results showed that the two viruses were widely distributed.Of the 10 strains tested,8 had endogenous RNA virus RsbEV-3740 and 7 had mitochondrial virus RsMV-328.4.Complete sequence of mitochondrial virus RsMV-328.After RACE analysis of the two end sequences of the DNA,we obtained the 5’and 3’ end sequences of mitochondrialvirus RsMV-328,and the full length sequence of 2855 nt was obtained after splicing.5.Biological study of endogenous RNA virus RsbEV-3740.RsbEV-3740-free strains Rs25 and Rs80 were obtained by means of mycelial cutting-tip detoxification.By comparing the growth rate and colony morphology of toxic and non-toxic strains,it was found that endogenous RNA viruses could promote the growth and pathogenicity of R.solani strain Rs25 and Rs80.The growth rate of virus-carrying strains was significantly faster,and the area of disease spots was larger after inoculation to tobacco and potato plants.Endogenous RNA virus RsbEV-3740 was observed to be able to enter and replicatein non-toxic strains by mycelial fusion,Moreover,the sclerotia from R.solani could also carry a large number of viruses.