Diversity Of Viruses In Rhizoctonia Spp. From Australia And Study On The Disease Control Potential Of A Hypovirulent Strain

Rhizoctonia spp.are a large group of soil-borne saprophytic pathogens with a wide host range and cause severe damage worldwide.Mycoviruses are a kind of viruses that can parasitically infect fungi.Some fungal viruses reduce host virulence and thus have great biocontrol potential for plant fungal diseases.In addition,exploring new viruses in Rhizoctonia spp.will help to study the diverdity and evolution of viruses.In this study,the biological characteristics of 37 strains of the genus Rhizoctonia isolated from Australia were investigated,the biocontrol potential of a hypovirulent strain to rapeseed damping off was determined,and the fungal viruses diversity were analyzed.Six new Bunyaviruses were identified from two hypovirulent strains.The results of this study provide a new biological material for the biological control of rapeseed damping off and a new virus resource for the classification and evolution of fungal viruses reserach.The main results are as follows:Among the 37 strains of the genus Rhizoctonia,17 were multinucleate Rhizoctonia strains and 20 were binucleate Rhizoctonia strains.These strains were different from virulent strains isolated from China in colony morphology,growth rate and other biological characteristics.Among them,24 strains showed avirulence or hypovirulence to rapeseed,and thus were referred as hypovirulent strains.Hypovirulent strain 4917 controlled damping off caused by a virulent strain by67.33% when applied to the vegetative soil near the roots of rapeseed seedlings at the one-tip one-leaf stage.From 1555 contigs in the virome data,fungal viruses were assembled and 78 were detected in the 24 hypovirulent strains by RT-PCR.According to the BLASTx searching results,the 78 viruses were found to be distributed in 12 known taxa.Among them,68 viruses were found to have less than 80% similarity to them best match viruses.Six new Bunyavirus were obtained from the hypovirulent strains 4917 and 8481 They were named as Rhizoctonia solani bunyavirus 1(Rs Bun V1),Rhizoctonia solani bunyavirus 2(Rs Bun V2),Rhizoctonia solani bunyavirus 3(Rs Bun V3),Rhizoctonia solani bunyavirus 4(Rs Bun V4),Rhizoctonia solani bunyavirus 5(Rs Bun V5),and Rhizoctonia solani bunyavirus 6(Rs Bun V6).And the full length of L-RNA segments suquences were cloned by using RACE terminal cloning technique.The six new Bunyavirus L-RNA segments were 12628 nt,12497 nt,12459 nt,12428 nt,12470 nt.They shared the same 5 ’ end and 3’ end sequences,forthemore,the 3’ end sequences were exactly reverse complementary to the 5 ’ end sequences.There was a complete Open Reading Frames(ORF)encoding an RNA-dependent RNA polymerase(Rd Rp)containing a conserved motif SDD in the complementary sequence of each L-RNA segment.According to the results of phylogenetic analysis based on viral Rd Rp,the Rd Rp of the six new Bunyavirus species and the Rd Rp of the three representative viruses whose evolutionary relationships ranged from near to far were analyzed by multiple comparison.The similarity was 12.55%～13.78% to Tulasnella bunyales like1 in the family Tulasviridae,and 11.30%～12.15% to Sclerotinia sclerotiorum bunyavirus 1,the nearest phylogenetically related fungal Bunyavirus,and6.78%～8.05% to Kigluaik phantom virus.Therefore,it is suggested that a new family should be proposed to accommodate these six Bunyaviruses.According to the conserved terminal sequence of L-RNA segments,a universal primer was designed and used to detect other segments of bunyaviruses in strains4917 and 8481.Six smaller segments were obtained.Two segments contained a Glycoprotein structural domain,one segment contained a Zinc-Lipoprotein structural domain,and three segments contained a Nucleoprotein protein structural domain,therefore,the six segments were presumed to be M-RNA segments and S-RNA segments of these newly discovered Bunyaviruses.The horizontal transmission test showed that the viruses in strain 4917 could not transfer to the virulent Rhizoctonia strains through hyphal fusion.In conclusion,biological characteristics of 37 strains of the genus Rhizoctonia from Australia were preliminarily investigated,the biocontrol potential of a hypovirulent strain to rapeseed damping off was determined.Six new Bunyaviruses were identified from hypovirulent strains 4917 and 8481.The findings enrich the diversity of fungal viruses and provide a new biological resource for the biological control of rapeseed damping off.