Identification And Biological Function Research Of Mycovirus From Trichoderma Spp.

Mycovirus,as the research hot area in recent years,is a kind of virus that infects filamentous fungi,mushrooms and yeasts and replicates and proliferates in the host.At present,the mycovirus from plant pathogens are the research focus for the capability to weaken the pathogenicity of the pathogen.but the research about mycoviruses of Trichoderma spp.is still in the initial stage.Until 2020,9 strains of mycoviruses were explored out,the resources of mycovirus are limited,so the researches of exploring mycovirus,obtaining the genome sequences and gene functions,diversity,and the affection on the biological characteristics and biocontrol function have the important theoretical and applied value.In 2014-2016,a large number of Trichoderma spp.strains were isolated from Xinjiang,Inner Mongolia and Heilongjiang.According to the morphological differences of the same species of Trichoderma spp.,it was speculated that mycoviruses should be existing.Three strains of Trichoderma carrying mycoviruses were obtained.Complete genome sequence analysis and classification status of Trichoderma T673 mycovirus were analyzed.At the same time,the effects of the virus on the biological characteristics and biocontrol function of Trichoderma were evaluated.The details of research contents and results are as the followings.1.Screening of Trichoderma strains with mycovirus.Morphological differences from the same strains were analysis for the Trichoderma spp.strains from Xinjiang,Inner Mongolia and Heilongjiang,collected during 2014 and 2016.120 strains of Trichoderma spp.were screened for ds RNA extraction.Three strains suspected carrying mycovirus were screened,namely,T.harzianum 673(T673),T.citrinopsis 703(T703)and T.citrinopsis 827(T827).2.Complete genome sequencing and classification status of mycovirus from T673.The extracted ds RNA was sequenced by Metagenome,and most of the genomic fragments were obtained.On this basis,the 5’ and 3’ terminal sequences were obtained by RACE technology,so the whole genome sequence of the virus was obtained.The mycovirus of T673 contains two subgenomic segments ds RNA1 and ds RNA2.ds RNA1 contains 1693 bp with one ORF encoding Rd Rp,ds RNA2 comtains1458 bp with one ORF encoding hypothetical protein.Phylogenetic analysis showed that the mycovirus belonged to an new species of partitiviridae,which was named Trichoderma Harzianum Partitivirus 2(Th PV2).3.The effect of mycovirus on the biological characteristics of Trichoderma.Through the combination methods of protoplast and ribavirin,the platform of mycovirus elimination from Trichoderma was built,and the mycovirus-free strain T673-F was obtained.Comparing with T673 and T673-F,on the media of PDA,CMD and CZA,the Th PV2 could slow down mycelial growth,and in the PD medium,the biomass of T673 could be reduced by Th PV2.However,on the medium of PDA,Th PV2 could improve the spore production of T673 1.12 times than T673-F.Adding 10% T673 fermentation broth into the T673-F fermentation broth,the conidiospore production of T673-F could be increased by 0.35 times,while adding 10% T673 fermentation broth into the T673-F induced fermentation broth,the chlamydospores production of T673-F could be increased by 0.3 times.4.Effect of mycovirus on Biocontrol characteristics of Trichoderma spp.In the antagonistic experiment with plant pathogens(Alternaria solanacearum,Fusarium oxysporum,Botrytis cinerea,Phytophthora capsici,Fusarium graminearum),T673 had higher inhibition rate than T673-F except for Rhizoctonia solani.and we also found there was the phenomenon of hyphal twining and fusion,so it was speculated that there might be horizontal transmission of Th PV2 between the Trichoderma and pathogen,however,Th PV2 was not detected from the pathogen,which indicated that the incompatibility of interspecies should be existed.In the interaction system of Trichoderma-Th PV2-cucumber-pathogen,both T673 and T673-F had the effect of disease resistance and growth promotion,but T673 possessing more capability to improve biocontrol effectivity of T673 to F.oxysporum f.sp.cucumerinum than T673-F,while T673-F could promote the growth of cucumber better than T673,moreover,T673 possesses more capability to promoting cucumber precocity than T673-F.In the field experiment,T673-F was detected the capability of promoting growth purple cabbage in the field.