Analysis Of Genetic Structure Of Rhizoctonia Solani In Anhui And Transcriptome Of Rice Response To Infection

Rice sheath blight is one of the three major diseases of rice in the world.With the promotion of hybrid rice and the replacement of varieties,single chemicals use and iteration of planting patterns,rice sheath blight has become increasingly serious.It has become one of the main factors restricting the steady production of rice in China,and Anhui province as a large rice-growing province bear the brunt.Therefore,in this study,we collected and isolated of Rhizoctonia solani in representative rice production areas of representative counties and cities in Anhui Province in 2018,and analyzed their biological characteristics,pathogenicity,genetic background differences,and pathogenicity differences in rice infection strains posttranscriptional differences in rice response mechanisms.The genetic structure of Rhizoctonia solani in different regions of Anhui Province was clarified,and the biological characteristics,pathogenicity and genetic background of the strains were clarified,and the transcriptome was combined to find Rhizoctonia solani infecting rice mechanism of action at the molecular level.It provides important theoretical basis and practical reference for the prevention and control of rice sheath blight.The results are as follows:1.Isolation and identification of rice sheath blight pathogen.Water agar rapid isolation method was used to isolate rice sheath blight disease samples collected from 15 cities and counties in Anhui Province,and verified by Koch’s rule,a total of 94 pathogenic bacteria were obtained.After morphological identification and molecular biological specialization identification,94 strains were identified as Rhizoctonia solani,and confirmed to be AG1-ⅠA hyphae fusion group.2.Biological characteristics and pathogenicity of Rhizoctonia solani.The growth rate,the number of sclerotia,and the quality of the sclerotia of 94 pathogenic bacteria were measured.There were some differences in the growth rate,the number of sclerotia,and the quality of the sclerotia of different strains.Among them,fast-growing strains,fewer sclerotia and a few strains are the dominant strains in Anhui Province.Cluster analysis of the data results using SPSS showed that the growth rate,the number of sclerotia,and the quality of the sclerotia were not significantly related to the geographic origin of their strains.There was a strong positive correlation between the number of sclerotia and the quality of the sclerotia,and the curve model conformed to the Logistic model.The pathogenicity of the strains was determined by in vitro leaf inoculation method and the results were analyzed by SPSS.It showed that the pathogenicity of different strains was significantly different,and there were different pathogenicity strains in various places.The most pathogenic and weakest strains were in Hexian and Langxi.It showed that there were significant differences in pathogenicity among different strains,and there was no correlation between the pathogenicity,growth rate and sclerotia of the strains.Cluster analysis showed that there was no significant correlation between the pathogenicity of strains and geographical origin.3.Analysis of genetic diversity and genetic differentiation of Rhizoctonia solani.The genetic diversity of 94 strains was analyzed by PCR amplification with three ISSR primers.A total of 51 loci were amplified,49 polymorphic loci,and the percentage of polymorphic loci was 96.08%.According to the analysis of Popgene software and NTsys software based on different geographical sources.Amount them,the genetic diversity of Rhizoctonia solani in Xiuning County was the most abundant.The Nei’s gene diversity index,Shannon information index,and effective allele number Ne were the highest,being 0.2980,0.4425 and 1.5244 respectively.The total population genetic diversity Ht was 0.3048,the average population genetic diversity Hs was 0.1327,and the average population differentiation coefficient Gst coefficient was 0.4040.The UPGMA method divides the strains into four groups under the genetic similarity coefficient of 0.65.Most strains of the same geographical origin are clustered into one group.For different populations,the population was divided into three groups under the genetic similarity coefficient of 0.85.Southern Anhui,Northern Anhui,and Central Anhui are distributed in all groups.The above results indicate that the genetic diversity among the populations of Rhizoctonia solani in Anhui Province is rich,the genetic differentiation is obvious,and the differentiation mostly comes from within the population,and there is a certain gene flow among the population.The genetic relationship between the strains is related to their geographic location,but the genetic relationship between different populations in Anhui is relatively close,and the genetic relationship is not related to pathogenicity,sclerotia and growth rate.4.RNA-seq analysis of rice sheath blight infecting rice.AHHX(strongly pathogenic strains)and AHLX(weakly pathogenic strains)with the most significant difference in pathogenicity were used to infect the main hybrid rice variety "Huiliangyou 996" of Anhui 48 hours later to extract RNA and perform RNA-seq sequencing analysis.Using Oryza sativa genome sequence NCBI(https://www.ncbi.nlm.nih.gov)as a reference,a total of 25028 expressed genes were identified through bioinformatics analysis.Using AHHX as a control,a total of 443 up-regulated differentially expressed genes and 738 down-regulated differentially expressed genes were obtained.Further analysis by databases such as GO and KEGG.The down-regulated differentially expressed genes are enriched in four pathway of photosynthesis,phenylalanine,tyrosine and tryptophan biosynthesis of carotenoids,ubiquinone and other terpenoid quinones.This indicates that these pathways or related genes are one of the main mechanisms for the defense response of rice to Rhizoctonia solani infection.