Genetic Diversity Of The AvrPit Alleles In Rice Blast Populations

Rice(Oryza sativa.L)is one of the most widely consumed food crops around the world.However,rice blast caused by the filamentous ascomycete fungus Magnaporthe oryzae,is one of the most devastating diseases of rice and caused severely lost throughout the world.Field test show that,due to the high variability of Magnaporthe oryzae,rice resistant varieties gradually lose their resistance to Magnaporthe oryzae after being cultivated widely and become susceptible varieties.Therefore,to breed resistant varieties and conduct rational distribution,it is necessary to study and understand the genetic structure and variation pattern of Magnaporthe oryzae.The AvrPit has been cloned(unpublished)in the previous study.In order to elucidate the genetic structure and variation pattern of AvrPit,we study the AvrPit from 6 populations of Magnaporthe oryzae in the rice cultivation areas in south China(Guangdong and Hunan),southwestern areas(Guizhou)and northeast areas(Liaoning,Jilin and Heilongjiang).The result is as follows:1.The distribution of AvrPit alleles360 isolates from 6 populations were used for the pathogenicity identification of cv.K59(contains Pit).The results showed that the pathogenicity of each population was 93.3%,63.3%,75.0%,6.7%,73.3% and 65.0% respectively.Except for Liaoning,the pathogenicity of the other regions was more than 50%,indicating that the distribution of AvrPit from Liaoning population is the most widely,and Guangdong is the least.2.The genotypes of AvrPit allelesAccording to the sequences and genomic structure of virulence(A)gene and virulence(V)gene,3 pairs of AvrPit-specific primers were designed to PCR amplify 360 isolates.The results showed that 294 isolates were amplified the specific bands by using the primers AvrPit-CDS-F/R(amplify the CDS region),indicating that AvrPit is conservative in these isolates.The primers AvrPit-A-F/R(amplify the full-length avirulence gene)and AvrPit-VF/R(amplify the full-length virulence gene)can amplify specific bands in 152 and 175 isolates respectively,indicating that the obvious sequence differentiation of A/V in AvrPit.Combining with the analysis of genotypes and phenotypes,The 360 isolates were classified as 7 types(1,can amplify specific band;0,can't amplify specific band;the three numbers mean the CDS region,the full-length avirulence gene and the full-length virulence gene respectively): A101(1.9%),A110(35.3%),V000(7.8%),V001(10.5%),V100(1.7%),V101(36.1%)and V110(6.7%),the predominance types are A110 and V101.3.The sequence diversity of AvrPit allelesTo explore the genetic diversity and pathogenicity variation of AvrPit further,based on phenotypic/genotype combinations and geographical distribution,we selected 100 representative isolates from 6 populations for re-sequencing analysis.we compared the sequencing results with the reference isolates CHL357(avirulence)and 70-15(virulence),and analyzed the variation of AvrPit in 6 populations.The results showed that the function loss of AvrPit were caused by these reasons:(1)The insertion of transposons into CDS region;(2)The deletion of CDS region;(3)The occurrence of SNPs in CDS region;(4)The Insertion of the transposon upstream region of CDS.