Identification Of IMD Immune Pathway Related Genes In Laodelphax Striatellus And Preliminary Study Of Its Antiviral Roles In Response To Rice Stripe Virus Infection

Small brown planthopper(Laodelphax striatellus)is an important rice pest,and rice stripe virus(RSV)transmitted by L.striatellus.It has a serious impact on the rice production in China.IMD immune pathway is an important mechanism for Drosophila to the infection of Gram-negative bacteria.Recent studies also shown that IMD immune pathway also plays an important role in the process of virus infection in arthropods such as fruit fly and shrimp.However,identification and function of IMD immune pathway related genes in planthopper have not been reported yet.To investigate the role of IMD pathway in the anti-RSV of L.striatellus,five IMD pathway-related genes(PGRP-LC,Duox,Relish,IKKα,and TBK1)were identified and its open reading frames(ORF)were cloned in this study.The expression patterns of these 5 genes in different developmental stages and different tissues from RSV-free and RSV-infected L.striatellus were analyzed,and the antiviral function of these IMD pathway-related genes in L.striatellus were further investigated using double-stranded RNA(dsRNA)interference technology.Key findings of this study are listed below:(1)Known IMD pathway-related genes were used as bait sequences,and bioinformatics methods were utilized to identify and clone the ORF of 5 important genes related to the IMD pathway from the genome of L.striatellus.The 5 identified genes,PGRP-LC,Duox,Relish,IKKα and TBK1,respectively,were further sequenced and phylogenetically analyzed.(2)The expression patterns of PGRP-LC,Duox,Relish,IKKα and TBK1 in different developmental stages and different tissues of RSV-free and RSV-infected L.striatellus were confirmed by qRT-PCR.Our results indicated that all these genes were widely expressed in various developmental stages and tissues of L.striatellus,and each gene has spatiotemporal expression specificity.In addition,RSV infection has a significant effect on the expression of these genes,indicating that IMD pathway may have antiviral role in L.striatellus responsive to RSV infection.(3)To investigate the role of the IMD pathway in L.striatellus in response to RSV infection,dsRNA injection method was used to interfere with IMD pathway related genes in virus free and RSV-infected L.striatellus.The results showed that the expression of the genes was significantly inhibited after RNA interference,and the virus accumulation in planthopper was significantly up-regulated,indicating that the IMD pathway might have an important role in antiviral response.Furthermore,the dsRNA injection method was also used to interfere with these 5 genes in virus free planthopper and viral acquisition percentage of L.striatellus was measured.Our results indicated that the virulence rate of planthopper was significantly increased in L.striatellus injected with dsRNA of IMD pathway related genes compare to the control(dsGFP injection).These results further demonstrated that PGRP-LC,Duox,Relish,IKKα and TBK1 are involved in the immune response of L.striatellus responsive to RSV.This study provides information for further research on the function of IMD immune pathway in the antiviral of L.striatellus,and also helps to understand the innate immune mechanism of vectors responsive to plant viruses.