Changes Of Susceptibility To Pesticides After E93 RNAi In Chilo Suppressalis

Chilo suppressalis Walker(Lepidoptera: Crambidae),which is one of the most serious rice pests in the world.The E93 gene is the primary response factor downstream of 20-Hydroxyecdysone(20E),which plays an important role in the metamorphosis of insects from pupa to adult.In this paper,the full-length sequence of E93 gene was obtained by reverse transcription PCR(RT-PCR),and the sequence characteristics of E93 gene was analyzed;Reverse transcription quantitative PCR(RT-q PCR)was used to detect the relative expression of E93 gene at different stages of C.suppressalis;The ds RNA of E93 was expressed in the bacterial and then fed to C.suppressalis,and the test insects were treated with chlorantraniliprole and methoxyfenozide,in order to know the sensitivity changes of test insects to insecticides after E93 RNAi.1.Cloning and molecular characteristics of E93 gene in C.suppressalis.The complete transcript sequence of E93 gene was successfully cloned by RT-PCR.The E93 gene sequences of several important model insects were obtained from NCBI database,and multiple sequence alignment was performed by DNAMAN.The results showed that the E93 gene sequence of C.suppressalis was highly similar to that of other insects.The phylogenetic analysis showed that the E93 genetic relationship between C.suppressalis and Bombyx mori was the closest,and the genetic relationship between C.suppressalis and Drosophila melanogaster was more far than others.Besides,the HTH domain exists in the E93 sequences of these insects,which indicates that HTH domain is highly conservative among insect species.2.Expression level of E93 gene in different developmental stages of C.suppressalis.The relative m RNA expression of E93 gene in different stages of C.suppressalis was determined by RT-q PCR.The results showed that the expression of E93 gene increased gradually with the increase of developmental instar of C.suppressalis.Further,expression of E93 gene in the 5th,6th and pupa stages of C.suppressalis increased significantly compared with the 1st to 4th instar larvae.3.The effects of E93 RNAi in C.suppressalis.In this study,the ds RNA injection method and ds RNA feeding method were used for RNAi.And,the feeding method is better in this study.RT-q PCR results showed that the interference efficiency of injection method was not stable,and the interference effect of feeding method was obvious.After ds RNA feeding,the E93 expression decreased significantly,while compared with the control group,the weight of the test group fed with the bacterial solution increased faster,and there was no significant difference in the molting rate.4.Changes of susceptibility to insecticides after E93 RNAi in C.suppressalis.The RT-q PCR results showed that the relative m RNA expression of E93 gene decreased significantly after RNAi and feeding with sublethal dose of chlorantraniliprole.While,E93 expression increased significantly after RNAi and feeding with sublethal dose of methoxyfenozide.Our data indicated that chlorantraniliprole inhibit the E93 expression while methoxyfenozide induce E93 expression in the rice stem borer.Besides,both the mortality rates of chlorantraniliprole and methoxyfenozide feeding increased after E93 RNAi.