Expression Characteristics And Functional Analysis Of Olfactory Receptors In Of Galeruca Daurica(Coleoptera:Chrysomelidae)

Galeruca daurica(joannis)is a new pest that has exploded in the Inner Mongolia prairies in recent years,mainly damaging the Allium plants of Liliaceae such as Allium mongolium,Allium ramosum,Allium polyrizum,and so on,which has seriously damaged grassland ecological environments.The olfactory sensory system plays a crucial role for the life activity of insects,which perceives volatiles in the external environment by olfaction,thus making information exchange.At present,the G.daurica olfactory receptor(OR)genes have been seldom studied.In this study,we investigated the functions of OR genes by analyzing the expression profiles of Gdau ORs and detecting the electroantennographical responses of male and female antennae to host volatiles and insect volatiles before and after interfering with Gdau ORco and Gdau ORs,which will provide a necessary foundation for further research on the molecular mechanisms of chemosensation of G.daurica and be of great importance in developing strategies to control its occurrence by using attractants or repellents.The main findings are as follows:1.Spatiotemporal expression patterns of the Gdau ORsq RT-PCR analysis revealed that the expression levels of 21 OR genes were significantly different in different tissues of G.daurica adults.The 15 OR genes were highly expressed in antennae,among which Gdau OR4 and Gdau ORco were most abundantly expressed.The expression levels of the 5 OR genes were higher in wings of G.daurica whereas Gdau OR2 is expressed at high levels in the abdomen.Expression analysis at different days after eclosion showed that the expression levels of ORs in the antennae were affected by days of eclosion.There were 7 Gdau ORs that showed the female-biased expression,suggesting that they may play a key role in the female’s behavior of searching for host plants or suitable oviposition sites.In addition,6Gdau ORs showed the male-biased expression,suggesting that they may be involved in mate seeking behavior by male recognition pheromone.Expression profiling of the G.daurica ORs laid a necessary foundation to uncover its olfactory recognition mechanism.2.The expression level changes of 21 Gdau ORs induced by two host volatilesThe results showed that both diallyl sulfide and diallyl disulfide could up-regulate the expression of Gdau OR4 and Gdau ORco in the antennae of male and female adults of G.daurica,and the expression levels reached the peak at 24 h,and then almost no longer increased.These results indicated that Gdau OR4 and Gdau ORco might play an important role in the chemoception process of host volatiles,which laid a foundation for further revealing the functions of Gdau ORs.3.RNAi effect of olfactory receptor genes in G.dauricaBy injecting ds OR4,ds OR11,ds OR15 and ds ORco,respectively,into the male and female adults of G.daurica,the RNA interference efficiency was detected by q RT-PCR,and the optimal interference time for the OR genes of G.daurica was determined to be48 h after injection.After silencing a single gene,the RNAi efficiencies of the 4Gdau OR genes reached more than 83 %,and had no effect on the expression levels of other OR genes.The EAG responses of male and female adults to 20 odor compounds(13 host volatiles,2 female volatiles,and 5 male volatiles)after different injection treatments were detected.The results showed that after ds OR4 treatment,the EAG responses of female antennae to 6 host volatiles were significantly reduced,while the responses of male antennae to only 4 of them were significantly reduced,indicating that Gdau OR4 may be involved in the process of host plant localization.Silencing Gdau OR11 significantly reduced the EAG response of male antennae to one female volatile,suggesting that Gdau OR11 may be a key receptor in the process of males searching for females.After ds OR15 treatment,only the EAG responses of female antennae to the three host volatiles were significantly reduced,and OR15 may play an odor recognition role in the behavior of females looking for oviposition sites.After silencing Gdau ORco,the EAG responses of female antennae to 8 host volatiles were significantly reduced,while the EAG responses of male antennae to seven compounds(1 female and 6 host volatiles)were significantly reduced.These results indicate that Gdau ORco has a functional synergistic effect with Gdau OR4,Gdau OR11 and Gdau OR15 in the olfactory recognition process of G.daurica.