Functional Characterization Of Odorant Receptors For β-caryophyllene In The Asian Corn Borer,Ostrinia Furnacalis

Insects developed a highly complex olfaction system to recognize various chemical signals in the environment for living,such as mate recognition,food searching,host plant finding,oviposition-site selection,avoidance against natural enemy and so on.Therefore,clarifying the mechanism of odorant perception can help us to develop insect behavior regulators such as attractants and mate disrupters.Ostrinia furnacalis is a pest of maize,which causes great losses and affects quality of maize in China.In this species,54 ORs have been identified,in which functions of 8 pheromone receptors(PRs)and Orco had been analyzed in details.Other ORs should be same important but only OfurOR27 was identified to be sensitive to three compounds,nonanal,octanal and 1-octanol.In this study,three female-biased odorant receptors(OfurOR15,OfurOR39 and OfurOR53)were selected to identify the odorants that associated with female behaviors.Furthermore,ORs in perception of these odorants were screened to clarify the molecular mechanism of chemosensation for these two compounds.This study will provide a theoretical basis for the development of new pest control methods.The main research results are as follows:1.Functional analysis was carried out for the three female-biased odorant receptors of O.furnacalis.The three female-biased odorant receptors(OfurOR15,OfurOR39 and OfurOR53)was cloned and constructed to expression vectors.The in vitro functions of these three odorant receptors were analyzed by Xenopus oocytes heterologous expression system.As the results,OfurOR15 and OfurOR53 which in the new PRs clade did not responded to the tested 235 odorants(including 43 sex pheromones,51 egg surface odorants,and 141 plant volatiles);while OfurOR39 was identified in responding of two plant defense volatiles,β-caryophyllene and methyl jasmonate,with the response values of 31.67±4.773 nA and 18.33±4.014 nA,respectively.EAG results showed that the female antennae responded differently to β-caryophyllene and methyl jasmonate at different concentrations(10 ng、100 ng、1 μg、10 μg、100 μg、1 mg).2.Identified the effect of β-caryophyllene for oviposition behavior in O.furnacalis.As the results,β-caryophyllene significantly repelled the oviposition behavior in a low concentration(4.4×102 ng/cm2).Interestingly,the effect was reduced while increasing the concentration of β-caryophyllene.At the concentration of 4.4×10-3 ng/cm2(total amount is 1 ng),no significant difference was found between CK and test compounds.49.43±9.640%females laid eggs on the plastic film with β-caryophyllene;at the concentration of 4.4×10-2 ng/cm2(total amount is 10 ng),27.76±6.023%females laid eggs on the plastic film withβ-caryophyllene,displaying significant repel effect;at the concentration of 0.44 ng/cm2(total amount is 100 ng),44.56±1.696%females laid eggs on the plastic film withβ-caryophyllene,displaying significant repel effect.3.Other ORs in responding of β-caryophyllene and methyl jasmonate were screened.In total,19 odorant receptors of O.furnacalis other than OfurOR15 and OfurOR53 were selected and functionally analyzed using Xenopus oocytes expression system.Among them,OfurOR14 was identified to be a sensitive receptor in responding of β-caryophyllene,whereas no more ORs were found for detecting methyl jasmonate.The average response values of OfurOR14 in responding of β-caryophyllene was about 434.0±70.61 nA and the EC50 value was about 1.324×10-5 M。In summary,this study targeted female-biased odorant receptors in O.furnacalis,and identified that OfurOR39 was the receptor in responding of two important plant defense volatiles,β-caryophyllene and methyl jasmonate,by Xenopus oocytes expression system.Behavioral analysis indicated that β-caryophyllene was a repellent against the oviposition behavior of female in O.furnacalis.Furthermore,19 ORs were screened in which OfurOR14 was identified to be a high-sensitive odorant receptor for β-caryophyllene.However,no more receptors were found in responding of methyl jasmonate.This study provides a basis for clarifying the olfactory mechanism in detecting plant defense volatiles in O.furnacalis,and provides theoretical guidance for the development of new green pest control methods for this pest.