The bean bug, Riptortus pedestris, is a major pest of soybeans in East Asian countries. Male-released aggregation pheromones attract both adults and nymphs, offering potential for eco-friendly pest control. However, the molecular mechanisms underlying the detection of the aggregation pheromones remain unclear. In the present study, functional analysis using the Xenopus oocyte expression system demonstrated that two ORs (OR23h and OR109d) were responsible for sensing aggregation pheromones, with the primary component (E)-2-hexenyl (E)-2-hexenoate (E2HE2H) being shared by the two ORs. Further quantitative PCR (qPCR) profiling indicated that OR109d was expressed only in male antennae, while OR23h was expressed in both sexes at similar levels. RNA interference (RNAi) assays demonstrated that dsOR23h-treatment significantly reduced the electroantennographic (EAG) response of (E)-2-hexenyl (Z)-3-hexenoate (E2HZ3H) in both sexes. Furthermore, simultaneous RNAi knockdown of the two ORs significantly reduced the male EAG response to E2HE2H and abolished male attraction to this compound. These results were consistent with the sex expression profile, demonstrating the sex and functional differentiation between the two ORs. Taken together, this study characterizes the ORs responsible for chemical perception and the associated aggregation behaviors driven by these pheromones. Thus, this study enhances our understanding of olfactory signaling in a hemipteran insect and contributes to the knowledge required for improved pest management.

odorant receptors  /  Xenopus oocyte expression and two-electrode voltage clamp  /  RNA interference  /  synergism  /  EAG response  /  behavioral preference