Male-biased odorant receptors mediate dual perception of sex pheromones and floral volatiles in mirid bugs

Plant volatiles serve as critical cues for mate location in herbivorous insects, often interacting with sex pheromones to enhance attraction. For male Apolygus lucorum, sex pheromone lures were found to be highly attractive in field traps but showed no significant effect in laboratory assays, suggesting that host plant volatiles may influence the detection of sex pheromones. To investigate this hypothesis, we first simulated field conditions using an H-type olfactometer and obtained consistent results. Subsequently, 15 odorant receptors (ORs) exhibiting high or male-biased expression in male antennae were selected for functional characterization. These AlucORs were heterologously expressed in Xenopus oocytes, and their responses to 64 compounds were analyzed using two-electrode voltage clamp (TEVC) recordings. Among the 15 AlucORs examined, nine displayed functional responses to the tested compounds. Of these responsive AlucORs, six displayed a broad ligand spectrum, responding to both major sex pheromone component trans-2-hexenyl butyrate (E2HB) and structurally similar floral esters. In particular, recombinant AlucOR3 and AlucOR81 were highly sensitive to the two key floral esters, butyl propionate and butyl butyrate, as well as to E2HB. Phylogenetic analysis revealed a novel clade of ORs (AlucOR3-subclade) in mirid bugs that exhibited dual sensitivity to both sex pheromones and floral volatiles and was subject to strong purifying selection. Additionally, bioactive ligands combined with E2HB could influence the behavioral attractiveness of male bugs. Our findings provide molecular and behavioral evidence for a synergistic mate-location strategy where plant-derived cues enhance pheromone perception in this agricultural pest.