Mechanisms of CO ₂ sensing mediated by gustatory receptors in <i>Holotrichia oblita</i> larvae

Carbon dioxide (CO₂) is a critical cue for Holotrichia oblita larvae (white grubs), a widely distributed soil-dwelling pest, in locating host plant roots. However, the molecular mechanisms underlying the perception of CO₂ in white grubs remain elusive. Here, our study demonstrated that CO₂ could induce electrophysiological and behavioral responses in third-instar larvae. Twelve to fourteen S4 sensilla and one digitiform organ potentially involved in CO₂ detection were observed on the antenna and maxillary palp of H. oblita larvae, respectively. Three gustatory receptor (GR) genes-HoblGR1, HoblGR2, and HoblGR3-were cloned and were predominantly expressed in the antennae and maxillary palps of third-instar larvae. The two-electrode voltage clamp (TEVC) recording system demonstrated that oocytes co-expressing HoblGR1 and HoblGR3, or HoblGR1, HoblGR2, and HoblGR3, gave robust responses to CO₂. Additionally, silencing of HoblGR1 or HoblGR3 for 72 h significantly reduced the electrophysiological and behavioral responses of white grubs to commercial CO₂ as well as root-emitted CO₂. Our work reveals that HoblGR1 and HoblGR3 are indispensable and sufficient for CO₂ sensing in H. oblita larvae, and this finding may aid in the development of CO₂-based trapping strategies for white grubs.