Hawkmoths can smell with grooming organs on their legs

The antennae are the primary olfactory organs of insects, though other appendages, such as mouthparts and the female ovipositor, can also detect odors. A prerequisite for the olfactory function of an appendage is the presence of sensilla with porous walls and the expression of chemosensory receptors by sensory neurons housed in these sensilla. In the tobacco hawkmoth, Manduca sexta, we demonstrate that the epiphysis, a small process on the tibia of the forelegs that is used to clean the antennae, is an olfactory organ. The epiphysis carries approximately 150 sensilla with wall pores, suggesting an olfactory function. Additionally, the epiphysis expresses a variety of chemosensory receptor genes. We identified the expression of ORCo, the obligate co-receptor of odorant receptors (ORs), as well as 54 tuning ORs. Moreover, the epiphysis expresses 22 ionotropic receptors (IRs), including the co-receptors IR8a, IR25a, and IR76b, and 33 gustatory receptors (GRs). Several of these IRs and GRs had not previously been found in the antennae or other appendages. Electrophysiological recordings from isolated epiphyses revealed responses to odorants from several chemical classes, host plant leaves, and the female pheromone gland. The strongest responses were elicited by acids and the amine pyrrolidine. Epiphysectomy did not affect courtship or foraging behavior; however, epiphysectomized females were less likely to reach a host plant than controls. Our study reveals the epiphysis of M. sexta to be a previously unknown olfactory appendage with a broad and partly unique chemosensory repertoire. Because the epiphysis is a constitutive feature of lepidopteran insects, its olfactory function may be present in most moths and butterflies.