Insect chemosensory proteins (CSPs) have been proposed to capture and transport hydrophobic chemicals from air to olfactory receptors in the lymph of antennal chemosensilla. They may represent a new class of soluble carrier protein involved in insect chemoreception. However, their specific functional roles in insect chemoreception have not been fully elucidated. In this study, we report for the first time three novel CSP genes (AlinCSP1-3) of the alfalfa plant bug Adelphocoris lineolatus (Goeze) by screening the antennal cDNA library. The qRT-PCR examinations of the transcript levels revealed that all three genes (AlinCSP1-3) are mainly expressed in the antennae. Interestingly, these CSP genes AlinCSP1-3 are also highly expressed in the 5th instar nymphs, suggesting a proposed function of these CSP proteins (AlinCSP1-3) in the olfactory reception and in maintaining particular life activities into the adult stage. Using bacterial expression system, the three CSP proteins were expressed and purified. For the first time we characterized the types of sensilla in the antennae of the plant bug using scanning electron microscopy (SEM). Immunocytochemistry analysis indicated that the CSP proteins were expressed in the pheromone-sensitive sensilla trichodea and general odorant-sensitive sensilla basiconica, providing further evidence of their involvement in chemoreception. The antennal activity of 55 host-related semiochemicals and sex pheromone compounds in the host location and mate selection behavior of A. lineolatus was investigated using electroantennogram (EAG), and the binding affinities of these chemicals to the three CSPs (AlinCSP1-3) were measured using fluorescent binding assays. The results showed several host-related semiochemicals, (Z)-3-hexen-1-ol, (E)-2-hexen-1-al and valeraldehyde, have a high binding affinity with AlinCSP1-3 and can elicit significant high EAG responses of A. lineolatus antennae. Our studies indicate the three antennae-biased CSPs may mediate host recognition in the alfalfa plant bug A. lineolatus.

Insect odorant binding proteins (OBPs) are thought to involve in insects' olfaction perception. In the present study, we identified 38 OBP genes from the antennal transcriptomes of Spodoptera litura. Tissue expression profiles analysis revealed that 17 of the 38 SlitOBP transcripts were uniquely or primarily expressed in the antennae of both sexes, suggesting their putative role in chemoreception. The RPKM value analysis revealed that seven OBPs (SlitPBP1-3, SlitGOBP1-2, SlitOBP3 and SlitOBP5) are highly abundant in male and female antennae. Most S. litura antennal unigenes had high homology with Lepidoptera insects, especially genes of the genus Spodoptera. Phylogenetic analysis of the Lepidoptera OBPs demonstrated that the OBP genes from the genus Spodoptera (S. litura, Spodoptera littoralis and Spodoptera exigua) had a relatively close evolutionary relationship. Some regular patterns and key conserved motifs of OBPs in genus Spodoptera are identified by MEME, and their putative roles in detecting odorants are discussed here. The motif-patterns between Lepidoptera OBPs and CSPs are also compared. The SlitOBPs identified here provide a starting point to facilitate functional studies of insect OBPs at the molecular level both in vivo and in vitro.Insects use their sensitive and selective olfactory organs, usually their antennae, to smell the outside world. Species-specific pheromones and general plant volatiles can diffuse inside the sensilla via the multipores to penetrate the cuticular surface of antennae 1 . When the odorant molecules enter the sensillum lymph, two types of processes occur. First, perireceptor events 2 occur as the hydrophobic pheromones and/or the odorants bind to the soluble odorant binding proteins (OBPs), which deliver the pheromones and odorants to the membrane-bound olfactory receptors (ORs). After activating the ORs, the odorant molecules are subsequently quickly degraded by the odorant degrading enzymes (ODEs) 3 . After these processes, receptor events occur by the specific interaction of odorants with ORs, leading to the activation of the chemoelectric transduction machinery in the olfactory receptor neurons (ORNs) 4 . Although many researchers have focus on these two events, the events prior to membrane receptor stimulation are still poorly understood.Insect OBPs are highly concentrated in antennal sensilla and are proposed to be involved in the first biochemical step of perireceptor events in odorant reception 5,6 . Insect OBPs were first identified in the silkmoth Antheraea polyphemus, where they are uniquely expressed in male antennae and bind with the sex pheromones, so these OBPs are named as pheromone binding proteins (PBPs) 7 . Based on the

Insects use their sensitive and selective olfactory system to detect outside chemical odorants, such as female sex pheromones and host plant volatiles. Several groups of olfactory proteins participate in the odorant detection process, including odorant binding proteins (OBPs), chemosensory proteins (CSPs), odorant receptors (ORs), ionotropic receptors (IRs) and sensory neuron membrane proteins (SNMPs). The identification and functional characterization of these olfactory proteins will enhance our knowledge of the molecular basis of insect chemoreception. In this study, we report the identification and differential expression profiles of these olfactory genes in the black cutworm moth Agrotis ipsilon. In total, 33 OBPs, 12 CSPs, 42 ORs, 24 IRs, 2 SNMPs and 1 gustatory receptor (GR) were annotated from the A. ipsilon antennal transcriptomes, and further RT-PCR and RT-qPCR revealed that 22 OBPs, 3 CSPs, 35 ORs, 14 IRs and the 2 SNMPs are uniquely or primarily expressed in the male and female antennae. Furthermore, one OBP (AipsOBP6) and one CSP (AipsCSP2) were exclusively expressed in the female sex pheromone gland. These antennae-enriched OBPs, CSPs, ORs, IRs and SNMPs were suggested to be responsible for pheromone and general odorant detection and thus could be meaningful target genes for us to study their biological functions in vivo and in vitro.