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The olfactory sense is crucial for organisms, facilitating environmental recognition and interindividual communication. Ithomiini butterflies exemplify this importance not only because they rely strongly on olfactory cues for both inter- and intra-sexual behaviors, but also because they show convergent evolution of specialized structures within the antennal lobe, called macroglomerular complexes (MGCs). These structures, widely absent in butterflies, are present in moths where they enable heightened sensitivity to, and integration of, information from various types of pheromones. In this study, we investigate chemosensory evolution across six Ithomiini species and identify possible links between expression profiles and neuroanatomical. To enable this, we sequenced four new high-quality genome assemblies and six sex-specific antennal transcriptomes for three of these species with different MGC morphologies. With extensive genomic analyses, we found that the expression of antennal transcriptomes across species exhibit profound divergence, and identified highly expressed ORs, which we hypothesize may be associated to MGCs, as highly expressed ORs are absent in Methona, an Ithomiini lineage which also lacks MGCs. More broadly, we show how antennal sexual dimorphism is prevalent in both chemosensory genes and non-chemosensory genes, with possible relevance for behavior. As an example, we show how lipid-related genes exhibit consistent sexual dimorphism, potentially linked to lipid transport or host selection. In this study, we investigate the antennal chemosensory adaptations, suggesting a link between genetic diversity, ecological specialization, and sensory perception with the convergent evolution of MCGs. Insights into chemosensory gene evolution, expression patterns, and potential functional implications enhance our knowledge of sensory adaptations and sexual dimorphisms in butterflies, laying the foundation for future investigations into the genetic drivers of insect behavior, adaptation, and speciation., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Molecular Biology and Evolution.)

The northern bark beetle, Ips duplicatus , is an emerging economic pest, reportedly infesting various species of spruce ( Picea spp.), pine ( Pinus spp.), and larch ( Larix spp.) in Central Europe. Recent climate changes and inconsistent forest management practices have led to the rapid spread of this species, leaving the current monitoring strategies inefficient. As understanding the molecular components of pheromone detection is key to developing novel control strategies, we generated antennal transcriptomes from males and females of this species and annotated the chemosensory proteins. We identified putative candidates for 69 odorant receptors (ORs), 50 ionotropic receptors (IRs), 25 gustatory receptors (GRs), 27 odorant-binding proteins (OBPs), including a tetramer-OBP, 9 chemosensory proteins (CSPs), and 6 sensory neuron membrane proteins (SNMPs). However, no sex-specific chemosensory genes were detected. The phylogenetic analysis revealed conserved orthology in bark beetle chemosensory proteins, especially with a major forest pest and co-habitant, Ips typographus . Recent large-scale functional studies in I. typographus chemoreceptors add greater significance to the orthologous sequences reported here. Nevertheless, identifying chemosensory genes in I. duplicatus is valuable to understanding the chemosensory system and its evolution in bark beetles (Coleoptera) and, generally, insects.

Insects rely on olfaction for mating, finding oviposition sites, and locating hosts. Hyphantria cunea is a serious pest that severely damages forests. Differential expression analysis of olfactory-related genes between males and females is the basis for elucidating the functions of olfactory-related proteins in H. cunea . In this study, Illumina HiSeqTM 4000 high-throughput sequencing technology was used to perform transcriptome sequencing of the antennal tissues of adult male and female H. cunea . Functional annotation was conducted using the NR, Swiss-Prot, KOG, KEGG, and GO databases, and the results showed that the antennal transcriptome of adult H. cunea contained 50,158 unigenes. Differential expression analysis identified 3923 genes that were significantly differentially expressed between male and female antennae. A total of 221 olfactory-related genes were annotated, and 96 sex-biased genes were identified, including 13 odorant receptors (ORs), 48 odorant binding proteins (OBPs), 7 chemosensory proteins (CSPs), 10 ionotropic receptors (IRs), 10 sensory neuron membrane proteins (SNMPs), 2 gustatory receptors (GRs), and 6 odorant-degrading enzymes (ODEs), indicating that there were differences in olfaction between male and female H. cunea . Quantitative real-time PCR was used to verify the expression levels of 21 putative general odorant receptor genes in male and female antennae. HcunOR4 and HcunOR5 showed female-biased expression; HcunOR48 , HcunOR49 and HcunOR50 showed male-biased expression. The results were consistent with the transcriptome differential analysis. The screening of male-biased odorant receptor genes might provide a theoretical basis for the functional characterization of odorant receptors for recognizing sex pheromones in H. cunea .

Graphosoma rubrolineatum (Hemiptera: Pentatomidae) is an important pest of vegetables and herbs (e.g., Umbelliferae and Cruciferae) in China, Siberia, Korea, and Japan. Insects are highly dependent on their olfactory system to detect odorants. However, no molecular-mediated olfactory genes in G. rubrolineatum have yet been identified. In this study, we first established the antennal transcriptome of G. rubrolineatum and identified 189 candidate olfactory genes, including 31 odorant-binding proteins (OBPs), 15 chemosensory proteins (CSPs), four sensory neuron membrane proteins (SNMPs),94 odorant receptors (ORs), 23 ionotropic receptors (IRs), and 22 gustatory receptors (GRs). Additionally, phylogenetic trees were constructed for olfactory genes between G. rubrolineatum and other hemipteran insects. We also detected the expression profiles of ten OBPs, five CSPs, two SNMPs, five ORs, four IRs, and four GRs by real-time quantitative PCR. The results revealed that most genes (GrubOBP1/11/31, GrubCSP3/8, GrubSNMP1a/1b, GrubOrco/OR9/11/13, GrubGR1/4/22, GrubIR25/75h/76b/GluR1) were highly expressed in the antennae, GrubOBP13/31 and GrubCSP4/11/12 were highly expressed in the legs, while GrubOBP20 and GrubGR19 were highly expressed in the wings. Our results will enrich the gene inventory of G. rubrolineatum and provide further insight into the molecular chemosensory mechanisms of G. rubrolineatum., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Guo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

In the polyphagous insect Monolepta signata (M. signata) (Coleoptera: Chrysomelidae), antennae are important for olfactory reception used during feeding, mating, and finding a suitable oviposition site. Based on NextSeq 6000 Illumina sequencing, we assembled the antennal transcriptome of mated M. signata and described the first chemosensory gene repertoire expressed in this species. The relative expression levels of some significant chemosensory genes were conducted by quantitative real-time PCR. We identified 114 olfactory-related genes based on the antennal transcriptome database of M. signata, including 21 odorant binding proteins (OBPs), six chemosensory proteins (CSPs), 46 odorant receptors (ORs), 15 ionotropic receptors (IRs), 23 gustatory receptors (GRs) and three sensory neuron membrane proteins (SNMPs). Blastp best hit and phylogenetic analyses showed that most of the chemosensory genes had a close relationship with orthologs from other Coleoptera species. Overall, this study provides a foundation for elucidating the molecular mechanism of olfactory recognition in M. signata as well as a reference for the study of chemosensory genes in other species of Coleoptera., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 He et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Atherigona orientalis Schiner (1868) is an acknowledged agricultural pest owing to its feeding habits and breeding locations. This insect is a tropical and subtropical pest in fruits and vegetables, in which >50 varieties of fruits and vegetables in 26 families, such as Capsicum annuum, Lycopersicon esculentum, and Cucumis melo have been attacked. Moreover, A. orientalis may also develop in rotten crops and feces or insect carcasses, which are also considered one kind of sanitary pest and medical insect. At present, the invasion ranges of A. orientalis are still increasing and more preventive and management measures are to be processed. To gain a better understanding of the molecular mechanisms involved in olfactory reception in A. orientalis, the transcriptome of male and female antennae and legs was systematically analyzed. In total, 131 chemosensory-related genes, including 63 odorant receptors (ORs), 20 gustatory receptors (GRs), 18 ionotropic receptors (IRs), 27 odorant binding proteins (OBPs), 1 chemosensory protein (CSP), and 2 sensory neuron membrane proteins (SNMPs), were identified. The analysis focused on obtaining expression information of candidate olfactory genes at the transcriptomic level by examining the differentially expressed genes (DEGs) in all samples. Totally, 41 DEGs were identified between male antennae (MA) and female antennae (FA), including 32 ORs, 5 OBPs, 1 IR, 2 GRs and 1 SNMP. In MA versus male legs (ML), 78 DEGs were identified (45 ORs, 18 OBPs, 6 GRs, 6 IRs, 1 CSP and 2 SNMPs). In FA and female legs (FL), 96 DEGs were identified (51 ORs, 21 OBPs, 9 GRs, 12 IRs, 1 CSP and 2 SNMPs). For ML and FL, 3 DEGs were identified, including 2 ORs and 1 SNMP. Our results supplement valuable insights for future research on the chemoreception mechanisms in A. orientalis., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024. Published by Elsevier Inc.)

The Japanese pine sawyer Monochamus alternatus serves as the primary vector for pine wilt disease, a devastating pine disease that poses a significant threat to the sustainable development of forestry in the Eurasian region. Currently, trap devices based on informational compounds have played a crucial role in monitoring and controlling the M. alternatus population. However, the specific proteins within M. alternatus involved in recognizing the aforementioned informational compounds remain largely unclear. To elucidate the spatiotemporal distribution of M. alternatus chemosensory-related genes, this study conducted neural transcriptome analyses to investigate gene expression patterns in different body parts during the feeding and mating stages of both male and female beetles. The results revealed that 15 genes in the gustatory receptor (GR) gene family exhibited high expression in the mouthparts, most genes in the odorant binding protein (OBP) gene family exhibited high expression across all body parts, 22 genes in the odorant receptor (OR) gene family exhibited high expression in the antennae, a significant number of genes in the chemosensory protein (CSP) and sensory neuron membrane protein (SNMP) gene families exhibited high expression in both the mouthparts and antennae, and 30 genes in the ionotropic receptors (IR) gene family were expressed in the antennae. Through co-expression analyses, it was observed that 34 genes in the IR gene family were co-expressed across the four developmental stages. The Antenna IR subfamily and IR8a/Ir25a subfamily exhibited relatively high expression levels in the antennae, while the Kainate subfamily, NMDA subfamily, and Divergent subfamily exhibited predominantly high expression in the facial region. MalIR33 is expressed only during the feeding stage of M. alternatus , the MalIR37 gene exhibits specific expression in male beetles, the MalIR34 gene exhibits specific expression during the feeding stage in male beetles, the MalIR8 and MalIR39 genes exhibit specific expression during the feeding stage in female beetles, and MalIR8 is expressed only during two developmental stages in male beetles and during the mating stage in female beetles. The IR gene family exhibits gene-specific expression in different spatiotemporal contexts, laying the foundation for the subsequent selection of functional genes and facilitating the full utilization of host plant volatiles and insect sex pheromones, thereby enabling the development of more efficient attractants.

Olfaction, one of the most important sensory systems governing insect behavior, is a possible target for pest management. Therefore, in this study, we analyzed the antennal transcriptome of the cowpea beetle, Callosobruchus maculatus (F.) (Coleoptera: Chrysomelidae: Bruchinae), which is a major pest of stored pulses and legumes. The de novo antennal RNA-seq assembly results identified 17 odorant, 2 gustatory, and 10 ionotropic receptors, 1 sensory neuron membrane protein, and 12 odorant-binding and 7 chemosensory proteins. Moreover, differential gene expression analysis of virgin male and female antennal samples followed by qRT-PCR revealed 1 upregulated and 4 downregulated odorant receptors in males. We also performed homology searches using the coding sequences built from previously proposed amino acid sequences derived from genomic data and identified additional chemosensory-related genes., Competing Interests: The authors have declared that no competing interests exist.

Crematogaster rogenhoferi (Hymenoptera: Formicidae), an omnivorous ant, is one of the dominant predatory natural enemies of a soft scale pest, Parasaissetia nigra Nietner (Homoptera: Coccidae), and can effectively control P. nigra populations in rubber forests. Olfaction plays a vital role in the process of predation. However, the information about the molecular mechanism of olfaction-evoked behaviour in C. rogenhoferi is limited. In this study, we conducted antennal transcriptome analysis to identify candidate olfactory genes. We obtained 53,892 unigenes, 16,185 of which were annotated. Based on annotations, we identified 49 unigenes related to chemoreception, including four odourant-binding proteins, three chemosensory proteins, 37 odourant receptors, two odourant ionotropic receptors and three sensory neuron membrane proteins. This is the first report on the molecular basis of the chemosensory system of C. rogenhoferi. The findings provide a basis for elucidating the molecular mechanisms of the olfactory-related behaviours of C. rogenhoferi, which would facilitate a better application of C. rogenhoferi as a biological control agent.

The olfactory system of insects facilitates their search for host and mates, hence it plays an essential role for insect survival and reproduction. Insects recognize odor substances through olfactory neurons and olfactory genes. Previous studies showed that there are significant sex-specific differences in how insects identify odorant substances, especially sex pheromones. However, whether the sex-specific recognition of odorant substances is caused by differences in the expression of olfaction-related genes between males and females remains unclear. To clarify this problem, the whole transcriptome sequence of the adult Helicoverpa assulta, an important agricultural pest of tobacco and other Solanaceae plants, was obtained using Pacbio sequencing. RNA-seq analysis showed that there were 27 odorant binding proteins (OBPs), 24 chemosensory proteins, 4 pheromone-binding proteins (PBPs), 68 odorant receptors and 2 sensory neuron membrane proteins (SNMPs) genes, that were expressed in the antennae of male and female H. assulta. Females had significantly higher expression of General odorant-binding protein 1-like, OBP, OBP3, PBP3 and SNMP1 than males, while males had significantly higher expression of GOBP1, OBP7, OBP13, PBP2 and SNMP2. These results improve our understanding of mate search and host differentiation in H. assulta., (© 2021 Wiley Periodicals LLC.)

Sensory genes play a key role in the host location of parasitoids. To date, the sensory genes that regulate parasitoids to locate gall-inducing insects have not been uncovered. An obligate ectoparasitoid, Quadrastichus mendeli Kim & La Salle (Hymenoptera: Eulophidae: Tetrastichinae), is one of the most important parasitoids of Leptocybe invasa, which is a global gall-making pest in eucalyptus plantations. Interestingly, Q. mendeli can precisely locate the larva of L. invasa, which induces tumor-like growth on the eucalyptus leaves and stems. Therefore, Q. mendeli-L. invasa provides an ideal system to study the way that parasitoids use sensory genes in gall-making pests. In this study, we present the transcriptome of Q. mendeli using high-throughput sequencing. In total, 31,820 transcripts were obtained and assembled into 26,925 unigenes in Q. mendeli. Then, the major sensory genes were identified, and phylogenetic analyses were performed with these genes from Q. mendeli and other model insect species. Three chemosensory proteins (CSPs), 10 gustatory receptors (GRs), 21 ionotropic receptors (IRs), 58 odorant binding proteins (OBPs), 30 odorant receptors (ORs) and 2 sensory neuron membrane proteins (SNMPs) were identified in Q. mendeli by bioinformatics analysis. Our report is the first to obtain abundant biological information on the transcriptome of Q. mendeli that provided valuable information regarding the molecular basis of Q. mendeli perception, and it may help to understand the host location of parasitoids of gall-making pests.

Chemoreception is critical for insect behaviors such as foraging, host searching and oviposition. The process of chemoreception is mediated by a series of proteins, including odorant-binding proteins (OBPs), gustatory receptors (GRs), odorant receptors (ORs), ionotropic receptors (IRs), chemosensory proteins (CSPs) and sensory neuron membrane proteins (SNMPs). The tephritid stem gall fly, Procecidochares utilis Stone, is a type of egg parasitic insect, which is an effective biological control agent for the invasive weed Ageratina adenophora in many countries. However, the study of molecular components related to the olfactory system of P. utilis has not been investigated. Here, we conducted the developmental transcriptome (egg, first-third instar larva, pupa, female and male adult) of P. utilis using next-generation sequencing technology and identified a total of 133 chemosensory genes, including 40 OBPs, 29 GRs, 24 ORs, 28 IRs, 6 CSPs, and 6 SNMPs. The sequences of these candidate chemosensory genes were confirmed by BLAST, and phylogenetic analysis was performed. Quantitative real-time PCR (qRT-PCR) confirmed that the expression levels of the candidate OBPs varied at the different developmental stages of P. utilis with most OBPs expressed mainly in the pupae, female and male adults but scarcely in eggs and larvae, which was consistent with the differentially expressed genes (DEGs) analysis using the fragments per kilobase per million fragments (FPKM) value. Our results provide a significant contribution towards the knowledge of the set of chemosensory proteins and help advance the use of P. utilis as biological control agents., (Copyright © 2020. Published by Elsevier Inc.)

The round-headed pine beetle Dendroctonus adjunctus, whose dispersion and colonization behaviors are linked to a communication system mediated by semiochemicals, is one of the five most critical primary pests in forest ecosystems in Mexico. This study provides the first head transcriptome analysis of D. adjunctus and the identification of the nonreceptor olfactory genes involved in the perception of odors. De novo assembly yielded 44,420 unigenes, and GO annotations were similar to those of antennal transcriptomes of other beetle species, which reflect metabolic processes related to smell and signal transduction. A total of 36 new transcripts of nonreceptor olfactory genes were identified, of which 27 encode OBPs, 7 encode CSPs, and 2 encode SNMP candidates, which were subsequently compared to homologous proteins from other bark beetles and Coleoptera species by searching for sequence motifs and performing phylogenetic analyses. Our study provides information on genes encoding nonreceptor proteins in D. adjunctus and broadens the knowledge of olfactory genes in Coleoptera and bark beetle species, and will help to understand colonization and aggregation behaviors for the development of tools that complement management strategies.

Understanding the interaction between the insect olfactory system and the environment is crucial for fully explaining the molecular mechanisms underlying insect behavior, and providing new strategies for integrated pest management. Although there is good evidence that olfactory proteins play a vital role in mediating insect behaviors, the olfactory mechanism of insects remains poorly understood. We identified a total of 71 chemosensory genes; 25 odorant-binding proteins (OBPs), 27 odorant receptors (ORs), 8 ionotropic receptors (IRs), 8 chemosensory proteins (CSPs) and 3 sensory neuron membrane proteins (SNMPs), in the antennae of male and female fall armyworms, Spodoptera frugiperda, an invasive global pest that causes significant economic damage worldwide. We used differential gene expression (DGE) and fragments per kilobase per million fragments (FPKM) values to compare the transcript levels of candidate chemosensory genes, and qRT-PCR to compare the expression levels of the OR gene, in male and female antennae. The expression of candidate OR genes in male and female antennae was consistent with the DGE data, and the expression of the SfruCL4419.Contig1-All and SfruUnigene1070-All genes was sex-biased. These results not only provide new information on the olfactory mechanism of S. frugiperda, and insects in general, but also suggest new gene targets for pest control., Competing Interests: Declaration of competing interest The authors have declared that no competing interest exists., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Cotesia vestalis (Hymenoptera: Braconidae) is an endoparasitoid wasp that attacks larvae of the diamondback moth, Plutella xylostella (Linnaeus), one of the most important pests of cruciferous plants in the world. This natural enemy has been used to control populations of P. xylostella. To more fully understand the molecular mechanisms of olfactory reception in C. vestalis, the transcriptomes of male antennae, female antennae, and mixed female and male legs were systematically analyzed, and the chemosensory genes were identified. A total of 253 olfactory genes, including candidates of 158 odorant receptors (ORs), 12 ionotropic receptors (IRs), 48 gustatory receptors (GRs), 22 odorant binding proteins (OBPs), 11 chemosensory proteins (CSPs), and two sensory neuron membrane proteins (SNMPs), were systematically identified and subjected to sequence and phylogenetic analysis. Then, the differentially expressed genes (DEGs) within female and male antennae were analyzed to obtain expression information on the candidate olfactory genes at the transcriptional level. The expression levels of 25 ORs, nine OBPs, eight GRs, and three IRs were significantly different between male and female antennae. Our results provide valuable information for further studies on the chemoreception mechanisms in C. vestalis, to ultimately improve the pest control measures by using natural enemies., Competing Interests: Declaration of competing interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Antennae are often considered to be the nostrils of insects. Here, we sequenced the transcriptome of the pheromone gland-ovipositor complex of Helicoverpa assulta and discovered that an odorant receptor (OR) gene, HassOR31 , had much higher expression in the ovipositor than in antennae or other tissues. To determine whether the ovipositor was involved in odorant detection, we co-expressed HassOR31 and its co-receptor, HassORco , in a Xenopus oocyte model system, and demonstrated that the OR was responsive to 12 plant odorants, especially Z-3-hexenyl butyrate. These odorants elicited electrophysiological responses of some sensilla in the ovipositor, and HassOR31 and HassORco were co-expressed within ovipositor sensilla. Two oviposition preference experiments showed that female moths lacking antennae still preferentially selected oviposition sites containing plant volatiles. We suggest that the expression of HassOR31 in the ovipositor of H. assulta helps females to determine precise egg-laying sites in host plants., Competing Interests: RL, LH, JD, CW No competing interests declared, (© 2020, Li et al.)

Animals often exhibit dramatically behavioral plasticity depending on their internal physiological state, yet little is known about the underlying molecular mechanisms. The migratory locust, Locusta migratoria, provides an excellent model for addressing these questions because of their famous phase polyphenism involving remarkably behavioral plasticity between gregarious and solitarious phases. Here, we report that a major insect hormone, juvenile hormone, is involved in the regulation of this behavioral plasticity related to phase change by influencing the expression levels of olfactory-related genes in the migratory locust. We found that the treatment of juvenile hormone analog, methoprene, can significantly shift the olfactory responses of gregarious nymphs from attraction to repulsion to the volatiles released by gregarious nymphs. In contrast, the repulsion behavior of solitarious nymphs significantly decreased when they were treated with precocene or injected with double-stranded RNA of JHAMT, a juvenile hormone acid O-methyltransferase. Further, JH receptor Met or JH-response gene Kr-h1 knockdown phenocopied the JH-deprivation effects on olfactory behavior. RNA-seq analysis identified 122 differentially expressed genes in antennae after methoprene application on gregarious nymphs. Interestingly, several olfactory-related genes were especially enriched, including takeout (TO) and chemosensory protein (CSP) which have key roles in behavioral phase change of locusts. Furthermore, methoprene application and Met or Kr-h1 knockdown resulted in simultaneous changes of both TO1 and CSP3 expression to reverse pattern, which mediated the transition between repulsion and attraction responses to gregarious volatiles. Our results suggest the regulatory roles of a pleiotropic hormone in locust behavioral plasticity through modulating gene expression in the peripheral olfactory system., Competing Interests: NO authors have competing interests.

Histia rhodope Cramer (Lepidoptera: Zygaenidae) is one of the most destructive defoliators of landscape tree Bischofia polycarpa (Levl.) Airy Shaw in China stretching to other Southeast Asia regions. Olfactory genes, encoding proteins such as odorant carrier proteins believed to initiate olfactory signal transduction in insects, have been acknowledged to be novel targets for pest control. In this study, we established antennal transcriptome of H. rhodope and ultimately identified 19 odorant binding proteins (OBPs), 23 chemosensory proteins (CSPs) and 4 Niemann-Pick type C2 proteins (NPC2s). The 19 OBPs, 6 CSPs and 4 NPC2s were assessed to validate the differential expressions between sexes, and between olfactory and non-olfactory tissues. 8 OBPs and 2 CSPs exhibited male-biased antennae expression, while 6 OBPs, 2 CSPs and HrhoNPC2a exhibited female-biased antennae expression. Moreover, 17 OBPs, 4 CSPs and 2 NPC2s were predominantly expressed in the antennae compared with non-olfactory tissues. HrhoOBP1 and HrhoOBP8 were predominantly expressed in the antennae and heads, HrhoCSP8 and HrhoCSP14 were highly expressed in abdomens and legs, HrhoNPC2c was highly expressed in abdomens, while HrhoNPC2d was expressed in all tissues. Phylogenetic analysis revealed that most H. rhodope proteins were closely related to proteins from other moths. Moreover, compared with other nocturnal moths, acting as a diurnal moth, we found that H. rhodope may have lost a PBP gene. Our results provide important molecular information for further studies on olfactory mechanisms of H. rhodope., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The rice bug, Leptocorisa acuta (Tunberg) (Hemiptera: Alydidae), is a notorious pest in Asia, and it is significantly attracted by the volatiles derived from host plants. However, it remains unknown how L. acuta recognizes host volatile compounds at the molecular level. Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) are thought to be responsible for the initial biochemical recognition during olfactory perception. Here, we followed the RNA Sequencing (RNA-Seq) approach to identify candidate genes encoding OBPs and CSPs from dissected antennae and mouthparts of L. acuta. In total, 26 unigenes were identified coding for OBPs (22 Classic OBPs and four Plus-C OBPs), and 17 unigenes coding for CSPs. Real-time quantitative PCR (RT-qPCR) revealed that 11 OBPs (LacuOBP1, 5, 6, 7, 9, 11, 13, 14, 17, 20 and 23) and nine CSPs (LacuCSP2, 3, 4, 5, 6, 8, 9, 10 and 12) were predominantly expressed in antennae, indicating that they might be essential for detection of general odorants and pheromones. Among these antennae-predominantly expressed genes, LacuOBP11 and LacuOBP13 showed male-biased expression and therefore may play crucial roles in the detection of sex pheromones. Seven LacuOBPs (LacuOBP4, 8, 10, 12, 21, 25 and 26) and two CSPs (LacuCSP7 and LacuOBP11) were predominantly expressed in mouthparts, suggesting that these genes might be involved in taste perception. Our work provides a starting point to facilitate functional study of these OBPs and CSPs in L. acuta at the molecular level in the future., Competing Interests: Declaration of competing interest The authors declared that they have no conflicts of interest to this work. We declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Olfaction plays key roles in insect survival and reproduction, such as feeding, courtship, mating, and oviposition. The olfactory-based control strategies have been developed an important means for pest management. Streltzoviella insularis is a destructive insect pest of many street tree species, and characterization of its olfactory proteins could provide targets for the disruption of their odour recognition processes and for urban forestry protection. In this study, we assembled the antennal transcriptome of S. insularis by next-generation sequencing and annotated the main olfactory multi-gene families, including 28 odorant-binding proteins (OBPs), 12 chemosensory proteins (CSPs), 56 odorant receptors (ORs), 11 ionotropic receptors (IRs), two sensory neuron membrane proteins (SNMPs), and 101 odorant-degrading enzymes (ODEs). Sequence and phylogenetic analyses confirmed the characteristics of these proteins. We further detected tissue- and sex-specific expression patterns of OBPs, CSPs and SNMPs by quantitative real time-PCR. Most OBPs were highly and differentially expressed in the antennae of both sexes. SinsCSP10 was expressed more highly in male antennae than in other tissues. Two SNMPs were highly expressed in the antennae, with no significant difference in expression between the sexes. Our results lay a solid foundation for understanding the precise molecular mechanisms underlying S. insularis odour recognition.

With the advent of next-generation sequencing, it is now possible to rapidly identify the entire repertoire of olfactory genes likely to be involved in chemical communication of an insect species. It remains, however, a challenge to identify olfactory proteins, such as odorant receptors and odorant-binding proteins (OBPs), vis-à-vis the odorants they detect. It has been reported that exposing the olfactory system to a physiologically relevant odorant alters the transcript levels of odorant receptor(s) involved in the detection of the tested odorant. We applied this paradigm in an attempt to identify putative OBPs from the scarab beetle Holotrichia oblita involved in the reception of plant-derived kairomones. Twenty-nine OBP genes were identified in the H. oblita transcriptome, 20 of which were enriched in antennae compared with nonolfactory tissues. Of these, 2 OBP genes, HoblOBP13 and HoblOBP9, were upregulated upon exposure to one of the female attractants (E)-2-hexenol and phenethyl alcohol; none of the OBP transcripts changed upon exposure to methyl anthranilate, which does not attract H. oblita females. Binding assays showed that HoblOBP13 and HoblOBP9 have high affinity for (E)-2-hexenol and phenethyl alcohol, respectively. RNAi treatment showed that transcripts of both HoblOBP13 and HoblOBP9 declined in a time-course manner 24-72 h postinjection. OBP-dsRNA-treated female beetles showed significantly lower attraction to (E)-2-hexenol and phenethyl alcohol than did water-injected beetles and those treated with GFP-dsRNA. We, therefore, concluded that HoblOBP13 and HoblOBP9 are essential for H. oblita reception of the plant-derived kairomones (E)-2-hexenol and phenethyl alcohol., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Olfaction and gustation play critical roles in the daily activities of insects by mediating important behaviors, such as locating host plants, mates and oviposition sites. These behaviors are mediated by a series of chemosensory receptor proteins, including odorant receptors (ORs), gustatory receptors (GRs) and ionotropic receptors (IRs). Holotrichia parallela is a worldwide pest on many crops and particularly on peanuts in China. In the present study, we produced Illumina-based transcriptomes from male and female antennae and ultimately identified 47 candidate ORs, 15 GRs and 27 IRs. Based on phylogenetic analyses, the relatedness with other species and possible functions of chemosensory genes were predicted and analyzed, such as the putative OR co-receptor (HparOR40), IR co-receptors (HparIR1 and HparIR27), carbon dioxide receptors (HparGR5 and HparGR14) and sugar receptors (HparGR8, HparGR11 and HparGR13). The differences in expression between genes and between sexes were analyzed using FPKM values and visualized in heat maps. Furthermore, the qRT-PCR results of 26 candidate genes were assessed to validate the differential expression analyses between sexes and between olfactory and non-olfactory tissues. Thirteen genes were more highly expressed in female than in male antennae, whereas four genes had higher expression in male than in female antennae. Twelve genes were predominantly expressed in the antennae as olfactory organs compared with expression in non-olfactory tissues. Our study was the first to annotate the chemosensory receptors of H. parallela, and it will provide a strong foundation for future work on physiology and ecology at the molecular level., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Olfaction in antennae is essential for regulating insect behaviors such as host preference and oviposition site selection. To better understand the olfactory mechanisms in the cicada Subpsaltria yangi that has a very narrow host range far fewer than diets of most other cicadas, an antennal transcriptome was constructed in this study. We identified 10 unigenes encoding putative odorant-binding proteins (OBPs), 10 chemosensory proteins (CSPs), 8 sensory neuron membrane proteins (SNMPs), 7 gustatory receptors (GRs), 29 odorant receptors (ORs) and 13 ionotropic receptors (IRs). We found that the OBPs were separated into two groups, Classic OBPs and Plus-C OBPs, according to their motifs. Based on sequence alignment and phylogenetic analysis, we found five Orcos in OR family. However, no pheromone receptor was identified, which may be related to that cicadas are sound-producing insects and acoustic signals other than pheromones play most important role in sexual communication. We used qRT-PCR to examine the expression profile of ten OBPs genes in various organs, and found that they were mainly expressed in olfactory organs such as antennae and mouthparts. Our results make it possible for future research of the olfactory system of S. yangi at the molecular level, and provide important bases for elucidation of the molecular mechanisms and evolution of chemosensation in sap-sucking insects., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Chemical senses are crucial for insect behaviors such as host preference, mate choice and oviposition site selection. Various protein families are involved in these processes, including odorant receptors (ORs), ionotropic receptors (IRs), gustatory receptors (GRs), chemosensory proteins (CSPs), odorant binding proteins (OBPs) and sensory neuron membrane proteins (SNMPs). To better understand the olfactory mechanism in Spodoptera exigua, we conducted transcriptome analysis of adult antennae and identified a total of 157 candidate chemosensory genes encoding 51 ORs, 20 IRs, 7 GRs, 32 CSPs, 45 OBPs and 2 SNMPs. Quantitative real time PCR (qPCR) analysis of the tissue- and sex-specific expression profiles of ORs, GRs and IRs revealed that these genes could be detected in at least one tissue tested. SexiOR6, 11, 13 and 16, which were predicted to be pheromone receptors based on phylogenetic analysis, exhibited male-specific antennae expression. SexiOR18, 26, 28, 30, 34, 39, and 40 exhibited female-biased expression. SexiGR1, SexiGR2 and SexiGR3, are predicted carbon dioxide receptors, and the former was expressed specifically in antennae, and the latter two were expressed both in antennae and labial palps. SexiIRs had diverse expression profiles. SexiIR8a and SexiIR25a were quite conserved and expressed at high levels. This work will greatly facilitate the understanding of olfactory system in S. exigua and provides valuable information for further functional studies of the chemoreception mechanism in Lepidopteran moths., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The grapevine moth, Lobesia botrana, is considered a harmful pest for vineyards in Chile as well as in North America and Europe. Currently, monitoring and control methods of L. botrana are based on its main sex pheromone component, being effective for low population densities. In order to improve control methods, antennal olfactory proteins in moths, such as odorant-binding proteins (OBPs) and odorant receptors (ORs) have been studied as promising targets for the discovery of new potent semiochemicals, which have not been reported for L. botrana. Therefore, the objective of this study was to identify the repertoire of proteins related to chemoreception in L. botrana by antennal transcriptome and analyze the relative expression of OBPs and CSPs in male and female antennae. Through next-generation sequencing of the antennal transcriptome by Ilumina HiSeq2500 we identified a total of 118 chemoreceptors, from which 61, 42 and 15 transcripts are related to ORs, ionotropic receptors (IRs) and gustatory receptors (GRs), respectively. Furthermore, RNA-Seq data revealed 35 transcripts for OBPs and 18 for chemosensory proteins (CSPs). Analysis by qRT-PCR showed 20 OBPs significantly expressed in female antennae, while 5 were more expressed in males. Similarly, most of the CSPs were significantly expressed in female than male antennae. All the olfactory-related sequences were compared with homologs and their phylogenetic relationships elucidated. Finally, our findings in relation to the improvement of L. botrana management are discussed., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Cyrtorhinus lividipennis Reuter (Hemiptera: Miridae) is an important egg predator of planthoppers which are destructive rice pests. The chemosensory genes in the mirid antennae play important roles in mating and prey-seeking behaviors. To gain a better understanding of the olfaction of C. lividipennis, we sequenced the antennal transcriptomes of the predator to identify the key olfaction genes. We identified 18 odorant binding proteins (OBPs), 12 chemosensory proteins (CSPs), 1 Niemann-Pick C2 protein (NPC2), 15 odorant receptors (ORs), 6 ionotropic receptors (IRs), 3 gustatory receptors (GRs) and 3 sensory neuron membrane proteins (SNMPs). Quantitative real-time PCR results showed that the relative transcript levels of three ClivORs (ClivOR6, 7 and 14) in the female antennae were 3 to 6 folds higher than that in the male antennae, indicating these genes were more related to oviposition site selection. The relative transcript levels of ClivCSP8 and ClivOR11 were 2.6 and 2.7 times higher in the male antennae than that of the female, respectively, indicating that these genes might be involved in mate searching. Moreover, the responses of dsorco treated predators to volatiles emitted from infested rice were significantly reduced, indicating these volatiles might serve as crucial cues in the host searching of C. lividipennis.

The coffee white stemborer, Xylotrechus quadripes Chevrolat (Coleoptera: Cerambycidae), feeds primarily on Coffea arabica L. (Gentianales: Rubiaceae) with its egg, larva, and pupa being developed within the trunk. The detection of chemosensory-related cues linked to adult mating, host seeking, and recognition is driven by three chemoreceptor gene repertoires of odorant (ORs), gustatory (GRs), and ionotropic (IRs) receptors as well as sensory neuron membrane proteins (SNMPs). Yet, information on these genes involved in chemoreception is unavailable in X. quadripes and relatively poor in the cerambycid beetles. Here, we presented the identification of four chemosensory transmembrane proteins from the antennal transcriptome of X. quadripes, including 33 ORs, five GRs, 18 IRs, and four SNMPs. Phylogenetic analysis classified the ORs into groups 1, 2, 3, 7, and olfactory coreceptor (Orco), showing three potential candidates (OR13, OR17, and OR21) for the sensing of male sex pheromones. The IRs were clustered into 10 orthologous groups, with additional copies for IR41a, IR64a, and IR75 clades. Four SNMPs were distributed in four independent clades, possibly representing a complete set in this species. Expression profiles revealed that all the genes were highly expressed in antennae, suggesting their olfactory roles. In addition, most of the genes showed the expression in nonantennal tissues including thoraxes, abdomens, wings, and legs, suggesting their involvement in nonchemosensory functions. Of notice, a highly conserved coreceptor IR25a displayed male-biased expression in the antennae, as the first presence in the cerambycid beetles. This study has established reference resources for understanding the mechanisms underlying the interactions between/within this beetle and its host plants.

The apple buprestid beetle, Agrilus mali Matsumura (Coleoptera: Buprestidae), can respond to various volatiles, but the underlying mechanism of odorant perception for this insect is poorly understood. Here, we cloned A. mali's odorant-binding proteins 3 (AmalOBP3) and 8 (AmalOBP8) and characterized their expression patterns and binding profiles. Sequence and phylogenetic analyses showed that AmalOBP3 and AmalOBP8 were distributed in the classic and minus-C OBP subfamily, respectively. AmalOBP3 was specifically and abundantly expressed in antennae of both sexes. AmalOBP8 displayed high transcript levels in antennae of both sexes, abdomens of males, and wings of both sexes. Both AmalOBPs exhibited much higher expression in male antennae than in female antennae, suggesting that they could be important in perception of male-specific olfactory cues (e.g., some sex pheromones). Out of the 40 odorant ligands tested, AmalOBP3 and AmalOBP8 bound to 15 and 21 different odorants, respectively, indicating a distinct and selective binding profile for them. Both AmalOBPs seemed to have very strong binding affinity to aliphatic alcohols and aldehydes with 12 to 15 carbon atoms. Alcohols, esters, and terpenoids were more likely to be good ligands for both AmalOBPs than aldehydes and alkanes. Together with its broad expression in different tissues, strong binding with higher numbers of putative ligands for AmalOPB8 means that this protein can have more extensive functional roles in chemosensation of A. mali. Our results provide insights into the molecular basis of chemosensation in A. mali, as well as a basis for developing detection, monitoring, and management tools for this serious pest.

Olfaction plays an indispensable role in mediating insect behavior, such as locating host plants, mating partners, and avoidance of toxins and predators. Olfactory-related proteins are required for olfactory perception of insects. However, very few olfactory-related genes have been reported in Plodia interpunctella up to now. In the present study, we sequenced the antennae transcriptome of P. interpunctella using the next-generation sequencing technology, and identified 117 candidate olfactory-related genes, including 29 odorant-binding proteins (OBPs), 15 chemosensory proteins (CSPs), three sensory neuron membrane proteins (SNMPs), 47 odorant receptors (ORs), 14 ionotropic receptors (IRs) and nine gustatory receptors (GRs). Further analysis of qRT-PCR revealed that nine OBPs, three CSPs, two SNMPs, nine ORs and two GRs were specifically expressed in the male antennae, whereas eight OBPs, six CSPs, one SNMP, 16 ORs, two GRs and seven IRs significantly expressed in the female antennae. Taken together, our results provided useful information for further functional studies on insect genes related to recognition of pheromone and odorant, which might be meaningful targets for pest management.

Adelphocoris suturalis Jakovlev (Hemiptera: Miridae) is an insect pest that causes severe agricultural damage to cotton and many other important crops. In insects, olfaction is very important throughout their lifetime. There are two groups of small soluble proteins, named odorant binding proteins (OBPs) and chemosensory proteins (CSPs), which are suggested to participate in the initial biochemical recognition steps of insect olfactory signal transduction. In this study, a total of 16 OBPs (12 classical OBPs and 4 plus-C OBPs) and 8 CSPs, were identified in the antennal transcriptome of A. suturalis. The sex- and tissue-specific profiles of these binding protein genes showed that 13 of the 16 OBP transcripts were highly expressed in the antennae of both sexes, and 4 OBPs (AsutOBP1, 4, 5 and 9) were expressed higher in the male antennae compared to the female antennae. Three CSPs (AsutCSP1, 4 and 5) were expressed specifically in the antennae of both sexes, and AsutCSP1 was expressed higher in the male antennae than in the female antennae. Our findings identify several novel OBP and CSP genes for further investigation of the olfactory system of A. suturalis at the molecular level., (Copyright © 2016. Published by Elsevier Inc.)

Olfaction in insects is essential for host identification, mating and oviposition, in which olfactory proteins are responsible for chemical signaling. Here, we determined the transcriptomes of male and female adult antennae of Anoplophora chinensis, the citrus longhorned beetle. Among 59,357 unigenes in the antennal assembly, we identified 46 odorant-binding proteins, 16 chemosensory proteins (CSPs), 44 odorant receptors, 19 gustatory receptors, 23 ionotropic receptors, and 3 sensory neuron membrane proteins. Among CSPs, AchiCSP10 was predominantly expressed in antennae (compared with legs or maxillary palps), at a significantly higher level in males than in females, suggesting that AchiCSP10 has a role in reception of female sex pheromones. Many highly expressed genes encoding CSPs are orthologue genes of A. chinensis and Anoplophora glabripennis. Notably, AchiPBP1 and AchiPBP2 showed 100% and 96% identity with AglaPBP1 and AglaPBP2 from A. glabripennis, with similar expression profiles in the two species; PBP2 was highly expressed in male antennae, whereas PBP1 was expressed in all three tissues in both males and females. These results provide a basis for further studies on the molecular chemoreception mechanisms of A. chinensis, and suggest novel targets for control of A. chinensis.

Antenna is the main chemosensory organ in mosquitoes. Characterization of the transcriptional changes after blood meal, especially those related to chemoreception, may help to explain mosquito blood sucking behavior and to identify novel targets for mosquito control. Anopheles sinensis is an Asiatic mosquito species which transmits malaria and lymphatic filariasis. However, studies on chemosensory biology in female An. sinensis are quite lacking. Here we report a transcriptome analysis of An. sinensis female antennae pre- and post- blood meal. We created six An. sinensis antenna RNA-seq libraries, three from females without blood meal and three from females five hours after a blood meal. Illumina sequencing was conducted to analyze the transcriptome differences between the two groups. In total, the sequenced fragments created 21,643 genes, 1,828 of them were novel. 12,861 of these genes were considered to be expressed (FPKM >1.0) in at least one of the two groups, with 12,159 genes expressed in both groups. 548 genes were differentially expressed in the blood-fed group, with 331 genes up-regulated and 217 genes down-regulated. GO enrichment analysis of the differentially expressed genes suggested that there were no statistically over represented GO terms among down-regulated genes in blood-fed mosquitoes, while the enriched GO terms of the up-regulated genes occurred mainly in metabolic process. For the chemosensory gene families, a subtle distinction in the expression levels can be observed according to our statistical analysis. However, the firstly comprehensive identification of these chemosensory gene families in An. sinensis antennae will help to characterize the precise function of these proteins in odor recognition in mosquitoes. This study provides a first global view in the changes of transcript accumulation elicited by blood meal in An. sinensis female antennae.

Studies on insect olfaction have increased our understanding of insect's chemosensory system and chemical ecology, and have improved pest control strategies based on insect behavior. In this study, we assembled the antennal transcriptomes of the lychee giant stink bug, Tessaratoma papillosa, by using next generation sequencing to identify the major olfaction gene families in this species. In total, 59 odorant receptors, 14 ionotropic receptors (8 antennal IRs), and 33 odorant binding proteins (28 classic OBPs and 5 plus-C OBPs) were identified from the male and female antennal transcriptomes. Analyses of tissue expression profiles revealed that all 59 OR transcripts, 2 of the 8 antennal IRs, and 6 of the 33 OBPs were primarily expressed in the antennae, suggesting their putative role in olfaction. The sex-biased expression patterns of these antenna-predominant genes suggested that they may have important functions in the reproductive behavior of these insects. This is the first report that provides a comprehensive resource to future studies on olfaction in the lychee giant stink bug.

Antennal and abdominal transcriptomes of males and females of the coconut hispine beetle Brontispa longissima were sequenced to identify and compare the expression patterns of genes involved in odorant reception and detection. Representative proteins from the chemosensory gene families likely essential for insect olfaction were identified. These include 48 odorant receptors (ORs), 19 ionotropic receptors (IRs), 4 sensory neuron membrane proteins (SNMPs), 34 odorant binding proteins (OBPs) and 16 chemosensory proteins (CSPs). Phylogenetic analysis revealed the evolutionary relationship of these proteins with homologs from Coleopterans or other insects, and led to the identification of putative aggregation pheromone receptors in B. longissima. Comparative expression analysis performed by calculating FPKM values were also validated using quantitative real time-PCR (qPCR). The results revealed that all ORs and antennal IRs, two IR co-receptors (BlonIR8a and BlonIR25a) and one SNMP (BlonSNMP1a) were predominantly expressed in antennae when compared to abdomens, and approximately half of the OBPs (19) and CSPs (7) were enriched in antennae. These findings for the first time reveal the identification of key molecular components in B. longissima olfaction and provide a valuable resource for future functional analyses of olfaction, and identification of potential targets to control this quarantine pest.

To better understand the olfactory mechanisms in the oriental armyworm Mythimna separate, one of the most serious pests of cereals, an antennal transcriptome was constructed in this study. A total of 130 olfactory related transcripts were identified. These transcripts were predicted to encode 32 odorant-binding proteins (OBPs), 16 chemosensory proteins (CSPs), 71 olfactory receptors (ORs), 8 ionotropic receptors (IRs), 1 gustatory receptor (GR) and 2 sensory neuron membrane proteins (SNMPs). Q-PCR analysis of the temporal expression profiles of seven OBPs in different tissues indicated that, except for MsepOBP19 which was highly expressed in the wings of 0-day-old adult and MsepOBP20 which was low expressed in all tissues, other tested MsepOBPs were significantly more highly expressed in the antenna than in the head (antenna excluded), thorax, abdomen, legs and wings. The expression levels of MsepOBPs were diverse in different life stages (differed on eclosion days). MsepOBP5 exhibited female-biased expression in 0- and 5-day-old adult, while no gender bias in 1- and 3-day-old adult was detected and similar expression profiles were found for MsepOBP7, 20, 24 and 26. In addition, we found that although the expression of MsepOBP22 was female biased in 0- and 5-day-old adult, in the 3-day-old adult it was male-biased. Our findings established a foundation for future studies of the functions of olfactory proteins in M. separata., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Meteorus pulchricornis is an endoparasitoid wasp which attacks the larvae of various lepidopteran pests. We present the first antennal transcriptome dataset for M. pulchricornis. A total of 48,845,072 clean reads were obtained and 34,967 unigenes were assembled. Of these, 15,458 unigenes showed a significant similarity (E-value <10 -5 ) to known proteins in the NCBI non-redundant protein database. Gene ontology (GO) and cluster of orthologous groups (COG) analyses were used to classify the functions of M. pulchricornis antennae genes. We identified 16 putative odorant-binding protein (OBP) genes, eight chemosensory protein (CSP) genes, 99 olfactory receptor (OR) genes, 19 ionotropic receptor (IR) genes and one sensory neuron membrane protein (SNMP) gene. BLASTx best hit results and phylogenetic analysis both indicated that these chemosensory genes were most closely related to those found in other hymenopteran species. Real-time quantitative PCR assays showed that 14 MpulOBP genes were antennae-specific. Of these, MpulOBP6, MpulOBP9, MpulOBP10, MpulOBP12, MpulOBP15 and MpulOBP16 were found to have greater expression in the antennae than in other body parts, while MpulOBP2 and MpulOBP3 were expressed predominately in the legs and abdomens, respectively. These results might provide a foundation for future studies of olfactory genes and chemoreception in M. pulchricornis., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Loxostege sticticalis Linnaeus is an economically important agricultural pest, and the larvae cause great damage to crops, especially in Northern China. However, effective and environmentally friendly chemical methods for controlling this pest have not been discovered to date. In the present study, we performed HiSeq2500 sequencing of transcriptomes of the male and female adult antennae, adult legs and third instar larvae, and we identified 54 candidate odorant receptors (ORs), including 1 odorant receptor coreceptor (Orco) and 5 pheromone receptors (PRs), 18 ionotropic receptors (IRs), 13 gustatory receptors (GRs), 34 odorant binding proteins (OBPs), including 1 general odorant binding protein (GOBP1) and 3 pheromone binding proteins (PBPs), 10 chemosensory proteins (CSPs) and 2 sensory neuron membrane proteins (SNMPs). The results of RNA-Seq and RT-qPCR analyses showed the expression levels of most genes in the antennae were higher than that in the legs and larvae. Furthermore, PR4, OR1-4, 7-11, 13-15, 23, 29-32, 34, 41, 43, 47/IR7d.2/GR5b, 45, 7/PBP2-3, GOBP1, OBP3, 8 showed female antennae-biased expression, while PR1/OBP2, 7/IR75d/CSP2 showed male antennae-biased expression. However, IR1, 7d.3, 68a/OBP11, 20-22, 28/CSP9 had larvae enriched expression, and OBP15, 17, 25, 29/CSP5 were mainly expressed in the legs. The results shown above indicated that these genes might play a key role in foraging, seeking mates and host recognition in the L. sticticalis. Our findings will provide the basic knowledge for further studies on the molecular mechanisms of the olfactory system of L. sticticalis and potential novel targets for pest control strategies.

Chemosensory membrane proteins, including odorant receptors (ORs), ionotropic receptors (IRs), gustatory receptors (GRs) and sensory neurone membrane proteins (SNMPs), are supposed to be crucial macromolecules in the insect olfactory signal transduction pathway. The alfalfa plant bug Adelphocoris lineolatus (Goeze) (Hemiptera: Miridae) is highly attracted to high-nitrogen or flowering plants and destroys many important agricultural crops. We assembled the antennal transcriptome of A. lineolatus using Illumina sequencing technology and identified a total of 108 transcripts encoding chemosensory membrane proteins (88 ORs, 12 IRs, four GRs and four SNMPs), amongst which 90 candidates appeared to be full length. Subsequently, both semiquantitative reverse transcription PCR and quantitative real-time PCR experiments were performed to investigate their tissue- and sex-biased expression profiles. The results showed that nearly all of the 108 candidate chemosensory membrane protein genes were largely expressed in adult antennae, and some genes additionally displayed significant differences in the expression levels between sexes. The results of our phylogenetic analysis and the detailed tissue- and sex-biased expression characteristics given here provide an important foundation for further understanding of the complex chemoreception system of the alfalfa plant bug and other Hemiptera species, which also could help us use chemosensory membrane proteins as targets to manipulate insect olfactory behaviour and broaden the applications of available tools for insect pest control., (© 2016 The Royal Entomological Society.)

Background: The oriental armyworm Mythimna separata (Walk) is a serious migratory pest; however, studies on its olfactory response and its underlying molecular mechanism are limited. To gain insights to the olfactory mechanism of migration, olfactory genes were identified using antennal transcriptome analysis. The olfactory response and the expression of olfactory genes for 1-day and 5-day-old moths were respectively investigated by EAG and RT-qPCR analyses., Results: Putative 126 olfactory genes were identified in M. separata, which included 43 ORs, 13 GRs, 16 IRs, 37 OBPs, 14 CSPs, and 3 SNMPs. RPKM values of IR75d and 10 ORs were larger than co-receptors IR25a and ORco, and the RPKM value of PR2 was larger than that of other ORs. Expression of GR1 (sweet receptor) was higher than that of other GRs. Several sex pheromones activated evident EAG responses where the responses of 5-day-old male moths to the sex pheromones were significantly greater than those of female and 1-day old male moths. In accordance with the EAG response, 11 pheromone genes, including 6 PRs and 5 PBPs were identified in M. separate, and the expression levels of 7 pheromone genes in 5-day-old moths were significantly higher than those of females and 1-day-old moths. PR2 and PBP2 might be used in identifying Z11-16: Ald, which is the main sex pheromone component of M. separata. EAG responses to 16 plant volatiles and the expression levels of 43 olfactory genes in 1-day-old moths were significantly greater than that observed in the 5-day-old moths. Heptanal, Z6-nonenal, and benzaldehyde might be very important floral volatiles for host searching and recognized by several olfactory genes with high expression. Some plant volatiles might be important to male moths because the EAG response to 16 plant volatiles and the expression of 43 olfactory genes were significantly larger in males than in females., Conclusions: The findings of the present study show the effect of adult age on olfactory responses and expression profile of olfactory genes in the migratory pest M. separate.

Hyphantria cunea (Drury) (Lepidoptera: Arctiidae) is an invasive insect pest which, in China, causes unprecedented damage and economic losses due to its extreme fecundity and wide host range, including forest and shade trees, and even crops. Compared to the better known lepidopteran species which use Type-I pheromones, little is known at the molecular level about the olfactory mechanisms of host location and mate choice in H. cunea, a species using Type-II lepidopteran pheromones. In the present study, the H. cunea antennal transcriptome was constructed by Illumina Hiseq 2500TM sequencing, with the aim of discovering olfaction-related genes. We obtained 64,020,776 clean reads, and 59,243 unigenes from the analysis of the transcriptome, and the putative gene functions were annotated using gene ontology (GO) annotation. We further identified 124 putative chemosensory unigenes based on homology searches and phylogenetic analysis, including 30 odorant binding proteins (OBPs), 17 chemosensory proteins (CSPs), 52 odorant receptors (ORs), 14 ionotropic receptors (IRs), nine gustatory receptors (GRs) and two sensory neuron membrane proteins (SNMPs). We also found many conserved motif patterns of OBPs and CSPs using a MEME system. Moreover, we systematically analyzed expression patterns of OBPs and CSPs based on reverse transcription PCR and quantitative real time PCR (RT-qPCR) with RNA extracted from different tissues and life stages of both sexes in H. cunea. The antennae-biased expression may provide a deeper further understanding of olfactory processing in H. cunea. The first ever identification of olfactory genes in H. cunea may provide new leads for control of this major pest., Competing Interests: The authors have declared that no competing interests exist.

The brown marmorated stink bug, Halyomorpha halys, is a devastating invasive species in the USA. Similar to other insects, olfaction plays an important role in its survival and reproduction. As odorant-binding proteins (OBPs) are involved in the initial semiochemical recognition steps, we used RNA-Sequencing (RNA-Seq) to identify OBPs in its antennae, and studied their expression pattern in different body parts under semiochemical stimulation by either aggregation or alarm pheromone or food odorants. Thirty full-length putative HhalOBPs were identified, corresponding to 22 'classic' OBPs and eight 'Plus-C' OBPs. The similarity amongst them ranged from 4.95-70.92%, and with another 325 hemipteran OBPs similarity ranged from 1.94-91.51%, the highest levels being with other stink bug OBPs. Phylogenetic analysis confirmed the monophyly of seven groups of stink bug and other hemipteran OBPs. All 30 HhalOBPs were expressed and about 2/3 were expressed primarily in antennae. The expression of 21 HhalOBPs was higher in the antennae under alarm pheromone stimulus, indicating that multiple OBPs may be responding to this pheromone. Two were highest in antennae under aggregation pheromone stimulus. These findings should provide a basis for understanding the physiological functions of HhalOBPs and the chemosensory perception of this pest, which may help to uncover new control targets for behavioural interference., (© 2016 The Royal Entomological Society.)

Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) of aphids are thought to be responsible for the initial molecular interactions during olfaction that mediate detection of chemical signals. Analysis of the diversity of proteins involved comprises critical basic research work that will facilitate the development of sustainable pest control strategies. To help us better understand differences in the olfactory system between winged and wingless grain aphids, we constructed an antennal transcriptome from winged and wingless Sitobion avenae (Fabricius), one of the most serious pests of cereal fields worldwide. Among the 133,331 unigenes in the antennal assembly, 13 OBP and 5 CSP putative transcripts were identified with 6 OBP and 3 CSP sequences representing new S. avenae annotations. We used qPCR to examine the expression profile of these genes sets across S. avenae development and in various tissues. We found 7 SaveOBPs and 1 SaveCSP were specifically or significantly elevated in antennae compared with other tissues, and that some transcripts (SaveOBP8, SaveCSP2 and SaveCSP5) were abundantly expressed in the legs of winged or wingless aphids. The expression levels of the SaveOBPs and SaveCSPs varied depending on the developmental stage. Possible physiological functions of these genes are discussed. Further molecular and functional studies of these olfactory related genes will explore their potential as novel targets for controlling S. avenae., Competing Interests: The authors have declared that no competing interests exist.

The Asian citrus psyllid, Diaphorina citri is the principal vector of the highly destructive citrus disease called Huanglongbing (HLB) or citrus greening, which is a major threat to citrus cultivation worldwide. More effective pest control strategies against this pest entail the identification of potential chemosensory proteins that could be used in the development of attractants or repellents. However, the molecular basis of olfaction in the Asian citrus psyllid is not completely understood. Therefore, we performed this study to analyze the antennal and abdominal transcriptome of the Asian citrus psyllid. We identified a large number of transcripts belonging to nine chemoreception-related gene families and compared their expression in male and female adult antennae and terminal abdomen. In total, 9 odorant binding proteins (OBPs), 12 chemosensory proteins (CSPs), 46 odorant receptors (ORs), 20 gustatory receptors (GRs), 35 ionotropic receptors (IRs), 4 sensory neuron membrane proteins (SNMPs) and 4 different gene families encoding odorant-degrading enzymes (ODEs): 80 cytochrome P450s (CYPs), 12 esterase (ESTs), and 5 aldehyde dehydrogenases (ADE) were annotated in the D. citri antennal and abdominal transcriptomes. Our results revealed that a large proportion of chemosensory genes exhibited no distinct differences in their expression patterns in the antennae and terminal abdominal tissues. Notably, RNA sequencing (RNA-seq) data and quantitative real time-PCR (qPCR) analyses showed that 4 DictOBPs, 4 DictCSPs, 4 DictIRs, 1 DictSNMP, and 2 DictCYPs were upregulated in the antennae relative to that in terminal abdominal tissues. Furthermore, 2 DictOBPs (DictOBP8 and DictOBP9), 2 DictCSPs (DictOBP8 and DictOBP12), 4 DictIRs (DictIR3, DictIR6, DictIR10, and DictIR35), and 1 DictCYP (DictCYP57) were expressed at higher levels in the male antennae than in the female antennae. Our study provides the first insights into the molecular basis of chemoreception in this insect pest. Further studies on the identified differentially expressed genes would facilitate the understanding of insect olfaction and their role in the interactions between olfactory system and biological processes.

Ionotropic receptors (IRs) mainly detect the acids and amines having great importance in many insect species, representing an ancient olfactory receptor family in insects. In the present work, we performed RNAseq of Microplitis mediator antennae and identified seventeen IRs. Full-length MmedIRs were cloned and sequenced. Phylogenetic analysis of the Hymenoptera IRs revealed that ten MmedIR genes encoded "antennal IRs" and seven encoded "divergent IRs". Among the IR25a orthologous groups, two genes, MmedIR25a.1 and MmedIR25a.2, were found in M. mediator. Gene structure analysis of MmedIR25a revealed a tandem duplication of IR25a in M. mediator. The tissue distribution and development specific expression of the MmedIR genes suggested that these genes showed a broad expression profile. Quantitative gene expression analysis showed that most of the genes are highly enriched in adult antennae, indicating the candidate chemosensory function of this family in parasitic wasps. Using immunocytochemistry, we confirmed that one co-receptor, MmedIR8a, was expressed in the olfactory sensory neurons. Our data will supply fundamental information for functional analysis of the IRs in parasitoid wasp chemoreception., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Periplaneta americana is a notorious urban pest prevalent in human habitats; very little is known about its chemosensory mechanism. Employing the advanced next-generation sequencing technique, in the present study, we conducted transcriptome sequencing and analysis of the antennae of the adult males and females as well as their mouthparts using an Illumina platform. This resulted in the discovery of a huge number of the members of all major known chemosensory receptor families in P. americana, including 96 odorant receptors (ORs), 53 ionotropic receptors (IRs), and 33 gustatory receptors (GRs). Tissue expression profiles showed most of them mainly expressed in antennae and phylogenetic analysis demonstrated the expansion in the clade distinguishing them from other functionally well-known Lepidoptera species. A high percentage of chemosensory receptor genes (ORs in particular) showing female antenna bias in mRNA expression was observed. Our results provide a basis for further investigations on how P. americana coordinates its chemosensory receptor genes in chemical communication with environments, and for development of novel pest management approaches.

Olfactory proteins form the basis of insect olfactory recognition, which is crucial for host identification, mating, and oviposition. Using transcriptome analysis of Anoplophora glabripennis antenna, we identified 42 odorant-binding proteins (OBPs), 12 chemosensory proteins (CSPs), 14 pheromone-degrading enzymes (PDEs), 1 odorant-degrading enzymes (ODE), 37 odorant receptors (ORs), 11 gustatory receptors (GRs), 2 sensory neuron membrane proteins (SNMPs), and 4 ionotropic receptor (IR). All CSPs and PBPs were expressed in antennae, confirming the authenticity of the transcriptome data. CSP expression profiles showed that AglaCSP3, AglaCSP6, and AglaCSP12 were expressed preferentially in maxillary palps and AglaCSP7 and AglaCSP9 were strongly expressed in antennae. The vast majority of CSPs were highly expressed in multiple chemosensory tissues, suggesting their participation in olfactory recognition in almost all olfactory tissues. Intriguingly, the PBP AglaPBP2 was preferentially expressed in antenna, indicating that it is the main protein involved in efficient and sensitive pheromone recognition. Phylogenetic analysis of olfactory proteins indicated AglaGR1 may detect CO2. This study establishes a foundation for determining the chemoreception molecular mechanisms of A. glabripennis, which would provide a new perspective for controlling pest populations, especially those of borers.

Olfactory transduction is a process by which olfactory sensory neurons (OSNs) transform odor information into neuronal electrical signals. This process begins with the binding of odor molecules to receptor proteins on olfactory receptor neuron (ORN) dendrites. The major molecular components involved in olfaction include odorant-binding proteins (OBPs), chemosensory proteins (CSPs), odorant receptors (ORs), gustatory receptors (GRs), ionotropic receptors (IRs), sensory neuron membrane proteins (SNMPs) and odorant-degrading enzymes (ODEs). More importantly, as potential molecular targets, chemosensory proteins are used to identify novel attractants or repellants for environmental-friendly pest management. In this study we analyzed the transcriptome of the flea beetle, Phyllotreta striolata (Coleoptera, Chrysomelidae), a serious pest of Brassicaceae crops, to better understand the molecular mechanisms of olfactory recognition in this pest. The analysis of transcriptomes from the antennae and terminal abdomens of specimens of both sexes identified transcripts from several key molecular components of chemoreception including 73 ORs, 36 GRs, 49 IRs, 2 SNMPs, 32 OBPs, 8 CSPs, and four candidate odorant degrading enzymes (ODEs): 143 cytochrome P450s (CYPs), 68 esterases (ESTs), 27 glutathione S-transferases (GSTs) and 8 UDP-glycosyltransferases (UGTs). Bioinformatic analyses indicated that a large number of chemosensory genes were up-regulated in the antennae. This was consistent with a potential role in olfaction. To validate the differential abundance analyses, the expression of 19 genes encoding various ORs, CSPs, and OBPs was assessed via qRT-PCR between non-chemosensory tissue and antennae. Consistent with the bioinformatic analyses, transcripts for all of the genes in the qRT-PCR subset were elevated in antennae. These findings provide the first insights into the molecular basis of chemoreception in the striped flea beetle.

Chouioia cunea Yang is an endoparasitic wasp that attacks pupae of Hyphantria cunea (Drury), an invasive moth species that severely damages forests in China. Chemosensory systems of insects are used to detect volatile chemical odors such as female sex pheromones and host plant volatiles. The antennae of parasite wasps are important for host detection and other sensory-mediated behaviors. We identified and documented differential expression profiles of chemoreception genes in C. cunea antennae. A total of 25 OBPs, 80 ORs, 10 IRs, 11 CSP, 1 SNMPs, and 17 GRs were annotated from adult male and female C. cunea antennal transcriptomes. The expression profiles of 25 OBPs, 16 ORs, and 17 GRs, 5 CSP, 5 IRs and 1 SNMP were determined by RT-PCR and RT-qPCR for the antenna, head, thorax, and abdomen of male and female C. cunea. A total of 8 OBPs, 14 ORs, and 8 GRs, 1 CSP, 4 IRs and 1 SNMPs were exclusively or primarily expressed in female antennae. These female antennal-specific or dominant expression profiles may assist in locating suitable host and oviposition sites. These genes will provide useful targets for advanced study of their biological functions.

Olfaction plays a crucial role in insect population survival and reproduction. Identification of the genes associated with the olfactory system, without the doubt will promote studying the insect chemical communication system. In this study, RNA-seq technology was used to sequence the antennae transcriptome of Athetis dissimilis, an emerging crop pest in China with limited genomic information, with the purpose of identifying the gene set involved in olfactory recognition. Analysis of the transcriptome of female and male antennae generated 13.74 Gb clean reads in total from which 98,001 unigenes were assembled, and 25,930 unigenes were annotated. Total of 60 olfactory receptors (ORs), 18 gustatory receptors (GRs), and 12 ionotropic receptors (IRs) were identified by Blast and sequence similarity analyzes. One obligated olfactory receptor co-receptor (Orco) and four conserved sex pheromone receptors (PRs) were annotated in 60 ORs. Among the putative GRs, five genes (AdisGR1, 6, 7, 8 and 94) clustered in the sugar receptor family, and two genes (AdisGR3 and 93) involved in CO2 detection were identified. Finally, AdisIR8a.1 and AdisIR8a.2 co-receptors were identified in the group of candidate IRs. Furthermore, expression levels of these chemosensory receptor genes in female and male antennae were analyzed by mapping the Illumina reads.

Background: Since chemosensory genes play key roles in insect behaviour, they can potentially be used as new targets for pest control. The cabbage beetle, Colaphellus bowringi, is a serious insect pest of cruciferous vegetables in China and other Asian countries. However, a systematic identification of the chemosensory genes expressed in the antennae has not been reported., Results: We assembled the antennal transcriptome of C. bowringi by using Illumina sequencing technology and identified 104 candidate chemosensory genes by analyzing transcriptomic data, which included transcripts encoding 26 odorant-binding proteins (OBPs), 12 chemosensory proteins (CSPs), four sensory neuron membrane proteins (SNMPs), 43 odorant receptors (ORs), nine ionotropic receptors (IRs), and ten gustatory receptors (GRs). The data obtained are similar to those found in other coleopteran species, suggesting that our approach successfully identified the chemosensory genes of C. bowringi. The expression patterns of 43 OR genes, some of which were predominately found in the antenna or associated with sex-biased expression, were analyzed using quantitative real time RT-PCR (qPCR)., Conclusions: Our study revealed that a large number of chemosensory genes are expressed in C. bowringi. These candidate chemosensory genes and their expression profiles in various tissues provide further information on understanding their function in C. bowringi as well as other insects, and identifying potential targets to disrupt the odorant system in C. bowringi so that new methods for pest management can be developed.