Candidate Pheromone Receptors Research Articles

Prediction of a conserved pheromone receptor lineage from antennal transcriptomes of the pine sawyer genus Monochamus (Coleoptera: Cerambycidae).

Longhorned beetles (Cerambycidae) are a diverse family of wood-boring insects, many species of which produce volatile pheromones to attract mates over long distances. The composition and structure of the pheromones remain constant across many cerambycid species, and comparative studies of those groups could, therefore, reveal the chemoreceptors responsible for pheromone detection. Here, we use comparative transcriptomics to identify a candidate pheromone receptor in the large and economically important cerambycid genus Monochamus, males of which produce the aggregation-sex pheromone 2-(undecyloxy)-ethanol ("monochamol"). Antennal transcriptomes of the North American species M. maculosus, M. notatus, and M. scutellatus revealed 60-70 odorant receptors (ORs) in each species, including four lineages of simple orthologs that were highly conserved, highly expressed in both sexes, and upregulated in the flagellomeres where olfactory sensilla are localized. Two of these orthologous lineages, OR29 and OR59, remained highly expressed and conserved when we included a re-annotation of an antennal transcriptome of the Eurasian congener M. alternatus. OR29 is also orthologous to a characterized pheromone receptor in the cerambycid Megacyllene caryae, suggesting it as the most likely candidate for a monochamol receptor and highlighting its potential as a conserved lineage of pheromone receptors within one of the largest families of beetles.

Identification of Chemosensory Genes, Including Candidate Pheromone Receptors, in Phauda flammans (Walker) (Lepidoptera: Phaudidae) Through Transcriptomic Analyses.

Olfactory and gustatory systems play an irreplaceable role in all cycles of growth of insects, such as host location, mating, and oviposition. Many chemosensory genes in many nocturnal moths have been identified via omics technology, but knowledge of these genes in diurnal moths is lacking. In our recent studies, we reported two sex pheromone compounds and three host plant volatiles that play a vital role in attracting the diurnal moth, Phauda flammans. The antennal full-length transcriptome sequence of P. flammans was obtained using the Pacbio sequencing to further explore the process of sex pheromone and host plant volatile recognition in P. flammans. Transcriptome analysis identified 166 candidate olfactory and gustatory genes, including 58 odorant-binding proteins (OBPs), 19 chemosensory proteins (CSPs), 59 olfactory receptors (ORs), 16 ionotropic receptors (IRs), 14 gustatory receptors (GRs), and 2 sensory neuron membrane proteins (SNMPs). Subsequently, a phylogenetic tree was established using P. flammans and other lepidopteran species to investigate orthologs. Among the 17 candidate pheromone receptor (PR) genes, the expression levels of PflaOR21, PflaOR25, PflaOR35, PflaOR40, PflaOR41, PflaOR42, PflaOR44, PflaOR49, PflaOR51, PflaOR61, and PflaOR63 in the antennae were significantly higher than those in other non-antennae tissues. Among these PR genes, PflaOR21, PflaOR27, PflaOR29, PflaOR35, PflaOR37, PflaOR40, PflaOR42, PflaOR44, PflaOR60, and PflaOR62 showed male-biased expression, whereas PflaOR49, PflaOR61, and PflaOR63 revealed female-biased expression. The functions of related OR genes were also discussed. This research filled the gap of the chemosensory genes of P. flammans and provided basic data for future functional molecular mechanisms studies on P. flammans olfaction.

A small number of male-biased candidate pheromone receptors are expressed in large subsets of the olfactory sensory neurons in the antennae of drones from the European honey bee Apis mellifera.

In the European honey bee (Apis mellifera), the olfactory system is essential for foraging and intraspecific communication via pheromones. Honey bees are equipped with a large repertoire of olfactory receptors belonging to the insect odorant receptor (OR) family. Previous studies have indicated that the transcription level of a few OR types including OR11, a receptor activated by the queen-released pheromone compound (2E)-9-oxodecenoic acid (9-ODA), is significantly higher in the antenna of males (drones) than in female workers. However, the number and distribution of antennal cells expressing male-biased ORs is elusive. Here, we analyzed antennal sections from bees by in situ hybridization for the expression of the male-biased receptors OR11, OR18, and OR170. Our results demonstrate that these receptors are expressed in only moderate numbers of cells in the antennae of females (workers and queens), whereas substantially higher cell numbers express these ORs in drones. Thus, the reported male-biased transcript levels are due to sex-specific differences in the number of antennal cells expressing these receptors. Detailed analyses for OR11 and OR18 in drone antennae revealed expression in two distinct subsets of olfactory sensory neurons (OSNs) that in total account for approximately 69% of the OR-positive cells. Such high percentages of OSNs expressing given receptors are reminiscent of male-biased ORs in moths that mediate the detection of female-released sex pheromone components. Collectively, our findings indicate remarkable similarities between male antennae of bees and moths and support the concept that male-biased ORs in bee drones serve the detection of female-emitted sex pheromones.

Identification and functional characterization of sex pheromone receptors in mirid bugs (Heteroptera: Miridae)

Mirid bugs are a group of important insect pests that cause large annual losses in agricultural production. Many studies have focused on the isolation and identification of sex pheromones in mirid bugs, and the components and biological activity of the sex pheromones have also been studied as a way to control these pests. However, few studies have focused on the mechanisms of pheromone perception. In this study, we identified the odorant receptor repertoire in three mirid bug species, Apolygus lucorum, Adelphocoris lineolatus, and Adelphocoris suturalis using antennal transcriptome sequencing and bioinformatics analysis. The candidate pheromone receptor (PR) genes were then identified by comparative transcriptomic and expression pattern analysis. Importantly, in vitro functional studies have shown that the candidate PRs have robust responses to the main mirid bug sex pheromone components (E)-2-hexenyl butyrate (E2HB) and hexyl butyrate (HB). Our study uncovered the mechanism of pheromone peripheral coding in these three species and elucidated the mechanism by which mirid bugs can specifically recognize a mate. Moreover, the results of our study will provide a theoretical basis for screening effective sex attractants or mating disturbance agents at the molecular and neural levels for enhanced control of these destructive pests.

Identification and expression profiles of candidate chemosensory receptors in Histia rhodope (Lepidoptera: Zygaenidae).

Insect olfaction and vision play important roles in survival and reproduction. Diurnal butterflies mainly rely on visual cues whereas nocturnal moths rely on olfactory signals to locate external resources. Histia rhodope Cramer (Lepidoptera: Zygaenidae) is an important pest of the landscape tree Bischofia polycarpa in China and other Southeast Asian regions. As a diurnal moth, H. rhodope represents a suitable model for studying the evolutionary shift from olfactory to visual communication. However, only a few chemosensory soluble proteins have been characterized and information on H. rhodope chemoreceptor genes is currently lacking. In this study, we identified 45 odorant receptors (ORs), nine ionotropic receptors (IRs), eight gustatory receptors (GRs) and two sensory neuron membrane proteins (SNMPs) from our previously acquired H. rhodope antennal transcriptomic data. The number of chemoreceptors of H. rhodope was less compared with that found in many nocturnal moths. Some specific chemoreceptors such as OR co-receptor (ORco), ionotropic receptors co-receptor, CO2 receptors, sugar receptors and bitter receptors were predicted by phylogenetic analysis. Notably, two candidate pheromone receptors (PRs) were identified within a novel PR lineage. qRT-PCR results showed that almost all tested genes (22/24) were predominantly expressed in antennae, indicating that they may be important in olfactory function. Among these antennae-enriched genes, six ORs, five IRs and two GRs displayed female-biased expression, while two ORs displayed male-biased expression. Additionally, HrhoIR75q.2 and HrhoGR67 were more highly expressed in heads and legs. This study enriches the olfactory gene inventory of H. rhodope and provides the foundation for further research of the chemoreception mechanism in diurnal moths.

Evolution of sex pheromone receptors in Dendrolimus punctatus Walker (lepidoptera: Lasiocampidae) is divergent from other moth species

Dendrolimus punctatus Walker (Lepidoptera: Lasiocampidae) is a pine caterpillar moth distributed in most areas of southern China and is an economically important pest of pine, due to its defoliation activity. Understanding fundamental sex pheromone perception mechanisms in D. punctatus may provide effective and sustainable options for novel control strategies. However, the identification and function of pheromone receptors, key genes that receipt the pheromone of this pest, are both unclear now. Previous researches suggested several candidate pheromone receptors whose expression levels were male antennae bias in D. punctatus. In this study, we cloned six candidate pheromone receptors (DpunOR 20/45/46/51/54/58) and Orco from D. punctatus. Phylogenetic tree analysis showed that lepidopteran PRs tend to be conserved and clustered together; however, D. punctatus candidate PRs were located in a distinct clade. Motif analysis of PRs showed clear sequences differences between Dendrolimus spp. and other tested moth species. To illustrate the ligand response properties of the candidate PRs of D. punctatus, each of the six genes was expressed with an Orco gene in Xenopus oocytes and using two-electrode voltage-clamp recordings. Finally, we successfully identified two sex pheromone receptors (PR45 and PR46). Our study, which identified a novel lineage of PRs tuned to Type I pheromones in Lepidoptera, provides evidence for the new evolution origin of sex pheromone communication in moths, and lays a foundation for the development of novel control strategies of D. punctatus.

A novel lineage of candidate pheromone receptors for sex communication in moths.

Sex pheromone receptors (PRs) are key players in chemical communication between mating partners in insects. In the highly diversified insect order Lepidoptera, male PRs tuned to female-emitted type I pheromones (which make up the vast majority of pheromones identified) form a dedicated subfamily of odorant receptors (ORs). Here, using a combination of heterologous expression and in vivo genome editing methods, we bring functional evidence that at least one moth PR does not belong to this subfamily but to a distantly related OR lineage. This PR, identified in the cotton leafworm Spodoptera littoralis, is highly expressed in male antennae and is specifically tuned to the major sex pheromone component emitted by females. Together with a comprehensive phylogenetic analysis of moth ORs, our functional data suggest two independent apparitions of PRs tuned to type I pheromones in Lepidoptera, opening up a new path for studying the evolution of moth pheromone communication.

Functional characterization of odorant receptors from the moth Eriocrania semipurpurella: A comparison of results in the Xenopus oocyte and HEK cell systems

The Xenopus oocyte and the Human Embryonic Kidney (HEK) 293 cell expression systems are frequently used for functional characterization (deorphanization) of insect odorant receptors (ORs). However, the inherent characteristics of these heterologous systems differ in several aspects, which raises the question of whether the two systems provide comparable results, and how well the results correspond to the responses obtained from olfactory sensory neurons in vivo. Five candidate pheromone receptors were previously identified in the primitive moth Eriocrania semipurpurella (Esem) and their responses were characterized in HEK cells. We re-examined the responses of these five EsemORs in Xenopus oocytes. We showed that in both systems, EsemOR1 specifically responded to the plant volatile β-caryophyllene. EsemOR3 responded stronger to the pheromone component (S,Z)-6-nonen-2-ol than to its enantiomer (R,Z)-6-nonen-2-ol, the second pheromone component. However, EsemOR3 also responded secondarily to the plant volatile β-caryophyllene in the oocyte system, but not in the HEK cell system. EsemOR4 was unresponsive in the HEK cells, but responded primarily to (R,Z)-6-nonen-2-ol followed by (S,Z)-6-nonen-2-ol in the oocytes, representing a discovery of a new pheromone receptor in this species. EsemOR5 was broadly tuned in both systems, but the rank order among the most active pheromone compounds and antagonists was different. EsemOR6 showed no response to any compound in either system. We compared the results obtained in the two different heterologous systems with the activity previously recorded in vivo, and performed in situ hybridization to localize the expression of these OR genes in the antennae. In spite of similar results overall, differences in OR responses between heterologous expression systems suggest that conclusions about the function of individual ORs may differ depending on the system used for deorphanization.

A Subset of Odorant Receptors from the Desert Locust Schistocerca gregaria Is Co-Expressed with the Sensory Neuron Membrane Protein 1.

In the desert locust Schistocerca gregaria (S. gregaria), pheromones are considered to be crucial for governing important behaviors and processes, including phase transition, reproduction, aggregation and swarm formation. The receptors mediating pheromone detection in olfactory sensory neurons (OSNs) on the antenna of S. gregaria are unknown. Since pheromone receptors in other insects belong to the odorant receptor (OR) family and are typically co-expressed with the “sensory neuron membrane protein 1” (SNMP1), in our search for putative pheromone receptors of S. gregaria, we have screened the OR repertoire for receptor types that are expressed in SNMP1-positive OSNs. Based on phylogenetic analyses, we categorized the 119 ORs of S. gregaria into three groups (I–III) and analyzed a substantial number of ORs for co-expression with SNMP1 by two-color fluorescence in situ hybridization. We have identified 33 ORs that were co-expressed with SNMP1. In fact, the majority of ORs from group I and II were found to be expressed in SNMP1-positive OSNs, but only very few receptors from group III, which comprises approximately 60% of all ORs from S. gregaria, were co-expressed with SNMP1. These findings indicate that numerous ORs from group I and II could be important for pheromone communication. Collectively, we have identified a broad range of candidate pheromone receptors in S. gregaria that are not randomly distributed throughout the OR family but rather segregate into phylogenetically distinct receptor clades.

Molecular Characterization of MbraOR16, a Candidate Sex Pheromone Receptor in Mamestra brassicae (Lepidoptera: Noctuidae).

Sex pheromone communication in Lepidoptera has long been a valuable model system for studying fundamental aspects of olfaction and its study has led to the establishment of environmental-friendly pest control strategies. The cabbage moth, Mamestra brassicae (Linnaeus) (Lepidoptera: Noctuidae), is a major pest of Cruciferous vegetables in Europe and Asia. Its sex pheromone has been characterized and is currently used as a lure to trap males; however, nothing is known about the molecular mechanisms of sex pheromone reception in male antennae. Using homology cloning and rapid amplification of cDNA ends-PCR strategies, we identified the first candidate pheromone receptor in this species. The transcript was specifically expressed in the antennae with a strong male bias. In situ hybridization experiments within the antennae revealed that the receptor-expressing cells were closely associated with the olfactory structures, especially the long trichoid sensilla known to be pheromone-sensitive. The deduced protein is predicted to adopt a seven-transmembrane structure, a hallmark of insect odorant receptors, and phylogenetically clustered in a clade that grouped a majority of the Lepidoptera pheromone receptors characterized to date. Taken together, our data support identification of a candidate pheromone receptor and provides a basis for better understanding how this species detects a signal critical for reproduction.

Cloning and functional characterization of three new pheromone receptors from the diamondback moth, Plutella xylostella

The highly specialized olfactory receptor neurons (ORNs) on the antennae of male moths can recognize blends of several pheromone components. In previous studies, a total of six candidate pheromone receptor (PR) genes were cloned and functionally characterized in the diamondback moth, Plutella xylostella. In the present work, we report on three novel candidate pheromone receptor genes: PxylOR8, PxylOR41, and PxylOR45 in the same species. Gene expression analysis revealed that PxylOR8 is specifically expressed in female adult antennae, while PxylOR41 and PxylOR45 are expressed in antennae in both sexes, but with a male bias. In situ hybridization revealed that PxylOR8, PxylOR41 and PxylOR45 are localized in long trichoid sensilla. Functional analyses on the three pheromone receptor genes were then performed using the heterologous expression system of Xenopus oocytes. PxylOR41 was tuned to two minor pheromone components Z9-14:Ac, Z9-14:OH, and their analog Z9-14:Ald. PxylOR8 and PxylOR45 did not respond to any tested pheromone components and analogs. These results may contribute to clarifying how pheromone detection works in P. xylostella.

Antennal transcriptome analysis of the piercing moth Oraesia emarginata (Lepidoptera: Noctuidae).

The piercing fruit moth Oraesia emarginata is an economically significant pest; however, our understanding of its olfactory mechanisms in infestation is limited. The present study conducted antennal transcriptome analysis of olfactory genes using real-time quantitative reverse transcription PCR analysis (RT-qPCR). We identified a total of 104 candidate chemosensory genes from several gene families, including 35 olfactory receptors (ORs), 41 odorant-binding proteins, 20 chemosensory proteins, 6 ionotropic receptors, and 2 sensory neuron membrane proteins. Seven candidate pheromone receptors (PRs) and 3 candidate pheromone-binding proteins (PBPs) for sex pheromone recognition were found. OemaOR29 and OemaPBP1 had the highest fragments per kb per million fragments (FPKM) values in all ORs and OBPs, respectively. Eighteen olfactory genes were upregulated in females, including 5 candidate PRs, and 20 olfactory genes were upregulated in males, including 2 candidate PRs (OemaOR29 and 4) and 2 PBPs (OemaPBP1 and 3). These genes may have roles in mediating sex-specific behaviors. Most candidate olfactory genes of sex pheromone recognition (except OemaOR29 and OemaPBP3) in O. emarginata were not clustered with those of studied noctuid species (type I pheromone). In addition, OemaOR29 was belonged to cluster PRIII, which comprise proteins that recognize type II pheromones instead of type I pheromones. The structure and function of olfactory genes that encode sex pheromones in O. emarginata might thus differ from those of other studied noctuids. The findings of the present study may help explain the molecular mechanism underlying olfaction and the evolution of olfactory genes encoding sex pheromones in O. emarginata.

Candidate pheromone receptors of codling moth Cydia pomonella respond to pheromones and kairomones

Olfaction plays a dominant role in the mate-finding and host selection behaviours of the codling moth (Cydia pomonella), an important pest of apple, pear and walnut orchards worldwide. Antennal transcriptome analysis revealed a number of abundantly expressed genes related to the moth olfactory system, including those encoding the olfactory receptors (ORs) CpomOR1, CpomOR3 and CpomOR6a, which belong to the pheromone receptor (PR) lineage, and the co-receptor (CpomOrco). Using heterologous expression, in both Drosophila olfactory sensory neurones and in human embryonic kidney cells, together with electrophysiological recordings and calcium imaging, we characterize the basic physiological and pharmacological properties of these receptors and demonstrate that they form functional ionotropic receptor channels. Both the homomeric CpomOrco and heteromeric CpomOrco + OR complexes can be activated by the common Orco agonists VUAA1 and VUAA3, as well as inhibited by the common Orco antagonists amiloride derivatives. CpomOR3 responds to the plant volatile compound pear ester ethyl-(E,Z)-2,4-decadienoate, while CpomOR6a responds to the strong pheromone antagonist codlemone acetate (E,E)-8,10-dodecadien-1-yl acetate. These findings represent important breakthroughs in the deorphanization of codling moth pheromone receptors, as well as more broadly into insect ecology and evolution and, consequently, for the development of sustainable pest control strategies based on manipulating chemosensory communication.

In Search for Pheromone Receptors: Certain Members of the Odorant Receptor Family in the Desert Locust Schistocerca gregaria (Orthoptera: Acrididae) Are Co-expressed with SNMP1.

Under given environmental conditions, the desert locust (Schistocera gregaria) forms destructive migratory swarms of billions of animals, leading to enormous crop losses in invaded regions. Swarm formation requires massive reproduction as well as aggregation of the animals. Pheromones that are detected via the olfactory system have been reported to control both reproductive and aggregation behavior. However, the molecular basis of pheromone detection in the antennae of Schistocerca gregaria is unknown. As an initial step to disclose pheromone receptors, we sequenced the antennal transcriptome of the desert locust. By subsequent bioinformatical approaches, 119 distinct nucleotide sequences encoding candidate odorant receptors (ORs) were identified. Phylogenetic analyses employing the identified ORs from Schistocerca gregaria (SgreORs) and OR sequences from the related species Locusta migratoria revealed a group of locust ORs positioned close to the root, i.e. at a basal site in a phylogenetic tree. Within this particular OR group (termed basal or b-OR group), the locust OR sequences were strictly orthologous, a trait reminiscent of pheromone receptors from lepidopteran species. In situ hybridization experiments with antennal tissue demonstrated expression of b-OR types from Schistocerca gregaria in olfactory sensory neurons (OSNs) of either sensilla trichodea or sensilla basiconica, both of which have been reported to respond to pheromonal substances. More importantly, two-color fluorescent in situ hybridization experiments showed that most b-OR types were expressed in cells co-expressing the “sensory neuron membrane protein 1” (SNMP1), a marker indicative of pheromone-sensitive OSNs in insects. Analyzing the expression of a larger number of SgreOR types outside the b-OR group revealed that only a few of them were co-expressed with SNMP1.In summary, we have identified several candidate pheromone receptors from Schistocerca gregaria that could mediate responses to pheromones implicated in controlling reproduction and aggregation behavior.

Molecular and functional characterization of a candidate sex pheromone receptor OR1 in Spodoptera litura.

Olfaction is primarily mediated by highly specified olfactory receptors (ORs). Here, we cloned and identified an olfactory receptor, named SlituOR1 (Genbank no. JN835269), from Spodoptera litura and found evidence that it is a candidate pheromone receptor. It exhibited male-biased expression in the antennae, where it was localized at the base of sensilla trichoidea, the antennal sensilla mainly responsive to pheromones in moths. Conserved orthologues of this receptor, found among known pheromone receptors within the Lepidoptera, and SlituOR1 were placed among a clade of candidate pheromone receptors in a phylogeny tree of insect OR gene sequences. SlituOR1 showed differential expression in S. litura populations attracted to traps baited with different ratios of the two sex pheromone components (9Z,11E)-tetradecadienyl acetate (Z9E11-14:OAc) and (9Z,12E)-tetradecadienyl acetate (Z9E12-14:OAc). Knocking down of SlituOR1 by RNA interference reduced the electroantennogram (EAG) response to Z9E11-14:OAc, and this result is consistent with the field trapping experiment. We infer that variation in transcription levels of olfactory receptors may modulate sex pheromone perception in male moths and could provide some of the flexibility required to maintain the functionality of communication with females when a population is adapting to a new niche and reproductive isolation becomes an advantage.

Alternative splicing produces two transcripts encoding female-biased pheromone subfamily receptors in the navel orangeworm, Amyelois transitella

Insect odorant receptors are key sensors of environmental odors and members of the lepidopteran pheromone receptor subfamily are thought to play important roles in mate finding by recognizing sex pheromones. Much research has been done to identify putative pheromone receptors in lepidopteran males, but little attention has been given to female counterparts. In this study, degenerate oligonucleotide primers designed against a conserved amino acid region in the C-terminus of lepidopteran pheromone receptors were used in 3’ RACE reactions to identify candidate pheromone receptors expressed in the antennae of female navel orangeworm. Two near full-length transcripts of 1469 nt and 1302 nt encoding the complete open reading frames for proteins of 446 and 425 amino acids, respectively, were identified. Based on BLAST homology and phylogenetic analyses, the putative proteins encoded by these transcripts are members of the lepidopteran pheromone receptor subfamily. Characterization of these transcripts indicates that they are alternatively spliced products of a single gene. Tissue expression studies indicate that the transcripts are female-biased with detection mainly in female antennae. To the best of our knowledge, these transcripts represent the first detection of alternatively spliced female-biased members of the lepidopteran pheromone receptor subfamily.

Evolution of two receptors detecting the same pheromone compound in crop pest moths of the genus Spodoptera

In moths, mate finding strongly rely on the detection of sex pheromones by pheromone receptors. Any modification in the functional properties of these receptors can have a drastic impact on reproduction. In the course of characterizing candidate pheromone receptors in the noctuid moth Spodoptera littoralis, we expressed them in Drosophila olfactory sensory neurons and stimulated them with a large panel of moth pheromone compounds. We found that two pheromone receptors detect (Z,E)-9,12-14:OAc, a minor component of the female pheromone blend. Whereas SlitOR6 is highly specific to this component, SlitOR13 is less sensitive and not strictly specific as it also detects (Z)9-14:OAc, another minor component of the sex pheromone. Interestingly, SlitOR13 expression is restricted to the distal part of male antennae, where we could identify a novel functional class of pheromone-sensitive neurons whose response spectrum matches that of SlitOR13. Based on a phylogenetic analysis of Lepidoptera pheromone receptors, we built an evolutionary scenario in which four different paralogous lineages emerged through gene duplications. The ability to bind (Z,E)-9,12-14:OAc appeared independently within three of these lineages, and an analysis of selective pressures revealed sites under positive selection that could have played a role in the emergence of functional properties of OR6 and OR13 in Spodoptera species.

Genes involved in sex pheromone biosynthesis of Ephestia cautella, an important food storage pest, are determined by transcriptome sequencing.

BackgroundInsects use pheromones, chemical signals that underlie all animal behaviors, for communication and for attracting mates. Synthetic pheromones are widely used in pest control strategies because they are environmentally safe. The production of insect pheromones in transgenic plants, which could be more economical and effective in producing isomerically pure compounds, has recently been successfully demonstrated. This research requires information regarding the pheromone biosynthetic pathways and the characterization of pheromone biosynthetic enzymes (PBEs). We used Illumina sequencing to characterize the pheromone gland (PG) transcriptome of the Pyralid moth, Ephestia cautella, a destructive storage pest, to reveal putative candidate genes involved in pheromone biosynthesis, release, transport and degradation.ResultsWe isolated the E. cautella pheromone compound as (Z,E)-9,12-tetradecadienyl acetate, and the major pheromone precursors 16:acyl, 14:acyl, E14-16:acyl, E12-14:acyl and Z9,E12-14:acyl. Based on the abundance of precursors, two possible pheromone biosynthetic pathways are proposed. Both pathways initiate from C16:acyl-CoA, with one involving ∆14 and ∆9 desaturation to generate Z9,E12-14:acyl, and the other involving the chain shortening of C16:acyl-CoA to C14:acyl-CoA, followed by ∆12 and ∆9 desaturation to generate Z9,E12-14:acyl-CoA. Then, a final reduction and acetylation generates Z9,E12-14:OAc. Illumina sequencing yielded 83,792 transcripts, and we obtained a PG transcriptome of ~49.5 Mb. A total of 191 PBE transcripts, which included pheromone biosynthesis activating neuropeptides, fatty acid transport proteins, acetyl-CoA carboxylases, fatty acid synthases, desaturases, β-oxidation enzymes, fatty acyl-CoA reductases (FARs) and fatty acetyltransferases (FATs), were selected from the dataset. A comparison of the E. cautella transcriptome data with three other Lepidoptera PG datasets revealed that 45 % of the sequences were shared. Phylogenetic trees were constructed for desaturases, FARs and FATs, and transcripts that clustered with the ∆14, ∆12 and ∆9 desaturases, PG-specific FARs and potential candidate FATs, respectively, were identified. Transcripts encoding putative pheromone degrading enzymes, and candidate pheromone carrier and receptor proteins expressed in the E. cautella PG, were also identified.ConclusionsOur study provides important background information on the enzymes involved in pheromone biosynthesis. This information will be useful for the in vitro production of E. cautella sex pheromones and may provide potential targets for disrupting the pheromone-based communication system of E. cautella to prevent infestations.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1710-2) contains supplementary material, which is available to authorized users.

Identification and Differential Expression of a Candidate Sex Pheromone Receptor in Natural Populations of Spodoptera litura.

Olfaction is primarily mediated by highly specific olfactory receptors (ORs), a subfamily of which are the pheromone receptors that play a key role in sexual communication and can contribute to reproductive isolation. Here we cloned and identified an olfactory receptor, SlituOR3 (Genbank NO. JN835270), from Spodoptera litura, to be the candidate pheromone receptor. It exhibited male-biased expression in the antennae, where they were localized at the base of sensilla trichoidea. Conserved orthologues of these receptors were found amongst known pheromone receptors within the Lepidoptera, and SlituOR3 were placed amongst a clade of candidate pheromone receptors in a phylogeny tree of insect ORs. SlituOR3 is required for the EAG responses to both Z9E11-14:OAc and Z9E12-14:OAc SlituOR3 showed differential expression in S. litura populations attracted to traps baited with a series of sex pheromone blends composed of different ratios of (9Z,11E)-tetradecadienyl acetate (Z9E11-14:OAc) and (9Z,12E)-tetradecadienyl acetate (Z9E12-14:OAc). The changes in the expression level of SlitOR3 and antennal responses after SlitOR3 silencing suggested that SlitOR3 is required for the sex pheromone signaling. We infer that variation in transcription levels of olfactory receptors may modulate sex pheromone perception in male moths and could affect both of pest control and monitoring efficiency by pheromone application after long time mass trapping with one particular ratio of blend in the field.

Transcriptome and expression profiling analysis link patterns of gene expression to antennal responses in Spodoptera litura.

BackgroundThe study of olfaction is key to understanding the interaction of insects with their environment and provides opportunities to develop novel tactics for control of pest species. Recent developments in transcriptomic approaches enable the molecular basis of olfaction to be studied even in species with limited genomic information. Here we use transcriptome and expression profiling analysis to characterize the antennal transcriptome of the noctuid moth and polyphagous pest Spodoptera litura.ResultsWe identify 74 candidate genes involved in odor detection and recognition, encoding 26 ORs, 21 OBPs, 18 CSPs and 9 IRs. We examine their expression levels in both sexes and seek evidence for their function by relating their expression with levels of EAG response in male and female antennae to 58 host and non-host plant volatiles and sex pheromone components. The majority of olfactory genes showed sex-biased expression, usually male-biased in ORs. A link between OR gene expression and antennal responses to odors was evident, a third of the compounds tested evoking a sex-biased response, in every case also male-biased. Two candidate pheromone receptors, OR14 and OR23 were especially strongly expressed and male-biased and we suggest that these may respond to the two female sex pheromone components of S. litura, Z9E11-14:OAc and Z9E12-14:OAc, which evoked strongly male-biased EAG responses.ConclusionsOur results provide the molecular basis for elucidating the olfactory profile of moths and the sexual divergence of their behavior and could enable the targeting of particular genes, and behaviors for pest management.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1375-x) contains supplementary material, which is available to authorized users.