No abstract

The lightbrown apple moth, Epiphyas postvittana is an increasingly global pest of horticultural crops. Like other moths, E. postvittana relies on olfactory cues to locate mates and oviposition sites. To detect these cues, moths have evolved families of genes encoding elements of the peripheral olfactory reception system, including odor carriers, receptors and degrading enzymes. Here we undertake a transcriptomic approach to identify members of these families expressed in the adult antennae of E. postvittana, describing open reading frames encoding 34 odorant binding proteins, 13 chemosensory proteins, 70 odorant receptors, 19 ionotropic receptors, nine gustatory receptors, two sensory neuron membrane proteins, 27 carboxylesterases, 20 glutathione-S-transferases, 49 cytochrome p450s and 18 takeout proteins. For the odorant receptors, quantitative RT-PCR corroborated RNAseq count data on steady state transcript levels. Of the eight odorant receptors that group phylogenetically with pheromone receptors from other moths, two displayed significant male-biased expression patterns, one displayed significant female-biased expression pattern and five were expressed equally in the antennae of both sexes. In addition, we found two male-biased odorant receptors that did not group with previously described pheromone receptors. This suite of olfaction-related genes provides a substantial resource for the functional characterization of this signal transduction system and the development of odor-mediated control strategies for horticultural pests.

Pheromone receptors (PRs) are essential in moths to detect sex pheromones for mate finding. However, it remains unknown from which ancestral proteins these specialized receptors arose. The oldest lineages of moths, so-called non-ditrysian moths, use short-chain pheromone components, secondary alcohols, or ketones, so called Type 0 pheromones that are similar to many common plant volatiles. It is, therefore, possible that receptors for these ancestral pheromones evolved from receptors detecting plant volatiles. Hence, we identified the odorant receptors (ORs) from a non-ditrysian moth, Eriocrania semipurpurella (Eriocraniidae, Lepidoptera), and performed functional characterization of ORs using HEK293 cells. We report the first receptors that respond to Type 0 pheromone compounds; EsemOR3 displayed highest sensitivity toward (2S, 6Z)-6-nonen-2-ol, whereas EsemOR5 was most sensitive to the behavioral antagonist (Z)-6-nonen-2-one. These receptors also respond to plant volatiles of similar chemical structures, but with lower sensitivity. Phylogenetically, EsemOR3 and EsemOR5 group with a plant volatile-responding receptor from the tortricid moth Epiphyas postvittana (EposOR3), which together reside outside the previously defined lepidopteran PR clade that contains the PRs from more derived lepidopteran families. In addition, one receptor (EsemOR1) that falls at the base of the lepidopteran PR clade, responded specifically to β-caryophyllene and not to any other additional plant or pheromone compounds. Our results suggest that PRs for Type 0 pheromones have evolved from ORs that detect structurally-related plant volatiles. They are unrelated to PRs detecting pheromones in more derived Lepidoptera, which, in turn, also independently may have evolved a novel function from ORs detecting plant volatiles.

Feces is an abundant, rich source of energy, utilized by a myriad of organisms, not least by members of the order Diptera, i.e., flies. How Drosophila melanogaster reacts to fecal matter remains unclear. Here, we examined oviposition behavior toward a range of fecal samples from mammals native to the putative Southeast African homeland of the fly. We show that D. melanogaster display a strong oviposition aversion toward feces from carnivorous mammals but indifference or even attraction toward herbivore dung. We identify a set of four predictor volatiles, which can be used to differentiate fecal from non-fecal matter, as well as separate carnivore from herbivore feces. Of these volatiles, phenol-indicative of carnivore feces-confers egg-laying aversion and is detected by a single class of sensory neurons expressing Or46a. The Or46a-expressing neurons are necessary and sufficient for oviposition site aversion. We further demonstrate that carnivore feces-unlike herbivore dung-contain a high rate of pathogenic bacteria taxa. These harmful bacteria produce phenol from L-tyrosine, an amino acid specifically enriched in high protein diets, such as consumed by carnivores. Finally, we demonstrate that carnivore feces, as well as phenol, is also avoided by a ball-rolling species of dung beetle, suggesting that phenol is a widespread avoidance signal because of its association with pathogenic bacteria.

The Hessian fly, Mayetiola destructor Say (Diptera, Cecidomyiidae), is a pest of wheat and belongs to a group of gall-inducing herbivores. This species has a unique life history and several ecological features that differentiate it from other Diptera such as Drosophila melanogaster and blood-feeding mosquitoes. These features include a short, non-feeding adult life stage (1–2 days) and the use of a long-range sex pheromone produced and released by adult females. Sex pheromones are detected by members of the odorant receptor (OR) family within the Lepidoptera, but no receptors for similar long-range sex pheromones have been characterized from the Diptera. Previously, 122 OR genes have been annotated from the Hessian fly genome, with many of them showing sex-biased expression in the antennae. Here we have expressed, in HEK293 cells, five MdesORs that display male-biased expression in antennae, and we have identified MdesOR115 as a Hessian fly sex pheromone receptor. MdesOR115 responds primarily to the sex pheromone component (2S,8E,10E)-8,10-tridecadien-2-yl acetate, and secondarily to the corresponding Z,E-isomer. Certain sensory neuron membrane proteins (i.e., SNMP1) are important for responses of pheromone receptors in flies and moths. The Hessian fly genome is unusual in that it encodes six SNMP1 paralogs, of which five are expressed in antennae. We co-expressed each of the five antennal SNMP1 paralogs together with each of the five candidate sex pheromone receptors from the Hessian fly and found that they do not influence the response of MdesOR115, nor do they confer responsiveness in any of the non-responsive ORs to any of the sex pheromone components identified to date in the Hessian fly. Using Western blots, we detected protein expression of MdesOrco, all MdesSNMPs, and all MdesORs except for MdesOR113, potentially explaining the lack of response from this OR. In conclusion, we report the first functional characterization of an OR from the Cecidomyiidae, extending the role of ORs as long-range sex pheromone detectors from the Lepidoptera into the Diptera.

The odorant receptors (ORs) of insects are crucial for host and mate recognition. In moths (Lepidoptera), specialized ORs are involved in male detection of the sex pheromone produced by females. Most moth sex pheromones are C-C acetates, alcohols, and aldehydes (Type I pheromones), and most pheromone receptors (PRs) characterized to date are from higher Lepidoptera (Ditrysia), responding to these types of compounds. With few exceptions, functionally characterized PRs fall into what has been called the "PR-clade", which also contains receptors that have yet to be characterized. While it has been suggested that moth PRs have evolved from plant odor-detecting ORs, it is not known when receptors for Type I pheromones arose. This is largely due to a lack of functionally characterized PRs from non-ditrysian Lepidoptera. The currant shoot borer moth, Lampronia capitella (Prodoxidae), belongs to a non-ditrysian lineage, and uses Type I pheromone compounds. We identified 53 ORs from antennal transcriptomes of this species, and analyzed their phylogenetic relationships with known lepidopteran ORs. Using a HEK293 cell-based assay, we showed that three of the LcapORs with male-biased expression (based on FPKM values) respond to Type I pheromone compounds. Two of them responded to pheromone components of L. capitella and one to a structurally related compound. These PRs are the first from a non-ditrysian moth species reported to respond to Type I compounds. They belong to two of the more early-diverging subfamilies of the PR-clade for which a role in pheromone detection had not previously been demonstrated. Hence, our definition of the monophyletic lepidopteran PR-clade includes these receptors from a non-ditrysian species, based on functional support.

No abstract

Insect olfactory receptors are routinely expressed in heterologous systems for functional characterisation. It was recently discovered that the essential olfactory receptor co-receptor (Orco) of the Hessian fly, Mayetiola destructor (Mdes), does not respond to the agonist VUAA1, which activates Orco in all other insects analysed to date. Here, using a mutagenesis-based approach we identified three residues in MdesOrco, located in different transmembrane helices as supported by 3D modelling, that confer sensitivity to VUAA1. Reciprocal mutations in Drosophila melanogaster (Dmel) and the noctuid moth Agrotis segetum (Aseg) Orcos diminish sensitivity of these proteins to VUAA1. Additionally, mutating these residues in DmelOrco and AsegOrco compromised odourant receptor (OR) dependent ligand-induced Orco activation. In contrast, both wild-type and VUAA1-sensitive MdesOrco were capable of forming functional receptor complexes when coupled to ORs from all three species, suggesting unique complex properties in M. destructor, and that not all olfactory receptor complexes are “created” equal.