The evolution of borneol repellency in culicine mosquitoes.

Insects have developed remarkable adaptations to effectively interact with plant secondary metabolites and utilize them as cues to identify suitable hosts. Consequently, humans have used aromatic plants for centuries to repel mosquitoes. The repellent effects of plant volatile compounds are mediated through olfactory structures present in the antennae, and maxillary palps of mosquitoes. Mosquito maxillary palps contain capitate-peg sensilla, which house three olfactory sensory neurons, of which two are mainly tuned to either carbon dioxide or octenol - two animal host odorants. However, the third neuron, which expresses the OR49 receptor, has remained without a known ecologically-relevant odorant since its initial discovery. In this study, we used odorant mixtures and terpenoid-rich Cannabis essential oils to investigate the activation of OR49. Our results demonstrate that two monoterpenoids, borneol and camphor, selectively activate OR49, and OR9-expressing neurons, as well as the MD3 glomerulus in the antennal lobe. We confirm that borneol repels female mosquitoes, and knocking out the gene encoding the OR49 receptor suppresses the response of the corresponding olfactory sensory neuron. Importantly, this molecular mechanism of action is conserved across culicine mosquito species, underscoring its significance in their olfactory systems.

Carboxylic acids that drive mosquito attraction to humans activate ionotropic receptors.

The mosquito, Aedes aegypti, is highly anthropophilic and transmits debilitating arboviruses within human populations and between humans and non-human primates. Female mosquitoes are attracted to sources of blood by responding to odor plumes that are emitted by their preferred hosts. Acidic volatile compounds, including carboxylic acids, represent particularly salient odors driving this attraction. Importantly, carboxylic acids are major constituents of human sweat and volatiles generated by skin microbes. As such, they are likely to impact human host preference, a dominant factor in disease transmission cycles. A more complete understanding of mosquito host attraction will necessitate the elucidation of molecular mechanisms of volatile odor detection that function in peripheral sensory neurons. Recent studies have shown that members of the variant ionotropic glutamate receptor gene family are necessary for physiological and behavioral responses to acidic volatiles in Aedes. In this study, we have identified a subfamily of variant ionotropic receptors that share sequence homology across several important vector species and are likely to be activated by carboxylic acids. Moreover, we demonstrate that selected members of this subfamily are activated by short-chain carboxylic acids in a heterologous cell expression system. Our results are consistent with the hypothesis that members of this receptor class underlie acidic volatile sensitivity in vector mosquitoes and provide a frame of reference for future development of novel mosquito attractant and repellent technologies.

Functional conservation of Anopheline linalool receptors through 100 million years of evolution.

Insects rely on olfactory receptors to detect and respond to diverse environmental chemical cues. Detection of semiochemicals by these receptors modulates insect behavior and has a direct impact on species fitness. Volatile organic compounds (VOCs) are released by animals and plants and can provide contextual cues that a blood meal host or nectar source is present. One such VOC is linalool, an enantiomeric monoterpene, that is emitted from plants and bacteria species. This compound exists in nature as one of two possible stereoisomers, (R)-(-)-linalool or (S)-(+)-linalool. In this study, we use a heterologous expression system to demonstrate differential responsiveness of a pair of Anopheline odorant receptors (Ors) to enantiomers of linalool. The mosquitoes Anopheles gambiae and Anopheles stephensi encode single copies of Or29 and Or53, which are expressed in the labella of An. gambiae. (S)-(+)-linalool activates Or29 orthologs with a higher potency than (R)-(-)-linalool, while the converse is observed for Or53 orthologs. The conservation of these receptors across a broad range of Anopheline species suggests they may function in the discrimination of linalool stereoisomers, thereby influencing the chemical ecology of mosquitoes. One potential application of this knowledge would be in the design of novel attractants or repellents to be used in integrated pest management practices.

Identification and functional characterization of olfactory indolergic receptors in Musca domestica.

In mosquitoes, indolic compounds are detected by a group of olfactory indolergic Odorant Receptors (indolORs). The ancient origin of indole and 3-methylindole as chemical signals suggest that they may be detected by insects outside the Culicidae clade. To test this hypothesis, we have identified potential indolOR genes in brachyceran flies based on sequence homology. Because of the crucial roles of indolic compounds in oviposition and foraging, we have focused our attention on the housefly Musca domestica. Using a heterologous expression system, we have identified indolOR transcript expression in the female antennae, and have characterized MdomOR30a and MdomOR49b as 3-methylindole and indole receptors, respectively. We have identified a set of 92 putative indolOR genes encoded in the genomes of Culicoidea, Psychodidae and brachycera, described their phylogenetic relationships, and exon/intron structures. Further characterization of indolORs will impact our understanding of insect chemical ecology and will provide targets for the development of novel odor-based tools that can be integrated into existing vector surveillance and control programs.

Carboxylic acid responses by a conserved odorant receptor in culicine vector mosquitoes.

Many mosquito behaviours that are critical for survival and reproduction depend upon timely responses to chemical cues. Of interest are the effects of volatile organic compounds like carboxylic acids (CAs) that are released by potential blood meal hosts. Short chain CAs are among the primary attractants for host-seeking females and influence host selection in vector species. Although the behavioural relevance of CA's has been established, less is known about the molecular receptive events that evoke responses to specific compounds, with the Ir family of chemoreceptors being broadly implicated in their detection. In this study, we demonstrate that Or orthologs from two vector species, Aedes aegypti (L.) and Aedes albopictus (Skuse), are selectively activated by straight chain carboxylic acids and that these responses are attenuated by the commercial insect repellant N,N-Diethyl-meta-toluamide. Our results suggest that multiple chemoreceptors, representing diverse families, are able to mediate molecular responses to CAs and may therefore underlie important behaviours that directly impact disease-transmission cycles.

Identification of Mosquito Bloodmeals Collected in Diverse Habitats in Malaysian Borneo Using COI Barcoding.

Land cover and land use change (LCLUC) acts as a catalyst for spillover of arthropod-borne pathogens into novel hosts by shifting host and vector diversity, abundance, and distribution, ultimately reshaping host-vector interactions. Identification of bloodmeals from wild-caught mosquitoes provides insight into host utilization of particular species in particular land cover types, and hence their potential role in pathogen maintenance and spillover. Here, we collected 134 blood-engorged mosquitoes comprising 10 taxa across 9 land cover types in Sarawak, Malaysian Borneo, a region experiencing intense LCLUC and concomitant spillover of arthropod-borne pathogens. Host sources of blood were successfully identified for 116 (87%) mosquitoes using cytochrome oxidase subunit I (COI) barcoding. A diverse range of hosts were identified, including reptiles, amphibians, birds, and mammals. Sixteen engorged Aedes albopictus, a major vector of dengue virus, were collected from seven land cover types and found to feed exclusively on humans (73%) and boar (27%). Culex tritaeniohynchus (n = 2), Cx. gelidus (n = 3), and Cx. quiquefasciatus (n = 3), vectors of Japanese encephalitis virus, fed on humans and pigs in the rural built-up land cover, creating potential transmission networks between these species. Our data support the use of COI barcoding to characterize mosquito-host networks in a biodiversity hotspot.