Pheromones and other semiochemicals play a crucial role in today's integrated pest and vector management strategies. These semiochemicals are typically discovered by bioassay-guided approaches. Here, we applied a reverse chemical ecology approach; that is, we used olfactory proteins to lead us to putative semiochemicals. Specifically, we used 7 of the top 10 odorant receptors (ORs) most expressed in the antennae of the southern house mosquito, Culex quinquefasciatus, and which are yet to be deorphanized. We expressed these receptors in the Xenopus oocyte recording system and challenged them with a panel of 230 odorants, including physiologically and behaviorally active compounds. Six of the ORs were silent either because they are not functional or a key odorant was missing. CquiOR36, which showed the highest transcript levels of all OR genes in female antennae, was also silent to all odorants in the tested panel, but yielded robust responses when it was accidentally challenged with an old sample of nonanal in ethanol. After confirming that fresh samples were inactive and through a careful investigation of all possible "contaminants" in the old nonanal samples, we identified the active ligand as acetaldehyde. That acetaldehyde is activating CquiOR36 was further confirmed by electroantennogram recordings from antennae of fruit flies engineered to carry CquiOR36. Antennae of female mosquitoes also responded to acetaldehyde. Cage oviposition and dual-choice assays demonstrated that acetaldehyde is an oviposition attractant in a wide range of concentrations and thus of potential practical applications.southern house mosquito | CquiOR36 | acetaldehyde | odorant receptors | EAG

Anecdotes related to preferential mosquito bites are very common, but to date there is no complete explanation as to why one out of two people systematically receives more mosquito bites than the other when both are equally accessible. Here we tested the hypothesis that two constituents of skin emanations, 6-methyl-5-heptan-2-one (6-MHO) and geranylacetone (GA), are natural repellents and may account for differential attraction in different ratios. We studied skin emanations from two human subjects, confirmed in behavioral assays that female southern house mosquitoes are significantly more attracted to subject A (attractant) than to subject N (non-attractant), and tested their 6-MHO/GA ratios in a dual-choice olfactometer. Although repelling at high doses, 6-MHO/GA mixtures were not active at the levels emitted by human skin. We found, however, differential attraction elicited by the aldehydes in the ratios produced by subjects A and N. When tested in a dose commensurate with the level released from human skin and in the ratio produced by subject A, the aldehyde mixture significantly attracted mosquitoes. By contrast, an aldehyde mixture at the same ratio released by subject N did not attract mosquitoes. We, therefore, hypothesized that aldehydes may play a role in the commonly observed differential attraction.

The insect repellent N,N-diethyl-3-methylbenzamide (DEET), is a multimodal compound that acts as a spatial repellent as well as an irritant (contact repellent), thus being perceived by the insect’s olfactory and gustatory systems as an odorant and a tastant, respectively. Soon after DEET was developed, almost 6 decades ago, it was reported that it reduced mosquito feeding on blood mixed with this repellent. It is now known that the mosquito proboscis senses contact repellents with the tips (labella) of the labium, which remain in direct contact with the outer layers of the skin, while the stylets, including the feeding deterrent sensor (labrum), penetrate the skin. We designed a behavioral assay that allowed us to tease apart contact repellency from feeding deterrence. First, we demonstrate here that when DEET was mixed with blood and covered by Parafilm® layers, it did not leak to the outer surface. In our assays, the mean number of landings and duration of contacts with surfaces covering blood mixed with DEET or blood plus solvent (dimethyl sulfoxide) did not differ significantly. The feeding times, however, were significantly different. When blood was mixed either with 0.1 or 1% DEET, female southern house mosquitoes spent significantly less time feeding than the time spent feeding on blood mixed only with the solvent. By contrast, there were no significant differences in the mean times of feeding on blood containing 1% picaridin and blood plus solvent. Like DEET, the contact repellent and insecticide, permethrin, caused a significant reduction in feeding time. We, therefore, concluded, that in this context, DEET and permethrin act as feeding deterrents.

The insect repellent N,N-diethyl-3-methylbenzamide (DEET), is a multimodal compound that acts as a spatial repellent as well as an irritant (contact repellent), thus being perceived by the insect’s olfactory and gustatory systems as an odorant and a tastant, respectively. Soon after DEET was developed, almost 6 decades ago, it was reported that it reduced mosquito feeding on blood mixed with this repellent. It is now known that the mosquito proboscis senses contact repellents with the tips (labella) of the labium, which remain in direct contact with the outer layers of the skin, while the stylets, including the feeding deterrent sensor (labrum), penetrate the skin. We designed a behavioral assay that allowed us to measure feeding deterrence without complications from contact or spatial repellency. Using the southern house mosquito, Culex quinquefasciatus, we demonstrate here that when DEET was mixed with blood and covered by Parafilm® layers, the mean number of landings and duration of contacts with surfaces covering blood mixed with DEET or blood plus solvent (dimethyl sulfoxide) did not differ significantly thus implying that DEET did not leak to the outer surface. The feeding times, however, were significantly different. When blood was mixed either with 0.1 or 1% DEET, female southern house mosquitoes spent significantly (P<0.0001) less time feeding than the time spent feeding on blood mixed only with the solvent. By contrast, significant differences in the mean times of feeding on blood containing 1% picaridin and blood plus solvent were significant at 5%, but not at 1% level. Like DEET, the contact repellent and insecticide, permethrin, caused a significant (P<0.0001) reduction in feeding time. We, therefore, concluded, that in this context, DEET, permethrin, and, to a lesser extent, picaridin, act as feeding deterrents.