SUMMARY Female mosquitoes that transmit deadly diseases locate human hosts by detecting exhaled CO2 and skin odor. The identities of olfactory neurons and receptors required for attraction to skin odor remain a mystery. Here we show that the CO2-sensitive olfactory neuron is also a sensitive detector of human skin odorants in both Aedes aegypti and Anopheles gambiae. We demonstrate that activity of this neuron is important for attraction to skin odor, establishing it as a key target for intervention. We screen ~0.5 million compounds in silico and identify several CO2-receptor ligands, including an antagonist that reduces attraction to skin and an agonist that lures mosquitoes to traps as effectively as CO2. Analysis of the CO2 receptor ligand space provides a foundation for understanding mosquito host-seeking behavior and identifies odors that are potentially safe, pleasant, and affordable for use in a new generation of mosquito control strategies worldwide.

Summary CO2 present in exhaled air is the most important sensory cue for female blood-feeding mosquitoes, causing activation of long-distance host-seeking flight, navigation towards the vertebrate host1, and, in the case of Aedes aegypti, increased sensitivity to skin odours2. The CO2 detection machinery is therefore an ideal target to disrupt host seeking. We use electrophysiological assays to identify a volatile odourant that causes an unusual, ultra-prolonged activation of CO2-detecting neurons in three major disease-transmitting mosquitoes: Anopheles gambiae, Culex quinquefasciatus and A. aegypti. Importantly ultra-prolonged activation of this neuron severely compromises its ability to subsequently detect CO2 for several minutes. We also identify odours that strongly inhibit the CO2-sensitive neuron as candidates for use in disruption of host-seeking behaviour, as well as an odour that evokes CO2-like activity and thus has potential use as a lure in trapping devices. Analysis of responses to panels of structurally related odours across the three mosquitoes and Drosophila, which have related CO2-receptor proteins, reveals a pattern of inhibition that is often conserved. We use video tracking in wind-tunnel experiments to demonstrate that the novel ultra-prolonged activators can completely disrupt CO2-mediated activation as well as source-finding behaviour in Aedes mosquitoes, even after the odour is no longer present. Finally, semi-field studies demonstrate that use of ultra-prolonged activators disrupts CO2-mediated hut entry behaviour of Culex mosquitoes. The three classes of CO2 response modifying odours offer powerful instruments for developing new generations of insect repellents and lures, which even in small quantities can interfere with the ability of mosquitoes to seek humans.

Summary There are major impediments to finding improved DEET alternatives because the receptors causing olfactory repellency are unknown, and new chemistries require exorbitant costs to determine safety for human use. Here we identify DEET-sensitive neurons in a pit-like structure in the Drosophila antenna called the sacculus. They express a highly conserved receptor Ir40a and flies in which these neurons are silenced or Ir40a is knocked down lose avoidance to DEET. We use cheminformatics to screen >400,000 compounds and identify >100 natural compounds as candidate repellents. We test several and find that most activate Ir40a+ neurons and are repellents for Drosophila. These compounds are strong repellents in mosquitoes as well. The candidates contain chemicals that do not dissolve plastic, are affordable, smell mildly like grapes, with three being considered safe for human consumption. Our findings pave the way to discover new generations of repellents that will help fight deadly insect-borne diseases worldwide.

Abstract. Laboratory studies were carried out to investigate the role of larval habitatderived microorganisms in the production of semiochemicals for oviposition site selection by Anopheles gambiae Giles sensu stricto mosquitoes. Dual-choice bioassays with gravid females were conducted in standard mosquito cages. Field-collected or laboratory-reared mosquitoes, individually or in groups, were offered a choice between unmodified (water or soil from a natural breeding site) or modified substrates (filtered water, autoclaved soil or sterile media to which bacterial suspensions had been added). Egg counts were used to assess oviposition preferences.Mosquitoes preferred to oviposit on unmodified substrates from natural larval habitats containing live microorganisms rather than on sterilized ones. Variable responses were observed when sterile substrates were inoculated with bacteria isolated from water and soil from natural habitats.We conclude that microbial populations in breeding sites produce volatiles that serve as semiochemicals for gravid An. gambiae. These signals, in conjunction with other (non-olfactory) chemical and physical cues, may be used by the female to assess the suitability of potential larval habitats in order to maximize the fitness of her offspring.Key words: oviposition site selection, soil microbiota, semiochemicals, Anopheles gambiae Résumé. Des études de laboratoire ont été conduites afin de déterminer le rô le de substances chimiques attractives de microorganismes présents dans les sites larvaires dans la préférence de ponte des femelles gravides d'Anopheles gambiae s.s. pour ces sites. Un test biologique à double choix a été mis en place dans des cages à moustiques standards contenant des femelles gravides. Un choix entre substrat naturel (eau et sol non modifiés provenant des sites larvaires naturels) et substrat modifié (eau filtrée, sol stérilisé, ajout de suspensions bactériennes) est offert individuellement ou en groupe aux femelles collectées sur le terrain ou élevées en laboratoire. Le nombre d'oeufs pondus dans chaque substrat a été utilisé comme critère pour évaluer la préférence des femelles gravides pour un substrat particulier. Les femelles ont préféré pondre dans le substrat naturel contenant des microorganismes vivants plutô t que dans le substrat stérile. Des réponses variables ont été observées lorsque des bactéries provenant des sites larvaires naturels ont été inoculées dans le substrat stérile. Nous concluons que la population microbienne, présente dans les site larvaires, produit des substances chimiques odorantes attirant les femelles gravides d'Anopheles gambiae s.s. Ces signaux, associés à d'autres substances non odorantes et à des facteurs physiques, peuvent être utilisés par les femelles pour évaluer le potentiel d'un site larvaire, en vue du développement optimal de leur progéniture.

Highlights d Diurnal/nocturnal mosquitoes have distinct circadian neural circuit and PER cycling d Diurnal versus nocturnal mosquitoes have distinct clockmodulated light preferences d Light preference is dependent on mosquito sex and species, time of day, and color d Circadian clock modulates timing and valence of mosquito light responses

There is an increased need for improved and affordable insect repellents to reduce transmission of rapidly spreading diseases with high mortality rates. Natural products are often used when DEET cannot be afforded or accessed and when consumers choose not to use a synthetic repellent. The essential oils from two newly bred Nepeta cataria (catnip) plants representing two different chemotypes and their respective isolated nepetalactone isomers were evaluated as mosquito repellents against Aedes aegypti mosquitoes that transmit the Zika and Dengue virus in a one choice landing rate inhibition assay. A dose response curve was generated for each treatment and a time course analysis of repellency was performed over 24 hours with a N. cataria essential oil sample. The results indicate that all essential oil samples and their respective purified nepetalactone isomers were able to achieve greater than 95% repellency. Between two and four hours, the ability to repel more than 95% of the mosquitoes diminished. At the lowest concentrations tested, the nepetalactones and crude essential oil samples were more effective than DEET at reducing the number of mosquito landings.

Natural anopheline populations exhibit much variation in ability to support malaria parasite development, but the genetic mechanisms underlying this variation are not clear. Previous studies in Mali, West Africa, identified two quantitative trait loci (QTL) in Anopheles gambiae mosquitoes that confer refractoriness (failure of oocyst development in mosquito midguts) to natural Plasmodium falciparum parasites. We hypothesize that new QTL may be involved in mosquito refractoriness to malaria parasites and that the frequency of natural refractoriness genotypes may be higher in the basin region of Lake Victoria, East Africa, where malaria transmission intensity and parasite genetic diversity are among the highest in the world. Using field-derived F 2 isofemale families and microsatellite marker genotyping, two loci significantly affecting oocyst density were identified: one on chromosome 2 between markers AG2H135 and AG2H603 and the second on chromosome 3 near marker AG3H93. The first locus was detected in three of the five isofemale families studied and colocalized to the same region as Pen3 and pfin1 described in other studies. The second locus was detected in two of the five isofemale families, and it appears to be a new QTL. QTL on chromosome 2 showed significant additive effects while those on chromosome 3 exhibited significant dominant effects. Identification of P. falciparum-refractoriness QTL in natural An. gambiae mosquitoes is critical to the identification of the genes involved in malaria parasite transmission in nature and for understanding the coevolution between malaria parasites and mosquito vectors.

BackgroundThe Asian Citrus Psyllid (ACP), Diaphorina citri, can transmit the bacterium Candidatus Liberibacter while feeding on citrus flush shoots. This bacterium causes Huanglongbing (HLB), a major disease of citrus cultivation worldwide necessitating the development of new tools for ACP surveillance and control. The olfactory system of ACP is sensitive to variety of odorants released by citrus plants and offers an opportunity to develop new attractants and repellents.ResultsIn this study, we performed single-unit electrophysiology to identify odorants that are strong activators, inhibitors, and prolonged activators of ACP odorant receptor neurons (ORNs). We identified a suite of odorants that activated the ORNs with high specificity and sensitivity, which may be useful in eliciting behavior such as attraction. In separate experiments, we also identified odorants that evoked prolonged ORN responses and antagonistic odorants able to suppress neuronal responses to activators, both of which can be useful in lowering attraction to hosts. In field trials, we tested the electrophysiologically identified activating odorants and identified a 3-odor blend that enhances trap catches by ∼230%.ConclusionThese findings provide a set of odorants that can be used to develop affordable and safe odor-based surveillance and masking strategies for this dangerous pest insect.