Our results describe a complete and highly concordant map of both the molecular and cellular olfactory components on the maxillary palp of the adult female An. gambiae mosquito. These results also facilitate the understanding of how An. gambiae mosquitoes sense olfactory cues that might be exploited to compromise their ability to transmit malaria.

Host odours play a major role in the orientation and host location of blood-feeding mosquitoes. Anopheles gambiae Giles sensu stricto, which is the most important malaria vector in Africa, is a highly anthropophilic mosquito species, and the host-seeking behaviour of the females of this mosquito is guided by volatiles of human origin. Ammonia, lactic acid and several carboxylic acids are known to be present in the human odour blend. We investigated the effect of these compounds on naive female mosquitoes using a dual-port olfactometer. Ammonia was an attractant on its own, whereas lactic acid was not attractive. Carboxylic acids, offered as a mixture of 12 compounds, were repellent at the concentration tested. The addition of ammonia to the carboxylic acid mixture overruled the repellent effect of the latter. Combining ammonia with either lactic acid or the carboxylic acids did not enhance the attractiveness of ammonia alone. However, a synergistic effect was found when ammonia, lactic acid and the carboxylic acids were applied as a blend. Our findings indicate that An. gambiae s.s. relies on the combination of ammonia, lactic acid and carboxylic acids in its orientation to human hosts. The role of lactic acid in this tripartite synergism differs from that reported for the yellow fever mosquito Aedes aegypti.

The African malaria mosquito Anopheles gambiae sensu stricto continues to play an important role in malaria transmission, which is aggravated by its high degree of anthropophily, making it among the foremost vectors of this disease. In the current study we set out to unravel the strong association between this mosquito species and human beings, as it is determined by odorant cues derived from the human skin. Microbial communities on the skin play key roles in the production of human body odour. We demonstrate that the composition of the skin microbiota affects the degree of attractiveness of human beings to this mosquito species. Bacterial plate counts and 16S rRNA sequencing revealed that individuals that are highly attractive to An. gambiae s.s. have a significantly higher abundance, but lower diversity of bacteria on their skin than individuals that are poorly attractive. Bacterial genera that are correlated with the relative degree of attractiveness to mosquitoes were identified. The discovery of the connection between skin microbial populations and attractiveness to mosquitoes may lead to the development of new mosquito attractants and personalized methods for protection against vectors of malaria and other infectious diseases.

Differences between human individuals in their attractiveness to female mosquitoes have been reported repeatedly, but the underlying mechanisms are not well understood. Skin emanations from 27 human individuals, collected on glass marbles, were tested against ammonia in a dual-choice olfactometer to establish their degrees of attractiveness to anthropophilic Anopheles gambiae s.s. Giles (Diptera: Culicidae) mosquitoes. Ammonia was used as a standard odour source because of its proven attractiveness to An. gambiae s.s. Skin emanations from most volunteers attracted significantly more mosquitoes than ammonia. There were clear differences in the attractiveness of skin emanations from different volunteers relative to that of ammonia, as well as in the strength of the trap entry response. Consistent differences were observed when emanations from the three most and the three least attractive volunteers were tested pairwise. No gender or age effect was found for relative attractiveness or trap entry response. Emanations from volunteers with higher behavioural attractiveness elicited higher electroantennogram response amplitudes in two pairs, but in a third pair a higher electroantennogram response was found for the less attractive volunteer. These results confirm that odour contributes to the differences in attractiveness of humans to mosquitoes.

Olfactory receptor neurons (ORNs) in the antenna of insects serve to encode odors in action potential activity conducted to the olfactory lobe of the deuterocerebrum. We performed an analysis of the electrophysiological responses of olfactory neurons in the antennae of the female malaria mosquito Anopheles gambiae s.s. and investigated the effect of blood feeding on responsiveness. Forty-four chemicals that are known to be present in human volatile emanations were used as odor stimuli. We identified 6 functional types of trichoid sensilla and 5 functional types of grooved-peg sensilla (GP) based on a hierarchical cluster analysis. Generalist ORNs, tuned to a broad range of odors, moderate specialist ORNs and 2 ORNs tuned to only one odor were identified in different sensilla types. Neurons in GP were tuned to more polar compounds including the important behavioral attractant ammonia and its synergist L-lactic acid, responses to which were found only in GP. Combinatorial coding is the most plausible principle operating in the olfactory system of this mosquito species. We document for the first time both up- and downregulation of ORN responsiveness after blood feeding. Modulation of host-seeking and oviposition behavior is associated with both qualitative and quantitative changes in the peripheral sensory system.

The role of aliphatic carboxylic acids in hostseeking response of the malaria mosquito Anopheles gambiae sensu stricto was examined both in a dual-choice olfactometer and with indoor traps. A basic attractive blend of ammonia + lactic acid served as internal standard odor. Single carboxylic acids were tested in a tripartite blend with ammonia + lactic acid. Four different airflow stream rates (0.5, 5, 50, and 100 ml/min) carrying the compounds were tested for their effect on trap entry response in the olfactometer. In the olfactometer, propanoic acid, butanoic acid, 3-methylbutanoic acid, pentanoic acid, heptanoic acid, octanoic acid, and tetradecanoic acid increased attraction relative to the basic blend. While several carboxylic acids were attractive only at one or two flow rates, tetradecanoic acid was attractive at all flow rates tested. Heptanoic acid was attractive at the lowest flow rate (0.5 ml/min), but repellent at 5 and 50 ml/min. Mixing the air stream laden with these 7 carboxylic acids together with the headspace of the basic blend increased attraction in two quantitative compositions. Subtraction of single acids from the most attractive blend revealed that 3-methylbutanoic acid had a negative effect on trap entry response. In the absence of tetradecanoic acid, the blend was repellent. In assays with MM-X traps, both a blend of 7 carboxylic acids + ammonia + lactic acid (all applied from low density polyethylene-sachets) and a simple blend of ammonia + lactic acid + tetradecanoic acid were attractive. The results show that carboxylic acids play an essential role in the host-seeking behavior of An. gambiae, and that the contribution to blend attractiveness depends on the specific compound studied.

Heat sensitivity is a sensory modality that plays a critical role in close-range host-seeking behaviors of adult female Anopheles gambiae, the principal Afrotropical vector for human malaria. An essential step in this activity is the ability to discriminate and respond to increases in environmental temperature gradients through the process of peripheral thermoreception. Here, we report on the characterization of the anopheline homolog of the transient receptor potential (TRP) A1/ANKTM1 channel that is consistent with its role as a heat-sensor in host-seeking adult female mosquitoes. We identify a set of distal antennal sensory structures that specifically respond to temperature gradients and express AgTRPA1. Functional characterization of AgTRPA1 in Xenopus oocytes supports its role in the molecular transduction of temperature gradients in An. gambiae, providing a basis for targeting mosquito heat responses as a means toward reducing malaria transmission.

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