Arm-in-cage testing of natural human-derived mosquito repellents.

BACKGROUND: Individual human subjects are differentially attractive to mosquitoes and other biting insects. Previous investigations have demonstrated that this can be attributed partly to enhanced production of natural repellent chemicals by those individuals that attract few mosquitoes in the laboratory. The most important compounds in this respect include three aldehydes, octanal, nonanal and decanal, and two ketones, 6-methyl-5-hepten-2-one and geranylacetone [(E)-6,10-dimethylundeca-5,9-dien-2-one]. In olfactometer trials, these compounds interfered with attraction of mosquitoes to a host and consequently show promise as novel mosquito repellents. METHODS: To test whether these chemicals could provide protection against mosquitoes, laboratory repellency trials were carried out to test the chemicals individually at different concentrations and in different mixtures and ratios with three major disease vectors: Anopheles gambiae, Culex quinquefasciatus and Aedes aegypti. RESULTS: Up to 100% repellency was achieved depending on the type of repellent compound tested, the concentration and the relative composition of the mixture. The greatest effect was observed by mixing together two compounds, 6-methyl-5-hepten-2-one and geranylacetone in a 1:1 ratio. This mixture exceeded the repellency of DEET when presented at low concentrations. The repellent effect of this mixture was maintained over several hours. Altering the ratio of these compounds significantly affected the behavioural response of the mosquitoes, providing evidence for the ability of mosquitoes to detect and respond to specific mixtures and ratios of natural repellent compounds that are associated with host location. CONCLUSION: The optimum mixture of 6-methyl-5-hepten-2-one and geranylacetone was a 1:1 ratio and this provided the most effective protection against all species of mosquito tested. With further improvements in formulation, selected blends of these compounds have the potential to be exploited and developed as human-derived novel repellents for personal protection.

To bite or not to bite! A questionnaire-based survey assessing why some people are bitten more than others by midges.

BACKGROUND: The Scottish biting midge, Culicoides impunctatus, responsible for more than 90% of biting attacks on human beings in Scotland, is known to demonstrate a preference for certain human hosts over others. METHODS: In this study we used a questionnaire-based survey to assess the association between people's perception of how badly they get bitten by midges and their demographic, lifestyle and health related characteristics. RESULTS: Most people (85.8%) reported being bitten sometimes, often or always with only 14.2% reporting never being bitten by midges when in Scotland. There was no association between level of bites received and age, smoking, diet, exercise, medication, eating strongly flavoured foods or alcohol consumption. However, there was a strong association between the probability of being bitten and increasing height (in men) and BMI (in women). A large proportion of participants (33.8%) reported experiencing a bad/severe reaction to midge bites while 53.1% reported a minor reaction and 13.1% no reaction at all. Also, women tend to react more than men to midge bites. Additionally, the results indicated that the susceptibility to being bitten by midges is hereditary. CONCLUSIONS: This study suggests that midges prefer to bite men that are tall and women that have a large BMI, and that the tendency for a child to be bitten or not could be inherited from their parent. The study is questionnaire-based; therefore, the interpretation of the results may be limited by the subjectivity of the answers given by the respondents. Although the results are relevant only to the Scottish biting midge, the approach used here could be useful for investigating human-insect interactions for other insects, particularly those which transmit pathogens that cause disease.

Identification of human-derived volatile chemicals that interfere with attraction of Aedes aegypti mosquitoes.

It is known that human individuals show different levels of attractiveness to mosquitoes. In this study, we investigated the chemical basis for low attractiveness. We recorded behaviors of Aedes aegypti toward the hands of human volunteers and toward the volatile chemicals produced by their bodies. Some individuals, and their corresponding volatiles, elicited low upwind flight, relative attraction, and probing activity. Analyzing the components by gas chromatography coupled to electrophysiological recordings from the antennae of Aedes aegypti, enabled the location of 33 physiologically relevant compounds. The results indicated that higher levels of specific compounds may be responsible for decreased "attractiveness." In behavioral experiments, five of the compounds caused a significant reduction in upwind flight of Aedes aegypti to attractive human hands. Thus, unattractiveness of individuals may result from a repellent, or attractant "masking," mechanism.

Rapid diagnostic PCR assays for members of the Culicoides obsoletus and Culicoides pulicaris species complexes, implicated vectors of bluetongue virus in Europe.

Biting midges of the Culicoides obsoletus Meigen and Culicoides pulicaris L. species complexes (Diptera: Ceratopogonidae) are increasingly implicated as vectors of bluetongue virus in Palearctic regions. However, predicting epidemiological risk and the spread of disease is hampered because whilst vector competence of Culicoides is expressed only in adult females, morphological identification of constituent species is only readily applicable to adult males and some species distinguishing traits have overlapping character states. Furthermore, adult males are typically rare in field collections, making characterisation of Culicoides communities impossible. Here we highlight the utility of mitochondrial cytochrome oxidase subunit I (COI) DNA sequences for taxonomic resolution and species identification of all species within C. obsoletus and C. pulicarus complexes. Culicoides were collected from 18 sites in the UK and Continental Europe, and identified to species level, or species complex level, based on morphological characters. The sample comprised four species from the C. obsoletus complex (n = 88) and five species from the C. pulicaris complex (n = 39). The DNA sequence of the 5' end of the COI gene was obtained from all individuals. Each member species formed a well-supported reciprocally monophyletic clade in a maximum likelihood phylogeny. Levels of DNA sequence divergence were sufficiently high between species to allow the design of species-specific PCR primers that can be used in PCR for identification of members of the C. pulicaris complex or in a multiplex PCR to identify members of the C. obsoletus complex. This approach provides a valuable diagnostic tool for monitoring species composition in mixed field collections of Culicoides.

Identification of a putative azadirachtin-binding complex from Drosophila Kc167 cells.

Cell-proliferation in Drosophila Kc167 cells was inhibited by 50% when cell cultures contained 1.7 x 10(-7) M azadirachtin for 48 h (a tertranortriterpenoid from the neem tree Azadirachta indica). Drosophila Kc167 cells exhibited direct nuclear damage within 6-h exposure to azadirachtin (5 x 10(-7) M and above) or within 24 h when lower concentrations were used (1 x 10(-9) M). Fractionation of an extract of Drosophila Kc167 cells combined with ligand overlay technique resulted in the identification of a putative azadirachtin binding complex. Identification of the members of this complex by Peptide Mass Fingerprinting (PMF) and N-terminal sequencing identified heat shock protein 60 (hsp60) as one of its components.

Laboratory and field responses of the mosquito, Culex quinquefasciatus, to plant-derived Culex spp. oviposition pheromone and the oviposition cue skatole.

Laboratory and field studies were conducted on the oviposition behavior of the pathogen-vectoring mosquito, Culex quinquefasciatus, in response to the oviposition pheromone 6-acetoxy-5-hexadecanolide, produced from a renewable plant resource, Kochia scoparia (Chenopodiaceae) (plant-derived pheromone, PDP), and via an established synthetic route (synthetic oviposition pheromone, SOP). Responses to the oviposition cue skatole (3-methylindole), presented individually and in combination with the plant-derived and synthetic oviposition pheromone, were also studied. Both laboratory and field assays showed that PDP and SOP were equally attractive. Synergistic effects were observed with one combination of PDP and skatole combinations in laboratory assays. Synergy was also observed under field conditions. SOP and skatole combinations showed additive effects in laboratory assays, but were not tested in field bioassays. Although synergism has been previously demonstrated with combinations of SOP and polluted waters, the work presented here is the first example of synergy between a specific oviposition attractant and the oviposition pheromone. Furthermore, the efficacy of mosquito pheromone produced from a cheap, renewable botanical source has been demonstrated.

Role of semiochemicals in mate location by parasitic sea louse, Lepeophtheirus salmonis.

The role of olfaction and diffusible pheromones in mate location behavior of sea lice, Lepeophtheirus salmonis, was assessed with Y-tube behavioral bioassays. The pheromone "emitting" animals were located in a chamber in one arm of a Y-tube arena, with artificial seawater flowing through both arms. Adult male sea lice displayed both activation and directional responses to seawater conditioned with preadult II virgin females, but were only activated by mated adult female conditioned water. Further, when males were given the choice of preadult II virgin females or mated adult females, a significant number of males chose the arm with the preadult II virgin females. Adult males showed activation responses when presented with water conditioned with adult males but were not attracted to them. When presented with adult males, preadult II virgin females showed only directional responses, but not activation responses. Preadult II virgin female conditioned water was extracted using solid-phase extraction (SPE) protocols pioneered for semiochemical isolation. Adult male sea lice showed significant directional responses to the preadult II virgin female SPE extract. Distillation under vacuum was performed on the extract to give a distillate comprising components with a molecular weight range and physical properties comparable to those of compounds utilized as volatile semiochemicals by terrestrial organisms and a residue comprising components with higher molecular weight range comparable to those utilized as involatile semiochemicals. Adult males were found to be both significantly activated and attracted to the distillate, but not to the residue. This research provides evidence that small, lipophilic organic molecules are used by sea lice as sex pheromone signals to locate a member of the opposite sex.