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United States Department of Agriculture

Agricultural Research Service


Location: Mosquito and Fly Research

2012 Annual Report

1a. Objectives (from AD-416):
1. Discover and evaluate new toxicants and biological control agents for control of biting Nematocera. 2. Develop new application methods for pesticides to biting Nematocera that minimize environmental exposure and that optimize lethal or repellent effect, including presentation on clothing, aerosol application in or outdoors, residual application, disinsection of aircraft, and delivery of spatially repellent compounds. 3. Conceive and test new methods of managing vector and pest populations through the use of behavior-altering chemicals, including repellents, attractants, and inhibitors. 4. Examine the parameters of behavioral bioassay methods that influence practical comparisons of personal protection products, with a view to determining those elements of commercial testing that influence reliability of results. 5. Determine chemical and other cues associated with regulation and orientation of specific behaviors by Aedes albopictus, Culex pipiens group species/hybrids, and other biting arthropods that can be applied to the solution of operational surveillance and control problems. 6. Discover and characterize environmental predictors of the distribution of mosquitoes in order to assess the risk of invasive species and pathogen transmission. Apply to the development of methods and techniques to accurately assess mosquito population density, to deploy vector surveillance systems, and to detect exotic invasive species.

1b. Approach (from AD-416):
1. High throughput bioassays will be used to screen toxicants from synthetic compound and natural product libraries. New formulations of bacterial toxins, baculoviruses, and biorationals will be evaluated in standard assays. Molecular methods (RNAi and dsRNA) will be used to identify targets for control and resistance management. Improved formulation and delivery of dsRNA will be developed to penetrate the mosquito cuticle. 2. Factors affecting measurement of bite protection by permethrin-treated clothing will be examined. Alternative repellents applied to military uniforms and improved binding of repellents to fabric will be researched, followed by laboratory validation of factory-treated fabric and semi-field studies. Aerosol application of control compounds, formulations, equipment, application techniques, and strategies will be explored. Existing and novel chemical compounds, formulations, equipment, impregnated materials, application techniques, and strategies for barrier applications will be evaluated under laboratory, semi-field and field conditions. Factors needed to achieve optimal control will be identified. Air curtains will be designed and evaluated in simulated aircraft fuselages. 3. Comparison of pathogen-infected and uninfected mosquito responses to behavior-altering chemicals will be examined. Baited commercial traps will be used as surrogates for human and livestock hosts. The manner in which compounds affect host-seeking behavior will be quantified. Catches in control and attractant-baited traps located near devices releasing spatial repellents or inhibitors will be compared. 4. The effect of mosquito fatigue on repellent protection time will be evaluated with female Aedes aegypti and a study comparing methods for measuring repellent efficacy will be conducted. Laboratory results of repellent efficacy will be compared to products tested in the semi-field environment. 5. Factors that result in positive resting site selection responses by adult mosquitoes, volatile chemical cues associated with mate location, and cues used for location and utilization of sugar and nectar sources will be determined. Plant-derived compounds that attract mosquitoes will be evaluated in the field. Factors that influence host-finding behavior based on host odors and olfactory cues will be used to enhance traps. Novel strategies for mosquito surveillance that utilize oviposition site cues will be devised. New chemical attractants will be identified and evaluated in the field using behavioral and chemical analyses. The response to toxicant exposure on arthropod behavior will be evaluated to improve toxicant impact. 6. Methods and techniques to assess mosquito population density will be developed. Environmental predictors of mosquito distribution will be characterized to provide improved strategies for assessment of mosquito populations. Discover ecologic and climatic factors to assess population densities of vector mosquitoes in the US and, using Rift Valley fever, evaluate the risk of exotic species and disease introduction into the U.S.

3. Progress Report:
Biting Nematocera transmit pathogens that cause diseases such as West Nile virus, dengue, malaria, leishmaniasis and filariasis to humans and animals in urban, suburban, and rural areas and in agricultural, recreational, and military environments. This research project is focused on improving the control of biting Nematocera through a better understanding of their biology and the development of novel products, technologies, and control strategies. New toxicants based on double-stranded RNA were discovered and patented. Studies showed that these highly specific toxicants can be delivered to mosquitoes in sugar-baited traps. The U.S. Army and U.S. Marine Corps are developing new uniform fabrics to prevent mosquitoes from biting through the fabric. The new type III U.S. Army fire resistant uniform contains a factory level permethrin finish that provides high protection from mosquito bites throughout the lifetime of the uniform. The U.S. Marine Corps has explored nonwoven fabrics that are nearly impenetrable to the bites of mosquitoes. A seminal study on the effects of ultra low volume (ULV) pesticides on Old World sand flies was completed. The results indicate that adulticiding combined with the use of permethrin-treated nets and clothing significantly reduced fly-human contact and therefore minimizes transmission of leishmaniasis to deployed military troops. An aircraft disinsection strategy that employs an air curtain rather than insecticides has been developed with the U.S. Department of Transportation and demonstrated to a major U.S. airline. Spatial repellents from plant sources were tested in a semi-field environment and found to reduce host finding by mosquitoes. Nearly 400 compounds were tested as candidate repellents. One compound was found to be over two times as potent as the standard N,N-diethyl-3-benzamide (DEET) repellent. The behaviors associated with mosquitoes finding resting sites were studied to improve techniques employed for the surveillance of mosquitoes. The attraction of the Asian Tiger mosquito to odors from over 40 plants was studied. Odors from butterfly bush and goldenrod were found to be the most attractive for this species. Factors that influence the ability of mosquitoes to locate traps were studied and it was determined that species specific trapping is achieved by manipulations of these cues. Research that compared trap catch with landing rates of mosquitoes on humans has produced an algorithm to improve the accuracy of assessing mosquito density in an area. A preliminary Geographic Information Systems (GIS)-based model has been developed for optimal trap deployment for surveillance of mosquitoes in a local area. Research was conducted that identified a correlation between years that El Niño effects are present, and the risk for outbreaks of Rift Valley fever and chikungunya virus in humans.

4. Accomplishments
1. Discovered new toxicants for mosquitoes. Disease causing organisms vectored by mosquitoes infect over 300 million people and large numbers of animals worldwide resulting in debilitating effects and often death. Researchers at Agricultural Research Service (ARS) in Gainesville, FL have been issued a patent entitled, “Pesticidal double stranded RNA composition and method of use thereof.” This technology describes a novel means of Aedes aegypti control by using double stranded RNA constructs to silence genes that encode inhibitor of apoptosis (IAP) proteins, which are critical to survival of these mosquitoes. Collaboration with a Cooperative Research and Development Agreement (CRADA) partner has resulted in development of dsRNA constructs for mosquito control. These highly specific (Hi-SPeC) compounds have been successfully delivered to adult mosquitoes via sugar baited traps and determined to knock-out the targeted protein. This approach can compliment current vector control strategies such as insecticide treated bednets (ITNs) and indoor residual spraying (IRS) programs to control important vectors of malaria and arboviruses to man and animals.

2. Discovered a link between El Niño events and incidence of outbreaks of mosquito-borne diseases. Researchers at ARS in Gainesville, FL discovered that extremes in climate conditions forced by the El Niño/Southern Oscillation (ENSO) lead to severe droughts or floods, ideal conditions for disease vectors to emerge, and resulted in epizootics and epidemics of 2 vector-borne diseases of humans and animals: Rift Valley fever and chikungunya. The results were reported in PLOS Neglected Tropical Diseases that as the frequency and severity of extremes in climate increase the potential for globalization of vectors and disease is likely to accelerate. The results of this work can be used to better protect humans and animals, on a global scale, from outbreaks of mosquito-borne illnesses.

3. Estimated accuracy of trap catches with mosquitoes that landed on humans. Scientists at ARS in Gainesville, FL, compared the number of female mosquitoes captured by suction traps, portable light traps (with CO2) and landing on a human subject. The results of this research showed that mosquito density from light trap samples was underestimated by 43-97% and over-estimated by 80-85% (depending on species), when compared with the mosquito landing rate on a human subject. Corresponding values for suction trap samples were 28-88% and 17-87%. A statistical algorithm that corrects mechanical trap-based estimates of adult mosquito density to the landing rate of mosquitoes on a human host was constructed. Use of this algorithm will provide a better estimate of mosquito density in a local area which in turn can be used to help mosquito control professionals determine the best method of mosquito control.

4. Completed seminal study on the efficacy of ultralow volume (ULV) pesticides on Old World sand fly species. Old world sand flies are responsible for the transmission of leishmania to humans in areas where U.S. military personnel are currently deployed. Researchers at ARS in Gainesville, FL successfully conducted the first study looking at the efficacy of ULV pesticide applications on Old World sand fly species in Leishmaniasis endemic regions in a natural setting in Africa. These findings, reported in the Journal of Medical Entomology, indicate that by combining ULV treatments of natural sand fly populations with treatment of camouflage netting, permethrin-treated clothing and the use of N,N-diethyl-3-methylbenzamide (DEET) could substantially minimize opportunities for human-sand fly contact and thus minimize opportunities for transmission of Leishmania to deployed troops. The outcome of this research leads to disease risk reduction to U.S. military personnel.

5. Demonstrated air curtain(s) to end users in a field situation and got feedback for product acceptance. Mosquitoes and flies can be frequent travelers on commercial aircraft and once on board the approved control method for these disease-carrying insects is use of residual or aerosol pesticides. Because of the general dislike of pesticides by the public and reports of passenger discomfort and death after aerosol applications, the U.S. Department of Transportation initiated cooperative research with ARS in Gainesville, FL to investigate non-pesticide methods for insect disinsection of aircraft. A developed system was evaluated in the field in cooperation with the U.S. Department of Transportation (DOT) and Delta Airlines in Accra, Ghana. Relatively high winds negated mosquito flight activity, but system acceptance by crew members and passengers was overwhelmingly positive.

6. Discovered novel mosquito repellents. Bites received from mosquitoes are often a nuisance, but can lead to transmission of disease pathogens from insect saliva injected into humans and other animals during the biting process. Discovery of new, highly potent and universal mosquito repellents is necessary because of concerns over the use of current mosquito repellents coupled with the problem that repellents vary in efficacy against different mosquito species. In conjunction with collaborators inside and outside of ARS, over 380 candidate repellents were evaluated at ARS in Gainesville, FL. Included in these, scientists at ARS Gainesville, FL, synthesized and evaluated novel fluorinated carboxamides as repellents and later as insecticides and fungicides. Out of 14 novel compounds, the best repellent was efficacious against Ae. aegypti mosquitoes at a concentration 2.3 times lower than that of the standard N,N-diethyl-3-methylbenzamide (DEET). A manuscript covering these findings has been drafted. The results of this research may lead to new products used to protect U.S. military and civilians from the bites of mosquitoes.

7. Discovered odor cues used by mosquitoes to locate nectar/sugar sources. Mosquitoes need sugar/nectar for survival. Plant odors alert mosquitoes to the location of sugar and nectar meals. A study on the phenology of flowering plants conducted by ARS scientists in Gainesville, FL on plants in north central Florida indicated that flowering plants were abundant during all seasons. Forty different species were collected and evaluated in dual-choice olfactometer studies. Intact flowers, solvent extracts and some identified volatile compounds from the butterfly bush and goldenrod were found to be very attractive to the invasive species, the Asian Tiger mosquito (Aedes albopictus). This information will be useful in the development of species-specific traps/targets for the population management of this mosquito species.

8. Managed vector and pest populations through the use of behavior altering spatial repellents. Mosquitoes and other pests vector disease to humans and livestock through transmission of disease agents. Therefore, approaches that minimize host-vector contact can provide an alternative strategy to reduce disease risk. Semi-field studies conducted by ARS researchers in Gainesville, FL, were conducted with various natural plant produced compounds to determine their ability to deter pest mosquito species from locating potential hosts. Several compounds were identified that achieve this objective. Additional studies will focus on optimization of release rates and development of delivery technology to use these compounds. The results of this work provide an alternative control technology for use against mosquitoes and other biting arthropods.

9. Determined mosquito resting site preferences. The resting behavior of mosquitoes and species composition in local areas of Florida are not well understood. Scientists at ARS in Gainesville, FL, inspected adult mosquito resting site structures in a north Florida swamp which revealed six species of mosquitoes in four genera. The faunal composition and density of mosquitoes varied significantly within and between resting structure types. Additional methods-development is necessary to standardize conventional mechanical aspiration-based equipment for sampling resting adult mosquito populations. In screened cage studies, resting adult mosquitoes showed cyclical entry and exit responses to 30 cm cubic "red boxes" that correlated with, respectively, changing light intensity and resting surface temperature. Other predictors of entry/exit activity were moisture content of the resting site substrate and compass direction of the box opening. Results from this research will lead to improved methods for surveillance of adult mosquitoes.

10. Optimization of insect traps based on factors that influence host-finding behavior. The control of mosquitoes and other biting flies in a local area is often dependent on the surveillance data. Efficient traps are needed to provide accurate surveillance data to provide the best information for control strategies. Mosquitoes utilize visual, physical and different combination of heat patterns, color patterns, chemical odor blends and delivery rates to locate the surrogate host (cylindrical traps). Scientists at ARS in Gainesville, FL, studied the interaction of various cues and the species composition that were trapped after modification of the various cues. These data indicate that species-specific traps can be developed if these factors are optimized for each species. Species specific traps will aid pest management professionals in determining the threat of disease in an area and the choice of control strategy to mitigate this disease risk.

11. Developed initial model for trap deployment to improve mosquito surveillance. Scientists at ARS in Gainesville, FL, sampled open, ecotone, and wooded habitats in a north Florida swamp for adult mosquitoes using light traps (with CO2) and suction traps. Mosquito capture responses were related to the physical and biological characteristics of each habitat type and a preliminary Geographic Information Systems (GIS)-based model for deployment of adult mosquito traps in a vector surveillance program developed. Ongoing methods development research is being conducted to determine the effect of trap type and location parameters, microclimatic factors, and semiochemical point source factors on the reliability of estimates of adult mosquito density by each sampling method. Improved assessment of the local density of adult mosquitoes can be used by mosquito control professionals for effective adulticiding of mosquitoes.

12. Evaluated mosquito bite protection ability of new U.S. Military uniforms. The U.S. Army and U.S Marine Corps factory treat all combat uniforms with permethrin to prevent mosquitoes from biting through the fabric. The treatment incorporates a binder that enhances permethrin retention throughout the lifetime of the uniform. The balance between binder and permethrin can only be assessed by bite protection evaluation of specimens of fabric from treated uniforms. Researchers at ARS in Gainesville, FL have completed optimization of the permethrin finish that prevents mosquito bites through the U.S. Army Fire Resistant Operation Enduring Freedom Camouflage (FROC) uniform. This uniform is now constructed of Fire Resistant Army Combat Uniform (FRACU) Type III fabric. This fabric construction is more durable than the Type I uniform fabric. The bite prevention of uniforms constructed with Type III fabric are superior to the protection from treated Type I fabric. The U.S. Marine Corps requested bite prevention evaluation of experimental nonwoven fabrics, this type of fabric is a candidate for the development of future US Marine Corps uniforms. It was demonstrated that nonwoven fabrics provide superior protection against mosquito bites compared to woven fabrics. The results of this research are used to guide the U.S. military in providing personnel with new uniforms that are optimized for prevention of bites from mosquitoes and other arthropods that spread disease.

13. Evaluation of host compounds for tick attraction. A series of chemicals previously identified from common tick hosts, were evaluated in the laboratory over a range of concentrations against brown dog ticks (Rhipicephalus sanguineus) in a Y-tube olfactometer. Several compounds were found to consistently activate and attract ticks. These compounds will provide the basis for development of a surveillance trap as well as an attract-and-kill device for ticks in residences or kennels where these ticks are pests. This approach should result in reduced pesticide use for these common residential and kennel pests.

Review Publications
Britch, S.C., Linthicum, K., Walker, T.W., Farooq, M., Gordon, S.W., Clark, J.W., Ngere, F., N'Gonga, D., Chepchieng, C. 2011. Evaluation of ULV applications against Old World sand fly (Diptera: Psychodidae) species in equatorial Kenya. Journal of Medical Entomology. 48(6):1145-1159.

Xue, R., Zhao, T., Muller, G.C., Linthicum, K., Linser, P., Moeller, J. 2012. 2nd International Forum for Surveillance and Control of Mosquitoes and Mosquito-borne Diseases. Wing Beats. 23(1):16-21.

Cohnstaedt, L.W., Allan, S.A. 2011. Effects of sublethal pyrethroid exposure on host-seeking behavior of female mosquitoes. Journal of Medical Entomology. 36(2):1-9.

Kovendan, K., Murugan, K., Vincent, S., Barnard, D.R. 2011. Studies on larvicidal and pupicidal activity of Leucas aspera Willd. (Lamiaceae) and bacterial insecticide, Bacillus sphaericus, against malarial vector, Anopheles stephensi Liston (Diptera: Culicidae). Parasitology Research. 10:1-6.

Aldridge, R.L., Wynn, W.W., Britch, S.C., Linthicum, K. 2012. Aspirator Gun for High-Throughput Mosquito Bioassays. Journal of the American Mosquito Control Association. 28(1):65-68.

Sims, K., Becnel, J.J., Funderburk, J. 2012. The morphology and biology of the entomophilic Thripinema fuscum (Tylenchida: Allantonematidae), and the histopathologic effects of parasitism on the host Frankliniella fusca (Thysanoptera: Thripidae). Journal of Natural History. 46(17-18):1111-1128.

Anyamba, A., Linthicum, K., Small, J., Collins, K., Tucker, C.J., Pak, E.P., Britch, S.C., Eastman, J.R., Pinzon, J.E., Russell, K. 2012. Climate teleconnections and recent patterns of human and animal disease outbreaks. PLOS Neglected Tropical Diseases. 6(1):1-14.

Anderson, J.F., Ferrandino, F.J., Dingman, D.W., Main, A.J., Andreadis, T.G., Becnel, J.J. 2011. Control of mosquitoes in catch basins in Connecticut with Bacillus thuringiensis israelensis, Bacillus sphearicus, and Spinosad. Journal of the American Mosquito Control Association. 27(1):45-55.

Becnel, J.J., Wei Pridgeon, Y. 2011. Susceptibility of Mosquito and Lepidopteran Cell Lines to the Mosquito Iridescent Virus (IIV-3) from Aedes taeniorhynchus. Journal of Invertebrate Pathology. 108:40-45. DOI: 10.1016/j.jip.2011.06.008.

Britch, S.C., Linthicum, K., Wynn, W.W., Aldridge, R.L., Walker, T.W., Farooq, M., Dunford, J.C., Smith, V.L., Robinson, C.A., Lothrop, B.B., Snelling, M., Gutierrez, A., Wittie, J., White, G. 2011. Longevity and efficacy of bifenthrin treatment on desert-pattern U.S. military camouflage netting against mosquitoes in a hot-arid environment. Journal of the American Mosquito Control Association. 27(3):272-279.

Clark, G.G., Rubio-Palis, Y. 2011. Mosquito vector biology and control in Latin America - A 21st symposium. Journal of the American Mosquito Control Association. 27(3):280-299.

Tucker, C.J., Yager, K., Anyamba, A., Linthicum, K. 2011. Climate Change, Extreme Weather Events, and Fungal Disease Emergence and Spread. Institute of Medicine. Fungal Diseases: An Emerging Threat to Human, Animal, and Plant Health. Washington, DC: The National Academies Press. 324-342.

Romoser, W.S., Oveido, M., Lerdthusnee, K., Patrican, L.A., Turell, M.J., Dohm, D.J., Linthicum, K., Bailey, C. 2011. Rift Valley fever virus-infected mosquito ova and associated pathology: possible implications for endemic maintenance. Journal of Research and Reports in Tropical Medicine. 2:121-127.

Murugan, K., Kovendan, K., Vincent, S., Barnard, D.R. 2012. Biolarvicidal and pupicidal activity of Acalypha alnifolia Klein ex Willd.(Family:Euphorbiaceae) leaf extract and microbial insecticide, Metarhizium anisopliae(Metsch.)against malaria fever mosquito Anopheles stephensi Liston. Parasitology Research. 110:263-270.

Sobolev, V.S., Khan, S.I., Tabanca, N., Wedge, D.E., Manly, S.P., Cutler, S.J., Coy, M.R., Becnel, J.J., Neff, S.A., Gloer, J.B. 2011. Biological Activity of Peanut (Arachis hypogaea) Phytoalexins and Selected Natural and Synthetic Stilbenoids. Journal of Agricultural and Food Chemistry. 59(5):1673-1682.

Amin, E., El-Hawary, S.S., Fathy, M.M., Mohammed, R., Ali, Z., Tabanca, N., Wedge, D.E., Becnel, J.J., Khan, I.A. 2011. Triterpenoidal saponins: bioactive secondary metabolites from Zygophyllum coccineum L. Planta Medica. 77:485-488.

Valles, S.M., Becnel, J.J., Pereira, R.M. 2011. Kneallhazia carolinensae sp. nov., a microsporidian pathogen of the thief ant, Solenopsis carolinensis. Journal of Invertebrate Pathology. 108:59-62.

Hall, A.C., Griffith, T.N., Tsikolia, M., Kotey, F.O., Gill, N., Humbert, D.J., Watt, E.E., Yermolina, Y.A., Goel, S., El-Ghendy, B., Hall, C.D. 2011. Cyclohexanol analogues are positive modulators of GABAA receptor currents and act as general anaesthetics in vivo. European Journal of Pharmacology. 667:175-181.

Kline, D.L., Bernier, U.R., Hogsette, Jr, J.A. 2012. Efficacy of three attractant blends tested in combination with carbon dioxide against natural populations of mosquitoes and biting flies at the lower suwannee wildlife refuge. Journal of the American Mosquito Control Association. 28(2):123-127.

Ozek, G., Ishmuratova, M., Tabanca, N., Radwan, M.M., Goger, F., Ozek, T., Wedge, D.E., Becnel, J.J., Cutler, S.J., Baser, K.H. 2012. One-step multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak. by preparative capillary GC with characterization by spectroscopic and spectrometric techniques and evaluation of biological activity. Journal of Separation Science. 35(5-6):650-660.

Kovendan, K., Murugan, K., Panneerselvam, C., Kumar, M., Amerasan, D., Subramaniam, J., Vincent, S., Barnard, D.R. 2011. Laboratory and field evaluation of medicinal plant extracts against filarial vector, Culex quinquefasciatus Say (Diptera: Culicidae). Parasitology Research. 110:2105-2115.

Coutinho, C.J., Alves, R., Sanscrainte, N.D., Viviani, A., Paulo, S.F., Souza, P.A., Carvalho-Mello, I.M., Becnel, J.J. 2012. Occurrence and phylogenetic characterization of a baculovirus isolated from Culex quinquefasciatus in São Paulo State, Brazil. Archives of Virology. DOI: 10.1007/s00705-012-1372-1.

Last Modified: 06/25/2017
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