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Field Evaluation of Arthropod Repellents Journal of the American Mosquito Control Association,12(2):172_176, 1996 FIELD EVALUATION OF ARTHROPOD REPELLENTS. DEET AND A PIPERIDINE COMPOUND. AI3-3722O. AGAINST ANOPHELES FUNESTUS AND ANOPHELES ARABIENSISIN WESTERN KENYA' TODD W. WALKER,' LEON L. ROBERT3 ROBERT A. COPELAND.4 ANDREW K. GITHEKO,s ROBERT A. WIRTZ,6 JOHN I. GITHURE? rNo TERRY A. KLEIN6 ABSTRACT A field evaluation of the repellents N,N-diethyl-3-methylbenzamide (deet) and 1-(3-cy- clohexen-1-yl-carbonyl)-2-methylpiperidine (AI3-3722O, a piperidine compound) was conducted againsr Anopheles funestus and An. arabiensis in Kenya. Both repellents provided significantly more protection (P < 0.001) than the ethanol control. AI3-3722O was significantly more effective (P < 0.001) than deet in repelling both species of mosquitoes. Aller t h, O.l mg/cm, of Al3-3722O provided 89.8Vo and,Tl.lVo protection against An. arabiensis and An. funestzs, respectively. Deet provided >807o protection for only 3 h, and protection rapidly decreased after this time to 6O.2Vo and 35.1Vo for An. irabienvi and Az. funestus, respectively, after t h. Anopheles funestu.r was significantly less sensitive (P < 0.001) to both repellents than An. arabiensis. The results of this study indicate that Al3-3722O is more effective than deet in repelling anophetine mosquitoes in westem Kenya. INTRODUCTION concerning the susceptibility of these mosquito species to arthropod repellent compounds. The vectors of human malaria parasites in The United States Army is currently evaluat- western Kenya are Anopheles gambiae Glles, ing a novel piperidine compound (AI3-3722O) An. arabiensir Patton, and An. Giles funestus that has been shown to be effective in the lab- (Githeko et al. 1993). Anopheles gambiae s.s. oratory against several anopheline species, in- and An. arabiensis are morphologically indistin- cluding laboratory-reared An. gambiae (Cole- guishable members of the An. gambiae complex man et al. 1993, Frances et al. 1993). In a recent (Gillies and de Meillon 1968, Thylor et al. field test, this compound was 1993). shown to be ef- fective against wild populations of Anopheles di- Githeko et al. (1993) reported that the intense rzs Peyton and Harrison in Thailand (S. P Fran- perennial transmission of malaria at Ahero, a ces, unpublished data) Al3-3722O is currently village in western Kenya, was primarily due to under consideration, either alone or in combi- An. funestus, not members of the An. gambiae nation with deet, as a U.S. Army repellent for- complex. Anopheles arabiensis, although pres- mulation. ent in this area, appears to play a minor role in The objective of this study was to determine malaria transmission. No field data are available if AI3-3722O provides more effective and longer lasting protection than deet against An. funestus rThe views of the authors do not purport to reflect and An. arabiensis mosquitoes in the field. Deet the position of the Department of the Army or the is the standard against which the efficacies of Department of Defense. Mention of a commercial candidate skin repellents are evaluated. Field product does not constitute an endorsement of the evaluation of new repellent compounds is nec- product by the Department of Defense. The volunteers essary because behavioral responses to repel- gave informed consent to participate in the study. lents differ between populations 'United States Army Center for Health Promotion feral arthropod and Preventive Medicine, Divisional Support Activity- and laboratory-reared populations (Frances et al. West, ATTN: MCHB-AW-ES, Aurora, CO 80045- 1993). 5001. 3 Department of Chemistry, United States Military MATERIAL AND METHODS Academy, West Point, New York 10996. 4 United States Army Medical Research Unit-Ke- Two repellents, MN-diethyl-3-methylbenza- nya, Unit 641O9, Box 401, APO AE 09831-4109. mide (deet), lOO7o (Morflex 5 Inc., Greensboro, Vector Biology and Control Research Centre, Ke- NC) and 1-(3-cyclohexen-l-yl-carbonyl)-2- nya Medical Research Institute, P O. Box 1578, Kis- methylpiperidine (AI3-3722O), 99Vo (synthe- umu, Kenya. sized by T, P McGovern, Insect Ecol- 6 Department of Entomology, Walter Reed Army In- Chemical stitute of Research, Washington, DC 2O3O7-510O- ogy Laboratory, USDA-ARS, Beltsville, MD) 7 Biomedical Sciences Research Centre, Kenya were evaluated. The study was conducted in Ny- Medical Research Institute, P O. Box 5484O, Nairobi, anza Province, western Kenya, adjacent to the Kenya. Ahero rice irrigation scheme. Ahero is located t72 Juue 1996 REPELLEI.i-TS PON ATTI. ARABIENSIS AND AN. FUNESTUS 173 23 km SE of Kisumu. located on the shore of 2100 h, I h prior to the start of each repellent Lake Victoria. test. At 22Wh. the volunteers entered their des- Twelve adult male residents (>21 years old) ignated houses, sat on chairs, and collected all of the area volunteered to participate in this mosquitoes biting them on their lower legs in study. Repellents were evaluated on the lower 20-mm scintillation vials for 45 min during each legs, using the area between the knee and ankle. hour of the night. A 15-min break followed each Both trouser legs were rolled up to just above collection period. The time, collector, and house the knees and a line was drawn at the knee with were recorded for each captured mosquito. This an ink marker to provide a line of demarcation collection procedure was repeated each hour un- for repellent application and mosquito collec- til approximately 0600 h the next morning so tion. Low-top canvas shoes, all of the same that 8 biting collections were made by each vol- brand, provided a demarcation line at the ankle. unteer. Ambient temperature and humidity were Volunteers wore hooded screened jackets (Bug recorded hourly throughout the test nights. This Out Outdoor Wear@Ltd., Wauwatsoa, WI) to re- 9-h repellent efficacy test was replicated for 6 strict mosquito feeding to the lower legs. test nights. Repellents were prep:red as 5Vo solutions in The collected mosquitoes were transported to absolute ethanol (50 mg/ml) and applied at a rate the Kenya Medical Research Institute (KEMRI), of 0.1 mg/cm2. The amount of repellent varied Vector Biology and Control Research Centre, at among volunteers and was based on the surface Kisian for processing and identification. In a area of application, previous study at this site, Githeko et al. (1993) reported that An. arabiensis comprised lNVo of A:[(a+b+c)t3]h, the mosquitoes captured from the An. gambiae calculated from the averaged measurements of complex. The ribosomal DNA polymerase chain leg height (ft = length between the crease of the reaction (PCR) amplification method described knee and the ankle bone) and circumference ([a by Taylor et al. (1993) was used to confirm the + b + cl/3, where a : circumference just below identification of mosquitoes identified morpho- the knee, b : circumference at mid-calf, and c logically as An. gambiae s.l. = circumference just above the ankle bone). Pri- Nightly collection totals were determined for or to treatment, the lower legs of each volunteer each of the 8 collection times. Totals for each were washed with TOqo ethanol to ensure that 45-min period were then summed and the per- the treatments were applied directly onto the cent protection at each collection time, defined skin. The ethanol was then allowed to evaporate as the number of bites received by an individual before the treatments were applied to the vol- in the treatment group relative to that of the con- * unteers' legs. The legs of the control volunteers trol, was calculated flO0(control treatment)/ were treated with absolute ethanol. contro[. Results of preliminary repellent tests The repellent tests were conducted inside lo- indicated that the data did not meet the distri- cal mud-walled houses, because An. arabiensis butional assumptions of an analysis of variance and An. funestus primarily bloodfeed indoors and that transformations would not normalize (Githeko et al. 1993). Due to the design of hous- the data. Statistical comparisons of repellent ef- es in western Kenya, mosquitoes have free ac- ficacy were made among the treatment groups cess to the interior through the open eves. Mos- using Kruskal-Wallis nonparametric analysis of quitoes can enter and exit freely even though all variance (Statistix 1987). doors and windows may be closed. Bednets were provided to the house occupants so that the RESULTS AND DISCUSSION volunteers were the only accessible blood-meal source. Nightly weather conditions throughout the The study was arranged in a 3-way factorial study were relatively constant. The mean tem- design (2 repellent treatments plus a control X perature was 20.6oC (range: 16.7-23.4"C) arld 8 time intervals X 6 collection nights). The 12 the mean percent relative humidity was 8l7o volunteers were selected at random for treatment (range: 63-lOOVo RH). There was little or no and house so that for each collection night there wind. were 4 participants per treatment and 3 treatment Three genera of mosquitoes (Anopheles, Ae- participants per house (deet, AI3-3722O, and des, and Culex) were collected. Ttwo species of control). Once assigned to a house, volunteer Anopheles (An. funestus and An. gambiae s.l.), treatment groups were evaluated in that house representing 72Vo of the total number, were col- throughout the study. Treatments were changed lected, providing sufficient numbers (5,029) to nightly so by the end ofthe study each volunteer evaluate repellent efficacy. Initially, 3,668 An. had received each treatment twice. funestus and 1,361 An. gambiae s./. were iden- The treatments were applied at approximately tified. Twenty percent (272) of the An. gambiae t74 JounNel-or rse Avtnrcltr Mosquno Cournol AssocnrloN Vor-.12, No. 2 ANOPHELESARABIENSIS 100 z 90 o F 80 ogJ F 70 o E o 60 zF uJ 50 o E 40 rlu z 30 ...€.-.
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