Journal of the Ameican Mosquito Control Association, 12(2):331-333, 1996

EFFECTIVENESS OF REPELLENT FORMULATIONS CONTAINING DEET AGAINST MOSQUITOES IN NORTHEASTERN '

S. P. FRANCES,''4 CHIRAPA EAMSILA,3 CHAIYAPHRUK PILAKASIRI3 ENO K. J. LINTHICUM,

ABSTRACT Formulations of the repellent diethyl methyl benzamide (deet) were evaluated against mosquitoes in field tests conducted in Province, northeastern Thailand. Thirty-three percent deet in an applicator stick provided >87.lVo protection against primarily Culex vishnui, Cx. gelidus' and Cx. tritaeniirhynchas for up to 5 h, and 5OVo deet in ethanol provided >93.3Vo protection against Anopheles sp. and culicines for up to 8 h. The U.S. Army Extended Duration Repellent Formulation (tEDRFl, containing 33Vodeet),5OVodeetinethanol, andT5Vo deetinethanolprovidedcompleteprotectionforup to 6 h against primarily Culex vishnui and Anopheles dirus.

The mosquito repellent diethyl methyl benz- The repellent formulations tested wete 33Vo amide (deet) was first discovered in 1954 and deet formulated into an applicator stick (Inter- has since been used throughout the world in nu- grade Trading Co., ), and a liquid con- merous formulations (Gupta and Rutledge taining 5OVo deet in ethanol (Intergrade Trading 1994). Although deet has a broad spectrum of Co.). A trial of the 33Vo deet stick formulation effectiveness against many arthropods, including was conducted on each of 3 nights inJuly 1992 mosquitoes, the response of different species and a second trial with 5OVodeet was conducted varies (Rutledge et al. 1978). Field tests con- on 2 nights in September 1992. Sixteen male ducted against mosquitoes in southeastern Thai- Thai soldiers acted as volunteers during the tri- land in 1965 reported the effectiveness of 6 re- als. All volunteers wore short trousers, and pellents (57o dilutions) and showed that only long-sleeve shirts during the trials. Repellent deet protected completely for 45 min or more was applied evenly to the exposed skin of the (Gilbert et al. 1970).In this note we report field legs, face, and hands of 8 volunteers; the re- studies on the effectiveness of topical repellent maining 8 volunteers were untreated. Both re- formulations containing deet against mosquitoes pellents were applied ad libertum by the vol- in northeastern Thailand. unteers to ensure an even coverage of the ex- During 1992, tests were conducted in a Roy- posed skin, but in this operational trial, the al Thai Army (RTA) camp situated about I km amount applied was not known. The volunteers from O'Trao refugee encampment, Phusing were assigned to 8 groups of 2 individuals (4 District, Sisaket Province, northeastern Thai- control groups, 4 repellent groups), and each land. The camp was situated in an area where member of a pair collected mosquitoes from his extensively for native forest had been cleared partner. In July 1992, repellent was applied to timber production. The relatively few trees that the test individuals at 1830 h, and in September remained were less than 20 years old and in- 1992 tests, repellent was applied at 1800 h on terspersed with secondary scrub vegetation. the first night and 1600 h on the second night of testing. For each test all volunteers assem- 'The views of the authors do not purport to reflect bled in their groups and walked about 50-100 the position of the Department of the Army or the m into the surrounding bushland. They sat in a Department of Defense. Mention of a commercial predetermined area and collected all biting product does not constitute an endorsement of the mosquitoes using small glass tubes for 20 min product by the Department ofDefense. The volunteers each hour from 1900 to 24OO h. The collected gave informed consent before participating in the study. mosquitoes for each pair and collection time 2 Department of Entomology, U.S. Army Medical were placed into a cup, counted, and returned Component, Armed Forces Research Institute of Med- to Bangkok for identification. ical Sciences, 315/6 Rajvithi Road, Bangkok 10400 In July 1992, a total of 390 mosquitoes, rep- Thailand. resenting 2l species from 4 genera, were col- 3 Research Division, Royal Thai Component, Armed lected on the control volunteers. The predom- Forces Research Institute of Medical Sciences, 315/6 inant species collected were Culex vishnui Rajvithi Road, Bangkok 1M0O Thailand. (42.7Vo gelidus a Present address: Army Malaria Research Unit, Liv- Theobald of total), Culex erpool Military Area, New South Wales 2174 Austra- Theobald (2OVo), Culex tritaeniorhynchus lia. Giles (12.77o), and Culex whitmorei (Glles)

331 332 JounNnr- oF THE AMERICIN Mosqurro CoNrnol AssocrerroN Vol. 12,No.2

-r Table 1. All mosquitoes(mean/pair/night SE) collected hourly on volunteers with skin either treated or untreatedwith 2 formulations of deet at O'Trao, Phusing District, Sisaket province, Thailand,July and September1992. Hours after Iuly repel- 1992 September1992 lent Protec- Protec- appli- 337o deet Untreated tion 5OVo deet Untreated tion cation (n : 22)1 (n : 39O) (7o) (n : 12) (n :722) (7o) 0 o.4 + O.2 3.1 -f 0.8 87.1 -f I 0.1 + 0.1 1.8 0.6 94.4 ; 9.3 -r 2.2 lo0 -r 'r 2 o.3 o.2 4.4 l.o 93.2 0 r2.o -r 2.0 100 -f 3 0.5 +- 0.4 5.1 1.1 90.2 o 16.7 -r 4.6 100 -r -+ 4 o.2 o.2 8.9 3.7 98.2 o 6.6 + 2.3 100 -r 5 0.4 0.3 9.2 + 4.2 95.4 1.5 + 1.5 22.4'r 5.8 93.3 6 0 17.4+ 4.8 100 7 0 14.8 -r 7.3 100 8 0 18.5 -'- 5.0 100 rTotal number of mosquitoes collected by each group.

(7.3Vo). All are considered potential vectors of small number of mosquitoes collected on the Japanese encephalitis (JE) in Thailand (Rat- treated volunteers may be attributed to a diffi- tanarithikul and Panthusiri 1994). In Septem- culty in applying the formulation uniformly on ber 1992, l2l Anophele.r sp. were identified the skin. Tlre 5OVo deet formulation was very and Anopheles karwari (James) (47.9Vo of effective in protecting treated volunteers against Anopheles sp.), Anopheles splendidas Koid- Anopheles sp. and culicines for up to 8 h without ntmi (31.47o), and Anopheles nivipes (Theo- any loss of activity. bald) (9.17o) were the predominant species, al- In November 1993, tests were conducted over though none of these are considered to be vec- 2 nights in an RTA camp located in Khukan Dis- tors of malaria in Thailand (Rosenberg et al. trict, Sisaket Province, Thailand. The camp was 1990). The remaining 6Ol culicines collected located in an area of mature native forest. near were not identified. a stream. Three formulations containing deet The protection provided by 2 formulations of were evaluated:5OVo deet in ethanol (Intergrade deet at O'Tfao is shown in Table l.'f};ie 337o Co.),75Vo deet in ethanol (), deet stick formulation provided >87.lvo protec- and the U.S. Army Extended Duration Repellent tion against primarily Culex spp. for up to 5 h, Formulation (EDRF), containing 33Va deet, ac- without a decrease of protection over time. The rylate polymers, and other inert ingredients (3M

Table 2. Mosquitoes (mean/pair/night -r SE) collected hourly by volunteers treated with 3 formulations of deet and untreatedvolunteers at an RTA camp in Khukan District, Sisaket Province. Thailand. November 1993. Hours after repel- Anopheles dirus All mosquitoes lent applica- EDRF 5OVo deet 75Vo deet Untreated EDRF tion (n : J)t (n=7) (n:5) (n = 57) (n=4) 40 o 0 0.5 -f 0.5 o 50 o 0 1.0 -f o 0 600.5 + 0.5 o 4.0 + 1.0 0 70 o o 6.5 + 2.5 o 8 1.5+ 0.5 2.O 4- 2.O 1.5 -f 0.5 6.5 -'- 0.5 1.5 + 0.5 901.0 + l.O 1.0 + 1.0 lo.o + 1.0 0.5 + o.5

I Total number of mosquitoes collected by each group. JuNe1996 OPERATIoNALAND SctENTlFlc NorEs JJJ

Corp., St Paul, MN). In these tests 4 pairs of of the current test also show that EDRF pro- volunteers, who wore military shirts buttoned at vides similar protection to repellents containing the wrist and long trousers rolled up to the knee, 5OVoandT5Vo deet in ethanol, and suggest that were used. Each pair of volunteers either tested these formulations are more effective against one of the 3 formulations or was untreated. The An. dirus than repellents containing lower con- repellent was applied under supervision at 140O centrations of deet. h. For liquid formulations, 1 ml of repellent was We thank soldiers from the Royal Thai applied evenly to each leg, including the foot, Army for acting as volunteers; Pradith Kaews- up to the knee, and I ml to the face, neck, and athien, Naruphon Kuttasingki, and Tawatchai hands. The EDRF was applied according to label Chaimanurat for field assistance; Chumnong instructions by applying a 2.5-ml strip of lotion Noigamol and Kol Mongkolpanya for identi- to each leg and 2.5-ml strip to the face, neck, fying mosquitoes; R. Rosenberg for advice and hands. Collections began at 1800 h when and encouragement; and R. D. Cooper for the volunteers entered the nearby forest and col- valuable comments on the manuscript. The lected all biting mosquitoes using small glass study was conducted under WRAIR protocol tubes for 45 min, followed by a 15-min rest. 396, and was approved by the WRAIR ethics Collections were then made each hour for a total committee and Director General of AFRIMS. of6h. This paper is published with the approval of The predominant mosquitoes collected during the Director General of Army Health Services these tests were C-x.vishnui (58.4Vo) and Anoph- (Australia). eles dirus Peyton and Harrison (34.7Vo). The low (total col- overall density of mosquitoes was REFERENCES CITED lected : 2O2) duing the tests; 5OVadeet provid- ed protection against all mosquitoes for up to 6 Frances,S. P, N. Eikarat,B. Sripongsaiand C. Eam- h, and EDRF andT5Vo deet provided protection sila. 1993. Responseof Anophelesdirus andAedes albopictus to repellents in the laboratory. J. Am. for up to 8 h (Table 2). Mosq. Control Assoc.9:47 4-47 6. Recent laboratory and field studies have Frances, S. P, T A. Klein, D. W. Hildebrandt, R. shown that An. dirus, the major vector of ma- Burge, C. Noigamol, N. Eikarat, B. Sripongsaiand laria in Thailand, is tolerant of formulations R. A. Wirtz. 1996. Laboratory and lield evaluation containing lower concentrations of deet. In the of the repellents, deet, CIC-4 and Al3-3'722O, laboratory, low concentrations of deet (=35Vo) against Anopheles dirus (Diptera: Culicidae) in provided protection against An. dirus for <90 Thailand. J. Med. Entomol. 33 (in press). min (Frances et al. 1993). In a subsequent field Gilbert, I. H., J. E. Scanlonand D. L. Bailey. 1970. test at , southeastern Repellentsagainst mosquitoesin Thailand. J. Econ. Entomol. 63:12O7-12O9 Thailand, 25Vo deet in ethanol provided >95Vo Gupta, R. K. and L. C. Rutledge. 1994. Role of re- protection against An. dirus for only I h after pellents in vector control and disease prevention. repellent application (Frances et al. 1996). Pre- Am. J. Trop. Med. Hyg. 5o(Suppl.):82-86. vious studies have shown EDRR which con- Gupta, R. K., A. W. Sweeney,L. C. Rutledge, R. D. tains 33Vo deet, provides similar protection Cooper,S. P Francesand D. R. Westrom. 1987. against mosquitoes as formulations containing Effectivenessof controlled-releasepersonal-use ar- higher concentrations of deet in ethanol (Gupta thropod repellents and permethrin-impregnated et al. 1987, Harbach et al. 1990). The results clothing in the field. J. Am. Mosq. Control Assoc. 3:556-560. Harbach,R. E., D. B. Thng, R. A. Wirtz and J. B. Table 2. Extended. Gingrich. 1990. Relative repellency of two formu- lations of N,N-diethyl-3-methylbenzamide(deet) and permethrin-treatedclothing againstCulex sitiens andAedes vigilax in Thailand.J. Am. Mosq. Control Assoc.6:641-644. All mosquitoes Rattanarithikul,R. and P. Panthusiri. 1994. Illustrated 5OVo deet 75Vo deet Untreated keys to the medically important mosquitosof Thai- (n:lO) (n:8) (4: 180) land. SoutheastAsian J. Trop. Med. Public Health 25(Suppl.l):l-66. 0029.O-+ l3.O Rosenberg,R., R. G. Andre and L. Somchit. 1990. 009.0 + 6.0 Highly efficient dry seasontransmission of malaria 1.5-f 1.5 0 8.5 -f 1.5 in Thailand. Tians. R. Soc. Tiop. Med. Hyg.84:22- 0013.5+ 1.5 28. M. Moussa,C. A. Lowe and R. K. 2.5 1- 2.5 2.O-+ O l3.o -+ 2.o Rutledge,L. C., A. + -r -f Sofield. 1978. Comparativesensitivity of mosquito 1.0 1.0 2.o 2.o 17.0 3.0 speciesand strains to the repellent diethyl toluam- ide. J. Med. Entomol. 14:536-541.