Journal of the American Mosquito Control Association, 24(2):270–280, 2008 Copyright E 2008 by The American Mosquito Control Association, Inc. EFFECTS OF SINGLE AND MULTIPLE APPLICATIONS OF MOSQUITO INSECTICIDES ON NONTARGET ARTHROPODS RYAN S. DAVIS AND ROBERT K. D. PETERSON1 Department of Land Resources and Environmental Sciences, 334 Leon Johnson Hall, Montana State University, Bozeman, MT 59717-3120 ABSTRACT. Mosquito management plans have been implemented in the United States and globally to manage mosquito vectors of West Nile virus and many other diseases. However, there is public concern about ecological risks from using insecticides to manage mosquitoes. Two studies were conducted during the late summers of 2004 through 2006 at Benton Lake National Wildlife Refuge near Great Falls, MT. The first experiment was conducted in 2004 and 2005 to assess acute impacts of mosquito adulticides (permethrin and d-phenothrin) and larvicides (Bacillus thuringiensis israelensis and methoprene) on nontarget aquatic and terrestrial arthropods after a single application. The second experiment was conducted in 2005 and 2006 to assess longer-term impacts of permethrin on nontarget terrestrial arthropods after multiple repeated applications. For aquatic samples, in the first study, no overall treatment effects were observed despite a potentially deleterious effect on amphipods on sample date 1 in 2004. During the same study, 1 of 54 responses had a significant overall treatment effect for sticky-card samples. Many of the responses for sticky- card samples suggested significant time effects and time 3 treatment effects. Three response variables were associated with fewer individuals present in the insecticide-treated plots in a multivariate analysis. For the multiple-spray study conducted in 2005 and 2006, 6 of the response variables collected via sticky cards exhibited significant overall treatment effects, but none was associated with fewer individuals in the insecticide-treated plots. None of the responses collected using sweep-net sampling suggested overall treatment effects. Time and time 3 treatment effects were prevalent in 2005, but no discernable pattern was evident. In general, nearly all of the responses evaluated for either study indicated few, if any, deleterious effects from insecticide application. KEY WORDS Mosquito control, adulticide, larvicide, ecological risk, nontarget organisms, West Nile virus INTRODUCTION increased from the margin of the insecticide application. Pankiw and Jay (1992) found that West Nile virus (WNV) has been a concern for honey bees in cages experienced significant people across the United States since the disease mortality from malathion spray drift. Hester et was initially observed in North America during the al. (2001) observed significant bee mortality in summer of 1999. Since that year, WNV has caused hives that were exposed to malathion both in the largest arboviral encephalitis epidemic in US open fields and in a forested environment. Zhong history (Huhn et al. 2003). The disease has resulted et al. (2003) found that aerially applied naled had in thousands of human morbidity cases and a negative effect on honey bees and reduced their hundreds of deaths (Huhn et al. 2003). Many honey production over a season. Tietze et al. people are concerned about the risks associated (1996) used sentinel crickets, rather than bees, to with managing mosquitoes that vector WNV using measure malathion deposition in a peri-domestic insecticides (Peterson et al. 2006). This concern is environment. Cricket mortality varied from related to the perception that ecological and 12.5% to 48.7%, depending on the location of human exposure to the insecticides will lead to the crickets in residential yards. risks that are more severe than from WNV itself. Other researchers have focused on aquatic Many of the studies conducted to measure invertebrates. In laboratory studies, Siegfried effects of mosquito management chemicals on (1993) found permethrin to be toxic at low arthropods have focused on honey bees (Apis concentrations (2.9–5.9 mg/liter) to several aquat- mellifera L.). Coldburn and Langford (1970) ic insects, including black flies, caddisflies, found high bee mortality from applications of mayflies, and damselflies. Milam et al. (2000) naled, malathion, and pyrethrum; although this found that when permethrin was applied at study did not use ultra-low volume (ULV) 219 ml/ha over test chambers (50-ml beakers) in applications, it suggests that these chemicals the field, the nontargets Daphnia pulex de Geer, may cause nontarget arthropod mortality. Caron Ceriodaphnia dubia Richard, and Pimephales (1979) exposed caged honey bees and beehives to promelas Rafinesque had 90% survival in 8 of ULV applications of malathion, naled, and 10 experiments. Few experimental data are pyrethrum. Bee mortality decreased as distance available for d-phenothrin. Risk assessments have been conducted to 1 To whom correspondence should be addressed. predict mosquito insecticide risk to humans and 270 JUNE 2008 ECOLOGICAL EFFECTS OF MOSQUITO INSECTICIDES 271 to ecological receptors. Peterson et al. (2006) MATERIALS AND METHODS found that risks from adulticides to humans was Two studies (representing 2 replications of 2 most probably negligible, especially in the context separate experiments) were conducted during the of a WNV outbreak. Davis et al. (2007) developed a screening-level ecological assessment. late summers of 2004 through 2006 at Benton Results from this assessment predicted that Lake National Wildlife Refuge near Great Falls, adulticide risks to terrestrial vertebrates and MT. The first experiment was conducted in 2004 aquatic organisms would also be negligible. and 2005 to assess acute impacts of mosquito The predominant larvicides now used in the adulticides and larvicides on nontarget aquatic United States are Bacillus thuringiensis israelensis and terrestrial arthropods after a single applica- (Bti) and the insect growth regulator methoprene. tion. The second experiment was conducted in The Bti endotoxins are almost nontoxic to 2005 and 2006 to assess longer-term impacts of a mammals and birds (Mittal 2003), although toxic mosquito adulticide on nontarget terrestrial to some aquatic receptors. Milam et al. (2000) arthropods after multiple applications. found that toxicity of Bti was much greater to Daphnia pulex than to Anopheles quadramiculatus Terrestrial and aquatic single-spray study, Say when applied as a liquid formulation. 2004–2005 Ecological effects have been monitored for Bti used for black fly and mosquito management. The site for the first experiment was near a road Merritt et al. (1989) found few changes in indices that parallels Pond 2 within the wildlife refuge used to measure treatment effects of Bti used for (47u41934.430N, 111u20947.690W–47u4195.680N, black fly management in a Michigan river. Drift 111u20954.010W). The experiment was arranged in samples taken at a control and treatment site did a randomized complete block design (RCBD) with 5 not differ for chironomids, baetids, gammarids, treatments replicated 3 times. The blocking factor or hydropsychids, but did have some treatment was location along the length of the experimental effects on perlid stoneflies and elmid beetles. site. Experimental units were 30.48 m in length, Similar results were observed in 10 stream trials 22.86 m in width, with a buffer zone of 15.24 m measuring stream insect density of selected taxa. between each unit. The total length of the site was Molloy (1992) observed that Bti applied for black approximately 686 m, and the area was approxi- 2 fly control within a New York stream affected mately 15,678 m . Treatments consisted of 2 adulti- filter-feeding chironomids, but not surface-dwell- cides applied at their maximum labeled rates, d- ing or tube-dwelling members of the same family. phenothrin (4 g/ha) + PBO (39.2 g/ha) (Anvil 10+10; Caddisflies and mayflies also showed no response Clark Mosquito Control, Roselle, IL) and permeth- to Bti treatments. rin (7.8 g/ha) + PBO (39.2 g/ha) (Aquareslin; Well- Although methoprene toxicity to terrestrial mark, Jayhawk, KS), 2 larvicides applied directly to vertebrates is very low, fish are susceptible to water, Bti (302.6 g/ha) (Vectobac, Valent BioSci- methoprene and Bti exposure. Methoprene de- ences, Walnut Creek, CA) and methoprene (14 g/ grades quickly in soil, groundwater, exposed ha) (Altosid Wellmark International, Schaum- water, and on vegetation (USEPA 1991). Meth- burg, IL), and an untreated control. Adulticides oprene degrades rapidly in water; its reregistra- were applied via ULV sprayer downwind to the tion eligibility document suggests that 80% will pond at dusk to represent a reasonable worst-case degrade within 13 days after application (USEPA acute exposure scenario. Larvicides were applied 1991). Methoprene has adverse effects on aquatic as liquids directly to water. arthropods, including the freshwater amphipod Samples were taken from each plot 1, 7, 14, and Gammarus sp. (Breaud et al. 1977), the mayfly 28 days after the treatments were applied, using a Callibaetis pacificus Seemann, the dytiscid beetle variety of techniques to capture both terrestrial Laccophilus sp., and chironomids (Norland and and aquatic invertebrates. For terrestrial arthro- Mulla 1975). pods, 2 Olson yellow sticky cards (Olson Products, We assessed some of the potential ecological Medina, OH) (7.62 3 12.7 cm) were placed in effects associated with mosquito management each plot at 1 m