Behavioral Adaptations of House Flies (Musca domestica L.) to Avoid the Insecticide Imidacloprid

Transcription

1 Behavioral Adaptations of House Flies (Musca domestica L.) to Avoid the Insecticide Imidacloprid Daniel A. Wasik, 2 Alec C. Gerry 1 1 Department of Entomology 2 Department of Environmental Science University of California, Riverside A B S T R A C T House fly (Musca domestica L.) resistance to insecticides is a growing problem for animal agriculture in southern California. Previous studies have demonstrated fly resistance to imidacloprid under laboratory and field conditions and suggested that resistance was partially due to changes in behavior. This study examined whether imidacloprid-resistant house flies detected and avoided imidacloprid prior to contact or whether resistance was due to reduced exposure following contact. Flies from insecticide-resistant and insecticide-susceptible laboratory colonies were placed into an enclosed arena with separate dishes containing either imidacloprid-treated or untreated sugar for food. Fly behaviors including visitation to the dishes and time spent on a dish were recorded using video cameras over a 15 minute period. House flies from both colonies similarly visited food dishes containing either untreated or imidaclopridtreated sugar, indicating that resistant flies were not detecting the presence of imidacloprid and avoiding contact. Following contact with the food dishes, flies from both colonies disengaged from imidacloprid-treated sugar dishes more often than from untreated sugar dishes. However, resistant flies disengaged from an imidacloprid-treated sugar dish significantly more often than susceptible flies. Also, while susceptible flies consumed both treated and untreated sugar equally, resistant flies consumed significantly less treated than untreated sugar. Imidacloprid appeared to act as a contact irritant causing locomotion stimulation in both susceptible and imidacloprid-resistant house flies, but the stimulation was significantly stronger in the resistant flies. Imidacloprid may also have acted as a feeding deterrent in resistant flies, but it was difficult to separate this effect from the irritancy effect. F A C U L T Y M E N T O R Alec C. Gerry Department of Entomology A U T H O R Environmental Sciences is a graduating senior who spent the last two years in the laboratory of Professor Alec Gerry examining the behavioral adaptations of insecticideresistant house flies adaptations which allow them to avoid lethal exposure to a toxicant. Dan is proud to see his research culminate with a journal article. He is excited to be returning to UCR next year to attend graduate school in the Environmental Toxicology Program. Dan is incredibly grateful to Dr. Gerry for serving as his mentor, the CNAS STEM Pathway Project, and the CNAS Undergraduate Research Scholarship Program for their enormous generosity which allowed him to focus on his studies while also having the opportunity to conduct research. Dan was initially introduced to my laboratory through the CNAS STEM Pathway Project which provides undergraduate students an opportunity to acquire research experience in an active laboratory at UC Riverside. When I first met Dan, he expressed an interest in chemistry and toxicology and wondered if there might be a project in my laboratory to match these interests. I agreed to mentor Dan, and we developed a research project that brought his interests together with my laboratory goals. Dan has a sharp mind and an inquisitive nature, and was soon discovering how insecticide-resistant house flies avoid lethal exposure to a toxicant commonly used for their control. I won t give away the answer here you must read the article! Dan presented part of his research at the 2009 Southern California Conference of Undergraduate Research and at the 2010 UCR Symposium for Undergraduate Research, Scholarship and Creative Activity. These are wonderful opportunities for undergraduate students to acquire important public speaking and presentation skills. There is little value in discovery if new knowledge cannot be communicated with others. I am certain that this article, his first research publication, will not be his last! U C R U n d e r g r a d u a t e R e s e a r c h J o u r n a l 3 9

2 INTRODUCTION The common house fly (Musca domestica L.) is a well known pest in animal agriculture and can also be common in urban settings in association with food industries and residential homes. 1 House flies are no longer thought of as only a nuisance, as they are known to mechanically transport disease-causing bacteria such as Salmonella enteritidis, Escherichia coli, and Shigella sonnei. 2,3 Following contact with these pathogens, flies are capable of spreading them through defecation, regurgitation, and contaminated body parts. 3 These pathogens are typically acquired from contact with livestock and can be distributed to other sources of food prior to human consumption. 3,4 It is important that measures are taken to reduce the population of house flies near food sources prone to contamination. Primarily, workers must practice good sanitation to eliminate fly breeding at animal facilities. To rapidly reduce the adult fly population insecticides are commonly used along animal feed lanes, outside milking parlors, or at other areas where flies congregate. 5 In order to achieve expected levels of adult fly control, increased application frequency and greater concentration of material is applied, and therefore greater environmental exposure to these insecticides. Due to the overuse of insecticides and to rapid generation time, flies have developed some level of resistance to most available insecticides, with some insecticides being essentially useless. 6 In 2003, Bayer Crop Sciences registered a house fly food bait containing imidacloprid. As the only effective bait product for control of house flies, it was used extensively throughout southern California in subsequent years. Reliance on one product for fly control has resulted in the very rapid development of resistance to imidacloprid by house flies. 2,7 House flies collected in 2008 from a Riverside County dairy were significantly more resistant to imidacloprid relative to house flies collected from the same location in When exposed to imidacloprid via a nochoice feeding assay, the only source of food is insecticide treated, resulted in a survival rate of 51% in 2008 compared to 17% in In both tests, the imidacloprid concentration was twice the lethal dose that results in 99% mortality for susceptible flies. 7 However, when tested in choice assays, giving flies access to both insecticide-treated and untreated food, the survival rate of flies collected in 2008 was even greater with majority of flies surviving even the highest imidacloprid applications. 7 Results from no-choice and choice assays provide evidence that house fly resistance to imidacloprid was due to evolutionary selection of house fly phenotypes conferring resistance through both altered physiological and behavioral mechanisms. Although physiological mechanisms of pesticide resistance have been well studied, it was only recently recognized that variation in insect behavior may similarly be acted on by selection to confer insecticide resistance. While previous studies have demonstrated that house flies are behaviorally resistant to imidacloprid, the specific behavioral changes associated with this resistance are unclear. Resistance may be due to detection and avoidance of the insecticide prior to house fly contact with the insecticide (volatile odor mediated repellency) or to irritancy and movement away from the insecticide following contact with it (excitatory response). Both responses would be expected to lead to resistance within the population over time as flies avoid a lethal exposure to the insecticide and survive to reproduce. The purpose of this study was to compare feeding behaviors of imidacloprid susceptible and resistant house fly populations to determine which behaviors have been altered leading to reduced contact with the insecticide. MATERIALS AND METHODS Fly Colonies Two separate house fly colonies were used in this study. A susceptible house fly colony (UCR colony) has been reared in a laboratory at UCR since their collection from a southern California dairy in 1982; this colony has never been exposed to imidacloprid. An imidaclopridresistant house fly colony (BS colony) has been reared in a laboratory at UCR since their more recent collection from a southern California dairy (BS Dairy) in Flies from the BS colony have been previously shown to be resistant to imidacloprid. 2,7 Both colonies were reared using standard methods U C R U n d e r g r a d u a t e R e s e a r c h J o u r n a l

3 Feeding Assay Differences in feeding behavior between susceptible and resistant fly colonies were examined using a choice feeding assay where flies from each colony were provided the opportunity to locate and feed on imidacloprid-treated or untreated sucrose. Groups of approximately 100, 3-5 day old adult house flies, estimated volumetrically (mixed sex), from each colony were placed into separate containers with only a water source and starved for 24 hours prior to the start of a feeding trial in order to increase fly response to sucrose during the trials. Starved imidacloprid resistant and susceptible flies were then introduced into separate clear plastic arenas (50x40x40 cm) with mesh windows to allow for air flow and 2 mounted video cameras to record fly activity at each food source. Flies were allowed to acclimate for 15 min before two sucrose treatments were introduced into each arena. Sucrose treatments were 3g of sucrose in a weigh dish to which was added either 1 ml of 250 μg/ml imidacloprid in acetone (treated dish) or 1 ml of acetone only (control). The acetone was allowed to evaporate for 24 hours from each dish prior to the start of every trial. Each sucrose dish was weighed before and after a trial to determine the amount of sucrose consumed by the flies. Each video camera recorded fly activity at a single sucrose dish with the number of landing events onto and departures from each sucrose dish recorded for 15 min after introducing the dishes to the arena. Flies within the arena had an equal opportunity to locate and land on either the treated or untreated sucrose dish. In order to ensure there was no bias for a dish due to unpredicted or unseen phenomenon, such as room lighting or undetected scent, dish positions were switched after every trial. If a fly lost contact with the dish and then returned it was considered a new visit even if it stayed within the video frame. If a fly walked along the outside edge of the dish and then returned to the inner edge or food source, it was considered the same visit. Choice trials were repeated nine times for each fly colony. For each fly colony, differences in landing and disengaging from the treated and untreated sucrose dishes were examined by Wilcoxon matched pairs test. Between fly colonies, differences were examined by Mann-Whitney Test on either count or proportional data. The W value given by the Wilcoxon matched pairs test and the U value given by the Mann-Whitney test indicate the strength of p. As the W or U value move away from zero, the p value becomes smaller. A p value of <0.05 was considered a significant difference between the experiment results being compared. Both tests were non-parametric, they do not make any assumptions as to the distribution of the data. RESULTS AND DISCUSSION The number of flies landing on either the imidaclopridtreated or untreated sucrose dishes was not significantly different (p>0.05) for either the susceptible (UCR) or resistant (BS) house fly populations (Figure 1). Landing behaviors were similarly not different (p>0.05) between the two fly populations. Although the BS flies are known to be resistant to imidacloprid, they exhibited no ability to detect and avoid the imidacloprid before landing on treated sugar as would be expected if the BS flies could detect volatiles associated with the toxicant. Figure 1. Mean ± standard error number of flies landing on dishes containing imidacloprid-treated sucrose or untreated sucrose was not significantly different (P>0.05) for either the UCR (susceptible) colony or the BS (imidacloprid-resistant) colony. Nine trials per colony were run. U C R U n d e r g r a d u a t e R e s e a r c h J o u r n a l 4 1

4 For both fly populations, there were significant differences between the number of flies disengaging from the treated and untreated sucrose dishes, with more flies disengaging from the treated dish relative to the untreated dish (UCR:W>37, p<0.03, BS:W>45, p<0.004) (Figure 2). However, the imidacloprid-resistant BS flies disengaged from the treated sucrose dish significantly more often that the susceptible UCR flies (U=15, p<0.03). The greater disengagement of susceptible UCR colony flies from the treated sucrose dish relative to the untreated sucrose dish appeared to show that behaviors were already available in susceptible house fly populations which could be acted upon by selection to increase avoidance of and therefore resistance to imidacloprid. The resistant BS flies have been selected to avoid a lethal exposure to imidacloprid by exhibiting increased disengagement from a food source containing imidacloprid relative to the susceptible UCR flies. Imidacloprid was acting as a locomotor stimulant, 9 a substance resulting in increased movement; in this case away from the imidacloprid-treated sugar. Figure 2. Mean ± standard error percent change of flies that disengaged from the treated sucrose dish following initial contact was far greater than that of flies disengaging from the untreated sucrose dish for both colonies. Nine trials per colony were run. Further evidence for altered contact behavior in resistant flies was clear in the significantly reduced consumption of treated sucrose relative to untreated sucrose (W=36, p<0.01), while susceptible flies showed no significant difference in consumption of treated and untreated sucrose (p>0.05). Figure 3. Mean ± standard error percent of sugar consumed by the susceptible UCR colony flies was not significantly different between treated and untreated sucrose. However, the resistant BS colony flies consumed much less of the treated sucrose than the untreated sucrose. Nine trials per colony were run. Given these results, consideration must be given to the management of both physiological as well as behavioral resistance whenever a new pesticide enters the commercial market. Previous methods of determining insecticide resistance in the field by counting flies on a treated food source at predetermined intervals may need to be reconsidered. The results of the study suggest that visitation data collected by time interval counts may not provide an accurate measurement of visitation due to excitatory repellency or locomotor stimulation following contact with the toxicant resulting in rapid disengagement from the food source. House flies may still be landing on the food as much before, but simply do not remain long enough to be counted. Although the susceptible UCR fly population shows an increased disengagement response to contact with imidacloprid, their similar consumption of treated and untreated sugar indicated that this disengagement response 4 2 U C R U n d e r g r a d u a t e R e s e a r c h J o u r n a l

5 was not protective from a lethal exposure to the toxicant in a food. In contrast, the BS flies limited their contact with and consumption of imidacloprid. Overuse of imidacloprid to control house flies in the field has resulted in selection for increased contact repellency behaviors such that consumption of the imidacloprid-treated sugar is greatly reduced. Because imidacloprid fly baits must be consumed for the toxicant to affect a fly, the evolution of behaviors to reduce consumption of imidacloprid, in conjunction with the previously determined physiological resistance to imidacloprid has resulted in the lack of effect of these fly baits under natural field conditions over the last two years. 2 REFERENCES 1. Butler, S.M., Gerry, A.C., and Mullens, B.A House fly (Diptera: Muscidae) activity near baits containing (Z)-9-tricosene and efficacy of commercial toxic baits on a southern California dairy. Journal of Economic Entomology. 100: Gerry, A. C. and Zhang, D Behavioral resistance of house flies, Musca domestica L (Diptera: Muscidae) to Imidacloprid. US Army Medical Department Journal. Jul-Sep: Ostrolenk, M. and Welch, H The common house fly (Musca domestica) as a source of pollution in food establishments. Food Research International. 7: Talley, J. L., A. C. Wayadande, L. P. Wasala, A. C. Gerry, J. Fletcher, U. DeSilva, and S. E. Gilliland Association of Escherichia coli O157:H7 with filth flies (Muscidae and Calliphoridae) captured in leafy greens fields and experimental transmission of E. coli O157:H7 to spinach leaves by house flies (Diptera: Muscidae). Journal of Food Protection. 72(7): Gerry, A. C., N. G. Peterson, and B. A. Mullens Predicting and controlling stable flies on California dairies. Oakland: University of California, Division of Agriculture and Natural Resources. Publication pp Kaufman, P. E., and Rutz, D.A Susceptibility of house flies (Diptera: Muscidae) exposed to commercial insecticides on painted and unpainted plywood panels. Pest Management Science. 58: Kaufman, P.E., Gerry, A.C., Rutz, D.A., and Scott, J.G Monitoring susceptibility of house flies (Musca domestica L) in the United States to Imidacloprid. Journal Agricultural and Urban Entomology. 23(4): Mandeville, J.D., Mullens, B.A., and Meyer, J.A Rearing and host age suitability of Fannia canicula`ris (L.) (Diptera: Muscidae) to parasitation by Muscidifurax zaraptor Kogan and Legner (Hymenoptera: Pteromalidae). Canadian Entomologist. 120: Miller, J. R.; Siegert, P. Y.; Amimo, F. A.; Walker, E. D Designation of Chemicals in Terms of the Locomotor Responses They Elicit from Insects: An Update of Dethier et al. (1960). Journal of Economic Entomology. 102(6): U C R U n d e r g r a d u a t e R e s e a r c h J o u r n a l 4 3

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