The susceptibility of Aedes aegypti populations displaying temephos resistance to Bacillus thuringiensis israelensis: a basis for management

Parasites & Vectors The susceptibility of Aedes aegypti populations displaying temephos resistance to Bacillus thuringiensis israelensis: a basis for management Ana Paula Arajo 0 Diego Felipe Araujo Diniz 0 Elisama Helvecio 0 Rosineide Arruda de Barros 0 Cludia Maria Fontes de Oliveira 0 Constncia Flvia Junqueira Ayres 0 Maria Alice Varjal de Melo-Santos 0 Lda Narcisa Regis 0 Maria Helena Neves Lobo Silva-Filha 0 0 Department of Entomology, Centro de Pesquisas Aggeu Magalhaes-FIOCRUZ , Recife, PE 50670-420 , Brazil Background: Aedes aegypti is the vector of dengue virus, and its control is essential to prevent disease transmission. Among the agents available to control this species, biolarvicides based on Bacillus thuringiensis serovar israelensis (Bti) are an effective alternative to replace the organophosphate temephos for controlling populations that display resistance to this insecticide. The major goal of this study was to determine the baseline susceptibility of Brazilian Ae. aegypti populations to Bti, taking into account their background in terms of larvicide exposure, status of temephos resistance and the level of activity of detoxifying enzymes involved in metabolic resistance to insecticides. Methods: Population samples were established under insectarium conditions. Larval susceptibility to temephos and Bti was evaluated through bioassays and lethal concentrations of these compounds were determined. Biochemical assays were performed to determine the specific activity of five detoxifying enzymes in these samples. Results: Fourteen populations were characterized and, except for one case, all displayed resistance to temephos. Most populations were classified as highly resistant. The populations also showed increased activity of one or more detoxifying enzymes (glutathione-S-transferases, esterases and mixed function oxidases), regardless of their temephos resistance status. All populations analyzed were susceptible to Bti, and the lethal concentrations were similar to those detected in two laboratory susceptible colonies. The response to Bti showed little variation. A maximum resistance ratio of 2.1 was observed in two untreated populations, while in two Bti-treated populations, the maximum resistance ratio was 1.9. No positive correlation was found between temephos resistance, increased activity of detoxifying enzymes, and susceptibility to Bti. Conclusions: Data from this study show that all populations were susceptible to Bti, including twelve untreated and two treated populations that had been exposed to this agent for more than ten years. The temephos resistance and increased activity of detoxifying enzymes observed in thirteen populations was not correlated with changes in susceptibility to Bti. Our data show a lack of cross-resistance between these two compounds; thus, Bti can be used in an integrated control program to fight Ae. aegypti and counteract the temephos resistance that was found among all populations analyzed. Vector control; Bti; Susceptibility; Temephos; Metabolic resistance; Cross-resistance - Background Aedes aegypti is the major vector of dengue virus (DENv) and has been responsible for an important disease burden in human populations worldwide in the last few decades [1,2]. This species has spread over most municipalities in Brazil, where it is the main DENv vector and has provoked frequent epidemics since 1986 [3]. Vector control remains the sole action to control dengue because vaccines and other prophylactic measures are not currently available. In this context, a National Program for Aedes aegypti Eradication (PEAa) was created in 1996 in Brazil and was replaced by the National Program for Dengue Control (PNCD) in 2002. The main goal of the PNCD is to fight this disease through integrated control actions, including the utilization of chemical larvicides and adulticides [4]. The organophosphate (OP) temephos has been the major larvicide used by the PNCD, and one of the most used compounds to control mosquitoes globally, despite its negative effects on nontarget organisms and reports of resistance. In mosquito, temephos resistance has been associated with the alteration of its target site in acetylcholinesterase and also with metabolic mechanisms associated with enzymes involved in the detoxification of xenobiotic compounds [5-7]. Biolarvicides based on the entomopathogenic bacteria Bacillus thuringiensis serovar israelensis (Bti) have been successfully used for dipteran control [8]. Bti was first introduced for controlling Simulium, and its utilization was later extended to Aedes species. Long-term programs carried out in many countries have demonstrated its effectiveness under field conditions [9-11]. Its larvicidal action is based on crystals produced upon bacterial sporulation, mainly composed of the four protoxins Cry11Aa, Cry4Aa, Cry4Ba and Cyt1Aa. Btis mode of action depends on the ingestion of these crystals by larvae. Crystal solubilization occurs at the alkaline pH of the midgut, and the protoxins released into the lumen are converted into active toxins by proteases [12]. The whole crystal displays optimal toxicity, whereas individual toxins, or their combinations, do not show comparable levels of activity [13]. Once activated, the Cry toxins bind to specific midgut receptors from Ae. aegypti larvae: cadherins, aminopeptidases and alkaline phosphatases have been identified as binding molecules [14]. Studies to elucidate the synergy among Bti toxins have demonstrated that Cyt1Aa can act as a surrogate receptor for Cry11Aa and Cry4Ba. Furthermore, binding between Cyt1Aa and Cry toxins induces conformational changes that improve the capacity of Cry to bind to the other receptors available in the midgut [15-17]. This complex action based on four toxins with the capacity to bind to different target molecules does not favor the selection of resistance. Previous reports have failed to demonstrate the development of resistance to whole Bti crystal after continuous exposure for resistance selection under laboratory conditions [18-21], and resistance to Bti-based larvicides in field populations has not been reported to date [22,23]. In light of the effectiveness of Bti to control Aedes species and the lack of resistance, its utilization is under expansion in control programs for the treatment of breeding sites. Bti has also been used in oviposition and adult traps to prevent the development of larvae in these devices when they are used for monitoring, or on a massive scale, to reduce mosquito populations [24-26]. Another factor that supports the increasing use of Bti is the worldwide Ae. aegypti resistance to temephos, as has frequently been reported. In Brazil, there is a serious resistance problem that compromises the effects expected from use of temephos by the PNCD [27-33]. Thus, Bti is a candidate to manage resistance to temephos; however, some reports have recently suggested a potential crossresistance between temep (...truncated)