Prasad Kona Madhavinadha, Raghavendra Kamaraju, Verma Vaishali, Velamuri Poonam Sharma, Pande Veena
ICMR-National Institute of Malaria Research, New Delhi, India.
Department of Biotechnology, Kumaun University, Nainital, Uttarakhand, India.
J Vector Borne Dis. 2017 Jul-Sep;54(3):226-232. doi: 10.4103/0972-9062.217613.
BACKGROUND & OBJECTIVES: Increase in prevalence and intensity of insecticide-resistance in vectors of vector-borne diseases is a major threat to sustainable disease control; and, for their effective management, studies on resistance mechanisms are important to come out with suitable strategies. Esterases are major class of detoxification enzymes in mosquitoes, which confers protection against insecticides in causing resistance. This study was aimed at biochemical characterization of esterases responsible for malathion resistance in Anopheles stephensi mosquitoes, along with its validation through biochemical techniques and native-PAGE assays.
Laboratory maintained susceptible and resistant An. stephensi mosquitoes were used for assessing the activity and effect of α - and β -esterases on malathion. Bioassay, synergist bioassay, biochemical assay and native- PAGE were employed to characterize the role of esterases in conferring malathion-resistance.
Notably significant (p < 0.0001) enhancement in α - and β -esterases activity was observed with 2-fold increase in resistant An. StephensiGOA compared to susceptible An. StephensiBB. native-PAGE depicted two major bands 'a' (Rf = 0.80) and 'b' (Rf = 0.72) in susceptible An. stephensiBB , while one intense band 'b' (Rf = 0.72) was visible in resistant An. stephensiGOA. Inhibition assay revealed complete inhibition of α - and β -esterases activity in presence of 1 mM malathion in susceptible strain compared to observed partial inhibition in resistant strain on native-PAGE.
INTERPRETATION & CONCLUSION: This study provides a better understanding on the role of esterase enzyme (carboxylesterase) in conferring malathion-resistance in An. stephensi mosquitoes, as evident from the native-PAGE assay results. The study results could be used in characterizing the resistance mechanisms in vectors and for suggesting alternative chemical insecticide based resistance management strategies for effective vector-borne disease control.
媒介传播疾病媒介中杀虫剂抗性的流行率和强度增加是对疾病可持续控制的重大威胁；为了有效管理这些抗性，对抗性机制的研究对于制定合适的策略很重要。酯酶是蚊子体内主要的解毒酶类，可赋予对杀虫剂的抗性保护。本研究旨在对斯氏按蚊中导致马拉硫磷抗性的酯酶进行生化特性分析，并通过生化技术和非变性聚丙烯酰胺凝胶电泳（native-PAGE）分析进行验证。
使用实验室饲养的敏感和抗性斯氏按蚊来评估α-和β-酯酶对马拉硫磷的活性和作用。采用生物测定、增效剂生物测定、生化测定和非变性聚丙烯酰胺凝胶电泳来表征酯酶在赋予马拉硫磷抗性中的作用。
值得注意的是，与敏感的斯氏按蚊BB相比，抗性的斯氏按蚊GOA中α-和β-酯酶活性显著增强（p < 0.0001），增加了2倍。非变性聚丙烯酰胺凝胶电泳显示，敏感的斯氏按蚊BB中有两条主要条带“a”（迁移率Rf = 0.80）和“b”（Rf = 0.72），而抗性的斯氏按蚊GOA中可见一条强烈的条带“b”（Rf = 0.72）。抑制试验表明，在敏感品系中，1 mM马拉硫磷存在时α-和β-酯酶活性完全被抑制，而在抗性品系的非变性聚丙烯酰胺凝胶电泳中观察到部分抑制。
本研究从非变性聚丙烯酰胺凝胶电泳分析结果可以明显看出，对酯酶（羧酸酯酶）在赋予斯氏按蚊马拉硫磷抗性中的作用有了更好的理解。研究结果可用于表征媒介中的抗性机制，并为有效的媒介传播疾病控制提出基于化学杀虫剂的替代抗性管理策略。
