Meyers Jacob I, Gray Meg, Foy Brian D
Department of Biomedical Sciences, Colorado State University, 1617 Campus Delivery, Fort Collins, CO 80523-1617, USA
Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, 1692 Campus Delivery, Fort Collins, CO 80523-1692, USA.
J Exp Biol. 2015 May 15;218(Pt 10):1487-95. doi: 10.1242/jeb.118596.
The glutamate-gated chloride channel (GluCl) is a highly sensitive insecticide target of the avermectin class of insecticides. As an alternative to using chemical insecticides to kill mosquitoes, we tested the effects of purified immunoglobulin G (IgG) targeting the extracellular domain of GluCl from Anopheles gambiae (AgGluCl) on the survivorship of three key mosquito disease vectors: Anopheles gambiae s.s., Aedes aegypti and Culex tarsalis. When administered through a single blood meal, anti-AgGluCl IgG reduced the survivorship of A. gambiae in a dose-dependent manner (LC50: 2.82 mg ml(-1), range 2.68-2.96 mg ml(-1)) but not A. aegypti or C. tarsalis. We previously demonstrated that AgGluCl is only located in tissues of the head and thorax of A. gambiae. To verify that AgGluCl IgG is affecting target antigens found outside the midgut, we injected it directly into the hemocoel via intrathoracic injection. A single, physiologically relevant concentration of anti-AgGluCl IgG injected into the hemocoel equally reduced mosquito survivorship of all three species. To test whether anti-AgGluCl IgG was entering the hemocoel of each of these mosquitoes, we fed mosquitoes a blood meal containing anti-AgGluCl IgG and subsequently extracted their hemolymph. We only detected IgG in the hemolymph of A. gambiae, suggesting that resistance of A. aegypti and C. tarsalis to anti-AgGluCl IgG found in blood meals is due to deficient IgG translocation across the midgut. We predicted that anti-AgGluCl IgG's mode of action is by antagonizing GluCl activity. To test this hypothesis, we fed A. gambiae blood meals containing anti-AgGluCl IgG and the GluCl agonist ivermectin (IVM). Anti-AgGluCl IgG attenuated the mosquitocidal effects of IVM, suggesting that anti-AgGluCl IgG antagonizes IVM-induced activation of GluCl. Lastly, we stained adult, female A. aegypti and C. tarsalis for GluCl expression. Neuronal GluCl expression in these mosquitoes was similar to previously reported A. gambiae GluCl expression; however, we also discovered GluCl staining on the basolateral surface of their midgut epithelial cells, suggesting important physiological differences in Culicine and Anopheline mosquitoes.
谷氨酸门控氯离子通道（GluCl）是阿维菌素类杀虫剂高度敏感的杀虫靶标。作为使用化学杀虫剂杀灭蚊子的替代方法，我们测试了靶向冈比亚按蚊（AgGluCl）GluCl胞外域的纯化免疫球蛋白G（IgG）对三种主要蚊媒病媒介蚊子存活的影响：冈比亚按蚊指名亚种、埃及伊蚊和致倦库蚊。通过单次血餐给药时，抗AgGluCl IgG以剂量依赖性方式降低了冈比亚按蚊的存活率（LC50：2.82 mg ml⁻¹，范围2.68 - 2.96 mg ml⁻¹），但对埃及伊蚊或致倦库蚊没有影响。我们之前证明AgGluCl仅位于冈比亚按蚊的头部和胸部组织中。为了验证AgGluCl IgG是否影响中肠外发现的靶抗原，我们通过胸腔内注射将其直接注入血腔。将单一的、生理相关浓度的抗AgGluCl IgG注入血腔同样降低了所有这三个物种蚊子的存活率。为了测试抗AgGluCl IgG是否进入这些蚊子中的每一种的血腔中，我们给蚊子喂食含有抗AgGluCl IgG的血餐，随后提取它们的血淋巴。我们仅在冈比亚按蚊的血淋巴中检测到IgG，这表明埃及伊蚊和致倦库蚊对血餐中发现的抗AgGluCl IgG的抗性是由于IgG跨中肠转运不足。我们预测抗AgGluCl IgG的作用方式是通过拮抗GluCl活性。为了验证这一假设，我们给冈比亚按蚊喂食含有抗AgGluCl IgG和GluCl激动剂伊维菌素（IVM）的血餐。抗AgGluCl IgG减弱了IVM的杀蚊效果，表明抗AgGluCl IgG拮抗IVM诱导的GluCl激活。最后，我们对成年雌性埃及伊蚊和致倦库蚊进行GluCl表达染色。这些蚊子中的神经元GluCl表达与先前报道的冈比亚按蚊GluCl表达相似；然而，我们还在它们中肠上皮细胞的基底外侧表面发现了GluCl染色，这表明库蚊属和按蚊属蚊子存在重要的生理差异。
