Department of Tropical Diseases, Faculty of Naval Medicine, Naval Medical University, Shanghai, 200433, China.
Yunnan Institute of Parasitic Diseases, Yunnan Provincial Key Laboratory of Vector-Borne Diseases Control and Research, Yunnan Provincial Center of Malaria Research, Yunnan Provincial Collaborative Innovation Center for Public Health and Disease Prevention and Control, Yunnan Institute of Parasitic Diseases Innovative Team of Key Techniques for Vector Borne Disease Control and Prevention (Developing), Pu'er, 665099, China.
Parasit Vectors. 2022 Jun 26;15(1):229. doi: 10.1186/s13071-022-05366-9.
Vector control is still a pivotal method for preventing malaria, and its potency is weakened by the increasing resistance of vectors to chemical insecticides. As the most abundant and vital malaria vector in Southeast Asia, the chemical insecticide resistance status in Anopheles sinensis remains elusive in Laos, which makes it imperative to evaluate the true nature of chemical insecticide resistance-associated genetic mutations in An. sinensis in Laos.
Adult An. sinensis were collected from three border regions in Laos. DNA was extracted from individual mosquitoes. PCR amplification and DNA sequencing of a fragment containing codon 1014 of the voltage-gated sodium channel (vgsc) gene were completed to study the kdr allele frequency distribution, kdr intron polymorphism, population genetic diversity, and the evolutionary status of the kdr codon. The mitochondrial cytochrome c oxidase subunit II gene (COII) was amplified and sequenced to examine population variations, genetic differentiation, spatial population structure, population expansion, and gene flow patterns.
Nine wild kdr haplotypes of the vgsc gene were detected in this study, and eight of them, namely 1014L1, 1014L2, 1014L4, 1014L7, 1014L9, 1014L10, 1014L11, and 1014L21, were discovered in the China-Laos border (northern Laos), while 1014L3 was only detected in the Thailand-Laos border (northwestern Laos) and Cambodia-Laos border (southern Laos). The newly identified haplotype, 1014L21, was uniquely distributed in the China-Laos border and was not identified in other countries. Based on sequence analysis of the mitochondrial COII genes, significant genetic differentiation and limited gene flow were detected between the China-Laos and Cambodia-Laos An. sinensis populations, which suggested that those two regions were genetically isolated. The distinct distribution of the kdr haplotype frequencies is probably the result of geographical isolation in mosquito populations.
Lack of kdr mutations in the vgsc gene was probably due to genetic isolation and the absence of intense selection pressure in the three border regions of Laos. This study reveals that pyrethroid-based chemical insecticides are still appropriate for battling An. sinensis in parts of Laos, and routine monitoring of chemical insecticide resistance should be continuously implemented and focused on more restricted areas as part of chemical insecticide resistance management.
病媒控制仍然是预防疟疾的关键方法，但由于媒介对化学杀虫剂的抗药性不断增强，其效果减弱。作为东南亚最丰富和最重要的疟疾媒介，老挝对中华按蚊的化学杀虫剂抗药性状况仍然难以捉摸，因此必须评估老挝中华按蚊化学杀虫剂抗性相关遗传突变的真实性质。
从老挝三个边境地区采集成年中华按蚊。从个体蚊子中提取 DNA。完成包含电压门控钠离子通道（vgsc）基因密码子 1014 的片段的 PCR 扩增和 DNA 测序，以研究 kdr 等位基因频率分布、kdr 内含子多态性、种群遗传多样性和 kdr 密码子的进化状态。扩增并测序线粒体细胞色素 c 氧化酶亚单位 II 基因（COII），以检查种群变异、遗传分化、空间种群结构、种群扩张和基因流动模式。
本研究共检测到 vgsc 基因的 9 种野生 kdr 单倍型，其中 8 种（1014L1、1014L2、1014L4、1014L7、1014L9、1014L10、1014L11 和 1014L21）在中国-老挝边境（老挝北部）发现，而 1014L3 仅在泰国-老挝边境（老挝西北部）和柬埔寨-老挝边境（老挝南部）发现。新发现的单倍型 1014L21 仅分布在中国-老挝边境，在其他国家未发现。基于线粒体 COII 基因序列分析，中国-老挝和柬埔寨-老挝中华按蚊种群之间存在显著的遗传分化和有限的基因流，表明这两个地区在遗传上是隔离的。kdr 单倍型频率的明显分布可能是蚊群地理隔离的结果。
老挝三个边境地区 vgsc 基因中缺乏 kdr 突变可能是由于遗传隔离和缺乏强烈的选择压力。本研究表明，在老挝部分地区，基于拟除虫菊酯的化学杀虫剂仍然适用于对抗中华按蚊，应持续进行化学杀虫剂抗性常规监测，并集中在更受限制的地区，作为化学杀虫剂抗性管理的一部分。
