Department of Parasite Disease Control and Prevention, Henan Center for Disease Control and Prevention, Zhengzhou, China.
Henan Provincial Medical Key Laboratory of Parasitic Pathogen and Vector, No. 105 South Agricultural Road, Zhengdong New District, Zhengzhou, 450016, China.
Parasit Vectors. 2023 Jun 2;16(1):180. doi: 10.1186/s13071-023-05796-z.
Historically, malaria due to Plasmodium vivax has been epidemic in Henan Province, China, with Anopheles sinensis as the main vector. The most effective measures to prevent malaria transmission are based on vector control through the use of insecticides. However, insecticides exert a strong selective pressure on mosquito populations for insecticide resistance. The aim of this study was to investigate the susceptibility profile and population genetic characteristics of An. sinensis to provide basic data and scientific guidance for the study of resistance mechanisms and the control of An. sinensis in Henan Province.
Adult Anopheles mosquitoes were collected at sites near local farmers' sheepfolds, pigsties and/or cowsheds located in Pingqiao, Xiangfu, Xiangcheng and Tanghe counties/districts of Henan Province during July-September 2021 for insecticide susceptibility testing. Molecular identification of collected mosquitoes as belonging to genus Anopheles was by PCR, and the frequencies of mutations in the knockdown resistance gene (kdr) and acetylcholinesterase-1 gene (ace-1) were detected using gene amplification. The mitochondrial DNA cytochrome oxidase subunit I (COI) gene was amplified in deltamethrin-resistant and deltamethrin-sensitive mosquitoes to analyze the genetic evolutionary relationship.
A total of 1409 Anopheles mosquitoes were identified by molecular identification, of which 1334 (94.68%) were An. sinensis, 28 (1.99%) were An. yatsushiroensis, 43 (3.05%) were An. anthropophagus and four (0.28%) were An. belenrae. The 24-h mortality rates of An. sinensis in Pingqiao, Tanghe, Xiangcheng and Xiangfu counties/districts exposed to deltamethrin were 85.85%, 25.38%, 29.73% and 7.66%, respectively; to beta-cyfluthrin, 36.24%, 70.91%, 34.33% and 3.28%, respectively; to propoxur, 68.39%, 80.60%, 37.62% and 9.29%, respectively; and to malathion, 97.43%, 97.67%, 99.21% and 64.23%, respectively. One mutation, G119S, was detected in the ace-1 gene. The frequencies of the main genotypes were 84.21% of specimens collected in Xiangfu (G/S), 90.63% of speciments collected in Xiangcheng (G/G) and 2.44% of speciments collected in Tanghe (S/S). Significantly higher G119S allele frequencies were observed in both propoxur- and malathion-resistant mosquitoes than in their sensitive counterparts in the Tanghe population (P < 0.05). Three mutations, L1014F (41.38%), L1014C (9.15%) and L1014W (0.12%), were detected in the kdr gene. The genotypes with the highest frequency in the populations of An. sinensis in Xiangfu and Tanghe were the mutant TTT (F/F) and wild-type TTG (L/L), at 67.86% (57/84) and 74.29% (52/70), respectively. In Pingqiao and Xiangfu, higher frequencies of the L1014F allele and lower frequencies of the L1014C allele were observed in mosquitoes resistant for beta-cyfluthrin than in those which were sensitive for this insecticide (P < 0.05). The results of Tajima's D and of Fu and Li's D and F were not significantly negative (P > 0.10), and each haplotype was interlaced and did not form two distinct branches.
High resistance to pyrethroids and propoxur was observed at four sites, but the resistance to malathion varied according to the location. Anopheles belenrae and the L1014W (TGG) mutation in An. sinensis were first discovered in Henan Province. The deltamethrin-resistant and deltamethrin-sensitive mosquito populations showed no genetic differentiation. The generation of resistance might be the result of the combination of multiple factors.
历史上，河南省疟疾流行，主要由间日疟原虫引起，媒介为中华按蚊。预防疟疾传播最有效的措施是通过杀虫剂控制蚊虫。然而，杀虫剂对蚊虫种群产生强烈的选择压力，导致抗药性。本研究旨在调查中华按蚊的敏感性特征和种群遗传特征，为河南省抗药性机制研究和中华按蚊的控制提供基础数据和科学指导。
2021 年 7 月至 9 月，在河南省平桥、襄县、项城和唐河县，在当地农民羊圈、猪圈和/或牛舍附近采集成蚊，进行杀虫剂敏感性测试。采用聚合酶链反应（PCR）对采集到的蚊子进行分类鉴定，通过基因扩增检测击倒抗性基因（kdr）和乙酰胆碱酯酶-1 基因（ace-1）的突变频率。在氯菊酯抗性和敏感的蚊子中扩增线粒体 DNA 细胞色素氧化酶亚基 I（COI）基因，分析遗传进化关系。
通过分子鉴定共鉴定出 1409 只按蚊，其中 1334 只（94.68%）为中华按蚊，28 只（1.99%）为嗜人按蚊，43 只（3.05%）为中华按蚊，4 只（0.28%）为雷氏按蚊。平桥、唐河、项城和襄县按蚊对氯菊酯的 24 小时死亡率分别为 85.85%、25.38%、29.73%和 7.66%，对溴氰菊酯分别为 36.24%、70.91%、34.33%和 3.28%，对残杀威分别为 68.39%、80.60%、37.62%和 9.29%，对马拉硫磷分别为 97.43%、97.67%、99.21%和 64.23%。在 ace-1 基因中检测到一个突变 G119S。主要基因型的频率分别为：襄县采集的标本（G/S）占 84.21%，项城采集的标本（G/G）占 90.63%，唐河采集的标本（S/S）占 2.44%。唐河县抗药性和敏感的残杀威种群中，G119S 等位基因频率明显较高（P<0.05）。在 kdr 基因中检测到 3 个突变 L1014F（41.38%）、L1014C（9.15%）和 L1014W（0.12%）。在襄县和唐河县的中华按蚊种群中，基因型频率最高的是突变型 TTT（F/F）和野生型 TTG（L/L），分别为 67.86%（57/84）和 74.29%（52/70）。在平桥和襄县，对溴氰菊酯抗性的蚊子中 L1014F 等位基因的频率较高，而对该杀虫剂敏感的蚊子中 L1014C 等位基因的频率较低（P<0.05）。Tajima's D 和 Fu 和 Li 的 D 和 F 的结果均不显著为负（P>0.10），每个单倍型相互交织，没有形成两个明显的分支。
在四个地点发现了对拟除虫菊酯和残杀威的高度抗性，但对马拉硫磷的抗性因地点而异。河南省首次发现了雷氏按蚊和中华按蚊的 L1014W（TGG）突变。氯菊酯抗性和敏感蚊虫种群无遗传分化。抗性的产生可能是多种因素共同作用的结果。
