Widespread occurrence of copy number variants and fixation of pyrethroid target site resistance in Anopheles gambiae (s.l.) from southern Côte d’Ivoire

Ruth M.A. Kouamé , Amy Lynd , Jackson K.I. Kouamé , Laura Vavassori , Kouabénan Abo , Martin J. Donnelly , Constant Edi , Eric Lucas
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Abstract

Resistance to pyrethroid and organophosphate insecticides in the malaria vector Anopheles gambiae (s.l.) is conferred by a variety of genetic mutations, including single nucleotide polymorphisms (SNPs) and copy number variants (CNVs). Knowledge of the distribution of these mutations in mosquito populations is a prerequisite for establishing better strategies for their management. In this study, a total of 755 Anopheles gambiae (s.l.) from southern Côte d’Ivoire were exposed to deltamethrin or pirimiphos-methyl insecticides and were screened to assess the distribution of SNPs and CNVs known or believed to confer resistance to one or other of the insecticide classes. Most individuals from the An. gambiae (s.l.) complex were identified by molecular tests as Anopheles coluzzii. Survival to deltamethrin (from 94% to 97%) was higher than to pirimiphos-methyl (from 10% to 49%). In An. gambiae (s.s.), the SNP in the Voltage Gated Sodium Channel (Vgsc) at the 995F locus (Vgsc-995F) was fixed, while other target site mutations were rare or absent (Vgsc-402L: 0%; Vgsc-1570Y: 0%, Acetylcholinesterase Acel-280S: 14%). In An. coluzzii, Vgsc-995F was the target site SNP found at highest frequency (65%) followed by other target site mutations (Vgsc-402L: 36%; Vgsc-1570Y: 0.33%; Acel-280S: 45%). The Vgsc-995S SNP was not present. The presence of the Ace1-280S SNP was found to be significantly linked to the presence of the Ace1-CNV, Ace1_AgDup. Significant association was found between the presence of the Ace1_AgDup and pirimiphos-methyl resistance in An. gambiae (s.s.) but not in An. coluzzii. The deletion Ace1_Del97 was found in one specimen of An. gambiae (s.s.). Four CNVs in the Cyp6aa/Cyp6p gene cluster, which contains genes of known importance for resistance, were detected in An. coluzzii, the most frequent being Dup 7 (42%) and Dup 14 (26%). While none of these individual CNV alleles were significantly associated with resistance, copy number in the Cyp6aa gene region in general was associated with increased resistance to deltamethrin. Elevated expression of Cyp6p3 was nearly associated with deltamethrin resistance, although there was no association of resistance with copy number. Use of alternative insecticides and control methods to arrest resistance spread in An. coluzzii populations is merited.

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Côte科特迪瓦南部冈比亚按蚊(s.l)拷贝数变异的广泛发生和拟除虫菊酯靶点抗性的固定
疟疾媒介冈比亚按蚊(s.l.)对拟除虫菊酯类和有机磷杀虫剂的耐药性是由多种基因突变引起的,包括单核苷酸多态性(SNPs)和拷贝数变异(CNVs)。了解这些突变在蚊子种群中的分布是制定更好的管理策略的先决条件。在这项研究中,共有755名来自科特迪瓦南部的冈比亚按蚊(s.l.)接触了溴氰菊酯或嘧啶米磷甲基杀虫剂,并对其进行了筛选,以评估已知或被认为会对一种或另一种杀虫剂产生耐药性的SNPs和CNVs的分布。冈比亚按蚊(s.l.)复合体中的大多数个体通过分子测试被鉴定为coluzzi按蚊。对溴氰菊酯的存活率(从94%到97%)高于对甲基吡嘧米磷的生存率(从10%到49%)。在冈比亚(s.s.)中,995F基因座(Vgsc-995F)的电压门控钠通道(Vgsc)中的SNP是固定的,而其他靶位点突变是罕见或不存在的(Vgsc-402L:0%;Vgsc-1570Y:0%,乙酰胆碱酯酶Acel-280S:14%)。在An.coluzzii中,Vgsc-995F是发现频率最高的靶位点SNP(65%),其次是其他靶位点突变(Vgsc-402L:36%;Vgsc-1570Y:0.33%;Acel-280S:45%)。Vgsc-995S SNP不存在。发现Ace1-280S SNP的存在与Ace1-CNV Ace1_AgDup的存在显著相关。在冈比亚(s.s.)中发现Ace1_AgDup的存在和吡瑞米磷的甲基抗性之间存在显著关联,但在coluzzii中没有。在冈比亚安的一个标本中发现了缺失Ace1_Del97。在coluzzi的Cyp6aa/Cyp6p基因簇中检测到四个CNV,其中包含已知对抗性具有重要意义的基因,最常见的是Dup 7(42%)和Dup 14(26%)。虽然这些单个CNV等位基因中没有一个与抗性显著相关,但Cyp6aa基因区域的拷贝数通常与对溴氰菊酯的抗性增加有关。Cyp6p3表达升高几乎与溴氰菊酯抗性有关,尽管抗性与拷贝数无关。使用替代杀虫剂和控制方法来阻止抗药性在珊瑚种群中的传播是值得的。
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