湖北省武汉市居民区白纹伊蚊电压门控钠离子通道基因突变分析

Analysis of voltage-gated sodium channel gene mutations in Aedes albopictus in the residential area of Wuhan,Hubei province,China

目的了解武汉市居民区生境白纹伊蚊种群电压门控钠离子通道(VGSC)基因突变情况,初步探索该市白纹伊蚊抗药性机制。方法2021年8-10月在武汉市居民区生境采集白纹伊蚊幼虫和蛹,于实验室饲养至成蚊,抽提单只成蚊的基因组DNA,PCR扩增VGSC基因部分片段,测序后分析VGSC基因型分布情况,并采用χ^(2)检验对雌雄蚊在VGSC基因各突变位点基因型分布进行比较分析。结果共检测武汉市居民区生境的白纹伊蚊238只,其中雌蚊126只,雄蚊112只。武汉市白纹伊蚊种群在VGSC基因V1016、I1532与F1534位点存在突变。V1016位点有2种等位基因,分别为野生型GTA/V(43.91%)和突变型GGA/G(56.09%);3种基因型分别为野生型纯合子V/V(19.33%)、野生/突变型杂合子V/G(49.16%)和突变型纯合子G/G(31.51%)。I1532位点有2种等位基因,分别为野生型ATC/I(99.16%)和突变型ACC/T(0.84%);2种基因型分别为野生型纯合子I/I(98.32%)和野生/突变型杂合子I/T(1.68%)。F1534位点发现3种等位基因,分别为野生型TTC/F(60.72%)、突变型TCC/S(33.19%)、TGC/C(6.09%);5种基因型分别为野生型纯合子F/F(35.30%)、野生/突变型杂合子F/S(41.60%)和F/C(9.24%)、突变型纯合子S/S(10.92%)与突变型杂合子S/C(2.94%)。VGSC基因D1763位点未检测到突变,仅存在野生型GAC/D(100%)等位基因。不同性别白纹伊蚊个体间VGSC基因V1016、I1532与F1534位点的基因型分布差异均无统计学意义(χ^(2)=0.198,P=0.656;χ^(2)=0.014,P=0.905;χ^(2)=2.210,P=0.137)。结论武汉市白纹伊蚊VGSC基因突变频率较高,可能是该市白纹伊蚊对拟除虫菊酯类杀虫剂抗药性迅速发展的重要机制之一。雌、雄蚊VGSC基因突变无明显差异。

Objective To investigate the mutations in the voltage-gated sodium channel(VGSC)gene of Aedes albopictus in the residential habitats of Wuhan,Hubei province,China,and to preliminarily explore the resistance development mechanism of Ae.albopictus in Wuhan.Methods Larvae and pupae of Ae.albopictus were collected from the residential habitats of Wuhan from August to October 2021 and raised to adults in the laboratory.Genomic DNA was extracted form each adult mosquito.The partial VGSC gene fragments were amplified by the polymerase chain reaction and sequenced to analyze the distribution of VGSC genotypes.The genotype distribution at each mutation site of the VGSC gene were compared for female and male mosquitoes by the Chi-square test.Results A total of 238 Ae.albopictus from the residential habitats of Wuhan,including 126 female mosquitoes and 112 male mosquitoes,were analyzed.Mutations were detected at the V1016,I1532,and F1534 loci of the VGSC gene.There were two alleles at the V1016 locus,namely wild-type GTA/V(43.91%)and mutant GGA/G(56.09%),and three genotypes,i.e.,the wild-type homozygote V/V(19.33%),wild/mutant heterozygote V/G(49.16%),and mutant homozygote G/G(31.51%).There were two alleles at the I1532 locus,namely wild-type ATC/I(99.16%)and mutant ACC/T(0.84%),and two genotypes,i.e.,wild-type homozygote I/I(98.32%)and wild/mutant heterozygote I/T(1.68%).At the F1534 locus,three alleles,i.e.,wild-type TTC/F(60.72%),mutant TCC/S(33.19%),and TGC/C(6.09%),and five genotypes,i.e.,wild-type homozygote F/F(35.30%),wild/mutant heterozygote F/S(41.60%)and F/C(9.24%),mutant homozygote S/S(10.92%),and mutant heterozygote S/C(2.94%),were detected.No mutant allele was found at the D1763 locus of the VGSC gene,at which only the wild-type allele GAC/D was detected(100%).There was no significant difference in the genotype distribution at the V1016,I1532,and F1534 loci of the VGSC gene between different sexes of Ae.albopictus(χ^(2)=0.198,P=0.656;χ^(2)=0.014,P=0.905;χ^(2)=2.210,P=0.137).Conclusions The rate of VGSC gene mutation in Ae.albopictus is high in Wuhan,which might be one of the important mechanisms of Ae.albopictus rapidly developing resistance towards pyrethroid insecticides in Wuhan.Anyway,no significant difference was found in VGSC gene mutations between female and male mosquitoes.