Department of Parasitology, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand.
Trop Med Int Health. 2011 Apr;16(4):501-9. doi: 10.1111/j.1365-3156.2011.02725.x. Epub 2011 Feb 22.
To develop rapid monitoring tools to detect the F1534C permethrin-resistance mutation in domain IIIS6 of the Aedes aegypti voltage-gated sodium channel gene and determine the frequency and distribution of this mutation in Thailand.
A TaqMan SNP genotyping and an allele specific PCR (AS-PCR) assay were developed and validated by comparison with DNA sequencing of homozygous susceptible and homozygous resistant laboratory strains, their reciprocal-cross progenies, and field-caught mosquitoes. To determine the resistance phenotype of wild-caught A. aegypti, mosquitoes were exposed to 0.75% permethrin paper. The AS-PCR assay was used to screen 619 individuals from 20 localities throughout Thailand.
Overall, both assays gave results consistent with DNA sequencing for laboratory strains of known genotype and for wild-caught A. aegypti. The only slight discrepancy was for the AS-PCR method, which overestimated the mutant allele frequency by 1.8% in wild-caught samples. AS-PCR assays of permethrin-exposed samples show that the mutant C1534 allele is very closely associated with the resistant phenotype. However, 19 permethrin-resistant individuals were homozygous for the wild-type F1534 allele. DNA sequencing revealed all these individuals were homozygous for two other mutations in domain II, V1016G and S989P, which are known to confer resistance (Srisawat et al. 2010). The F1534C mutation is widespread in Thailand with mutant allele frequencies varying among populations from 0.20 to 1.00.
These assays can be used for the rapid detection of the F1534C resistance mutation in A. aegypti populations. The F1534C, and other, mutations underlie an extremely high prevalence of pyrethroid resistance in Thailand.
开发快速监测工具，以检测埃及伊蚊电压门控钠离子通道基因 IIIS6 区的 F1534C 氯菊酯抗性突变，并确定该突变在泰国的频率和分布。
开发了 TaqMan SNP 基因分型和等位基因特异性 PCR（AS-PCR）检测方法，并通过与纯合敏感和纯合抗性实验室品系、它们的回交后代以及野外捕获的蚊子的 DNA 测序进行比较进行了验证。为了确定野外捕获的埃及伊蚊的抗药性表型，将蚊子暴露在 0.75%氯菊酯纸上。使用 AS-PCR 检测方法筛选了来自泰国 20 个地区的 619 个人。
总体而言，两种检测方法的结果与已知基因型的实验室品系和野外捕获的埃及伊蚊的 DNA 测序结果一致。唯一的细微差异是 AS-PCR 方法，它在野外捕获的样本中高估了突变等位基因频率 1.8%。暴露于氯菊酯的样本的 AS-PCR 检测表明，突变 C1534 等位基因与抗性表型非常密切相关。然而，有 19 个氯菊酯抗性个体是野生型 F1534 等位基因的纯合子。DNA 测序显示，所有这些个体均为已知具有抗性的 II 域中的两个其他突变（V1016G 和 S989P）的纯合子（Srisawat 等人，2010 年）。F1534C 突变在泰国广泛存在，突变等位基因频率在不同种群中从 0.20 到 1.00 不等。
这些检测方法可用于快速检测埃及伊蚊种群中的 F1534C 抗性突变。F1534C 和其他突变是泰国极高的拟除虫菊酯抗性的基础。
