背景:房子飞,家蝇,是可能影响公众健康的疾病的重要载体。反复使用拟除虫菊酯杀虫剂可能是家蝇电压敏感钠通道(VSSC)中突变和氨基酸取代的选择压力,这最终导致了抵抗。这项研究的目的是使用分子工具确定击倒抗性(kdr)突变的存在,并建立针对阿拉伯联合酋长国(UAE)家蝇的CDC瓶生物测定法,以筛选溴氰菊酯抗性。
方法:从阿布扎比的19个地点收集成年苍蝇,阿联酋,提取DNA,随后进行特定等位基因的PCR扩增(PASA)和使用几种引物的常规PCR以扩增VSSC基因的区域。对PCR产物进行Sanger测序。我们还设计了在逆转录酶(RT)-PCR中使用互补DNA(cDNA)检测四个kdr突变的引物,然后进行Sanger测序。此外,建立了CDC瓶生物测定法,用于检测成年家蝇中的溴氰菊酯抗性。
结果:在PASA中,引物成功扩增目标条带(480、280和200bp)。在19个地点中的18个地点收集的苍蝇中发现了kdr等位基因,患病率最高和最低,分别为46.9%和9.4%,分别。抗性纯合(RR)昆虫占测试种群的5.0%,杂合(RS)昆虫占36.5%。RR基因型在19个采样位置中的10个采集的家蝇中普遍存在。家蝇种群大多处于哈代-温伯格平衡状态,除了三个地方。除了验证先前鉴定的kdr突变L1014F的存在,在这项研究中,我们检测到两个kdr突变,L1014H和T929I,以前在阿联酋没有报道过。此外,第一次在阿联酋,使用CDC瓶生物测定溴氰菊酯抗性,发现60分钟和4.5微克/毫升是诊断时间和剂量,分别。使用这种方法,我们在16个地方中的两个地方的家蝇中检测到溴氰菊酯抗性,阻力水平为12.5%。
结论:使用DNA测序,我们证实了一个已知的kdr突变的存在,并在阿布扎比的家蝇中发现了两个新的kdr突变。此外,我们使用CDC瓶生物测定法检测了这些果蝇的溴氰菊酯抗性。建议进一步研究以全面识别阿联酋家蝇种群中的更多kdr突变,并评估其对控制策略的影响。
BACKGROUND: The house fly, Musca domestica, is a significant carrier of diseases that can impact public health. Repeated use of pyrethroid insecticides may act as a selection pressure for mutations and amino acid substitutions in the house fly voltage-sensitive sodium channel (VSSC), which ultimately confers resistance. The objectives of this study were to determine the presence of knockdown resistance (
kdr) mutations using molecular tools and to set up a CDC bottle bioassay specific for house flies in the United Arab Emirates (UAE) to screen for deltamethrin resistance.
METHODS: Adult flies were collected from 19 locations in Abu Dhabi, UAE, and DNA was extracted, followed by PCR amplification of specific alleles (PASA) and conventional PCR using several primers to amplify regions of the VSSC gene. Sanger sequencing was performed on PCR products. We also designed primers that detect four kdr mutations using complementary DNA (cDNA) in reverse transcriptase (RT)-PCR followed by Sanger sequencing. Additionally, a CDC bottle bioassay was set up for detecting deltamethrin resistance in adult house flies.
RESULTS: In PASA, the primers successfully amplified the target bands (480, 280 and 200 bp). The
kdr allele was found in flies collected from 18 of the 19 locations, at the highest and lowest prevalence of 46.9% and 9.4%, respectively. Resistant homozygous (RR) insects constituted 5.0% of the tested populations, and heterozygous (RS) insects accounted for 36.5%. The RR genotype was prevalent in house flies collected at 10 of 19 sampling locations. House fly populations were mostly in Hardy-Weinberg equilibrium, except in three locations. In addition to verifying the presence of the previously identified
kdr mutation L1014F, in this study we detected two
kdr mutations, L1014H and T929I, that have not previously been reported in the UAE. Also, for the first time in the UAE, a CDC bottle bioassay for deltamethrin resistance was used, which found that 60 min and 4.5 µg/ml were the diagnostic time and dose, respectively. Using this assay, we detected deltamethrin resistance in house flies from two of 16 locations, with a resistance level of 12.5%.
CONCLUSIONS: Using DNA sequencing, we confirmed the presence of a known kdr mutation and uncovered two new
kdr mutations in house flies from Abu Dhabi. Additionally, we detected deltamethrin resistance in these flies using a CDC bottle bioassay. Further research is recommended to comprehensively identify more kdr mutations in UAE house fly populations and assess their impacts on control strategies.