High frequency of Voltage-gated sodium channel (VGSC) gene mutations in Aedes albopictus (Diptera: Culicidae) suggest rapid insecticide resistance evolution in Shanghai, China

Abstract

Background Dengue fever is an infectious disease that is imported into Shanghai, China and requires prevention and control measures. Controlling the vector Aedes albopictus through insecticide use is a key approach to dengue control. However, the rapid evolution of insecticide resistance in Ae. albopictus has raised concerns about the failure of dengue control efforts. Knockdown resistance (kdr) caused by point mutations in the voltage-gated sodium channel (VGSC) gene is a primary mechanism of pyrethroid resistance. In this study, we investigated the kdr mutations of Ae. albopictus in Shanghai and evaluated the trend in its evolution. Methodology/principal findings We collected 17 populations of Ae. albopictus from 15 districts in Shanghai in 2020, extracted genomic DNA from individual mosquitoes, and amplified Domain II, III, and IV in VGSC using PCR. Following sequencing, we obtained 658 VGSC sequences. We detected the nonsynonymous mutations V1016G, I1532T, and F1534S/C/I, among which V1016G and F1534C/I were reported in Shanghai for the first time and F1534I was a novel mutant allele in Ae. albopictus. The overall mutation frequency was 84.65%, with individual mutation frequencies ranging from 46.81% to 100%, excluding the Fengxian District population, which had a frequency of 0%. The V1016G and I1532T mutation types accounted for 7.14% and 3.42%, respectively. The mutant allele at codon 1534 accounted for 63.98% of all mutations, including TCC/S (62.77%), TGC/C (1.06%), and ATC/I (0.15%). We identified and classified five intron types in Domain III by length, including A (83 bp, 12.07%), B (68 bp, 87.30%), C (80 bp, 0.16%), D (72 bp, 0.16%), and E (70 bp, 0.31%). Individuals with intron B had a significant mutation tendency at codon 1534 relative to intron A (chi-square test, p < 0.0001). We found no correlation between mutation frequency and the amount of pyrethroid used (Pearson correlation, p = 0.4755). Conclusions/significance In recent years, kdr mutations in the Ae. albopictus population in Shanghai have rapidly evolved, as evidenced by an increase in mutation types and significantly increased mutation frequency. The F1534I/ATC mutant allele was found to be a novel mutation, F1534C/TGC was reported for the first time in Shanghai, and intron B in Domain III was significantly associated with mutation frequency at codon 1534. Continuous monitoring of resistance changes and strict regulation of insecticide use are required.

Diversity and distribution of sodium channel mutations inAedes albopictus(Diptera: Culicidae)

“…One of the variants of F1534L requires 2 substitutions from the wild-type state. Heterozygote 1534C/1534S individuals have also been found in China ( Wei et al 2021 , Zheng et al 2022 , Yuan et al 2023 ) along with 1534L/1534S ( Liu et al 2020 , Wei et al 2021 ) and the first report of phenylalanine to isoleucine mutation appearing as a heterozygote with 1534S (1534S/1534I) ( Yuan et al 2023 ). Italy is another focal point for studies in this area and 9 Vssc mutations have been recorded from Ae .…”

“…Functional studies have confirmed a causative function of this allele to pyrethroid resistance ( Yan et al 2020 , Guo et al 2022 ). The detection of F1534S (TCC) in China, the United States, and possibly Vietnam is of concern, suggesting that this resistant genotype ( Chen et al 2016 , Xu et al 2016 , Gao et al 2018 , Li et al 2018 , Wei et al 2021 , Wu et al 2021 , Guo et al 2022 , Yuan et al 2023 ) is already spreading. A recent study by Chen et al (2021) included historical samples and revealed the presence of the F1534S mutation (TCC) in Ae .…”

“…Synonymous mutations have been identified in V1016, I1532, and F1534 from China ( Chen et al 2016 , Zhou et al 2019 , Zheng et al 2022 , Yuan et al 2023 ) and Italy ( Pichler et al 2021 ), coding for the same amino acid as in wild-type mosquitoes. We identified the same phenomenon for V1016 from Malaysia, Singapore, and Vietnam.…”

“… a ATA also recorded as wild type by Zhou et al (2019) in Beijing, China. b TTT also recorded as wild type by Chen et al (2016) in Hainan Island, China and by Yuan et al (2023) in Shanghai, China. c GTG also recorded as wild type by Pichler et al (2021) and Zheng et al (2022) .…”

“… b TTT also recorded as wild type by Chen et al (2016) in Hainan Island, China and by Yuan et al (2023) in Shanghai, China. …”