Knockdown resistance mutations distribution and characteristics of Aedes albopictus field populations within eleven dengue local epidemic provinces in China
Authors: Zhao Chunchun , Zhou Xinxin , Xue Chuizhao , Lun Xinchang , Li Wenyu , Liu Xiaobo , Wu Haixia , Song Xiuping , Wang Jun , Liu Qiyong , Meng Fengxia 
Published: February 2023

Background

Aedes albopictus, commonly known as the tiger mosquito, has attracted global attention because its bite can transmit several viruses, such as dengue virus. With the absence of an effective therapy and vaccine, mosquito control is the sole method for dengue fever control. However, Ae. albopictus has developed resistance to most insecticides, especially pyrethroids. Many scholars have conducted thorough research for the target-site of pyrethroids. The main target-site is the voltage-gated sodium channel gene (VGSC) whose mutation causes knockdown resistance (kdr). The spatial distribution of three locus kdr mutations in Ae. albopictus has not been comprehensively analyzed nationwide in China. In addition, the relationship between the frequency of kdr mutations and dengue fever has not yet been explored.

Methods

A total of 2,241 Ae. albopictus samples from 49 populations from 11 provinces of mainland China were collected in 2020 and analyzed for mutations in the VGSC gene. DNAstar 7.1. Seqman and Mega-X were used to compare the sequences and read the peak map to confirm the genotypes and alleles of each mutation. ArcGIS 10.6 software was used to make interpolation and extract meteorological data of collection sites and to conduct spatial autocorrelation analysis. R 4.1.2 software was used to conduct a chi-square test for kdr mutations and dengue area and to analyze the correlation between meteorological factors and kdr mutations.

Results

The overall frequencies of mutant alleles at 1016G, 1532T, and 1534S/C/L were 13.19%, 4.89%, and 46.90%, respectively. Mutations at the three loci were found at 89.80% (44/49), 44.90% (22/49), and 97.96% (48/49) of the field populations. At each of the loci V1016 and I1532, only one allele was detected, which was GGA(G) and ACC(T), respectively. Five mutant alleles were found at codon 1534: TCC/S (33.49%), TGC/C (11.96%), TTG/L (0.60%), CTC/L (0.49%), and TTA/L (0.58%). In total, 31 triple-locus genotype combinations were found, and the single locus mutation was the most common. We also found firstly triple-locus mutant individuals, whose genotypes were V/G+I/T+F/S and V/G+I/T+S/S. The 1016 and 1532 mutation rates were significantly negatively related to the annual average temperature (AAT), but the 1534 mutation rate was significantly positively related to AAT. The 1532 mutation rate was significantly positively related to the 1016 mutation rate but negatively related to the 1534 mutation rate. A relationship was observed between the 1534 codon mutation rate and dengue epidemic areas in this study. Furthermore, spatial autocorrelation analysis results showed that the mutation rates of different codons in different geographical areas had spatial aggregation and positive spatial correlation.

Conclusion

This study showed that the multiple kdr mutations at codon 1016, 1532 and 1534 of Ae. albopictus were found in most areas of China. Two novel triple-locus genotype combinations, V/G+I/T+F/S and V/G+I/T+S/S, were detected in this study. In addition, the relationship between mosquito resistance and dengue fever outbreak should be further explored, especially considering the insecticide-usage history in different areas. The characteristic of spatial aggregation of VGSC gene mutation rates reminds us to notice the gene exchange and similarity of insecticide usage in the adjacent areas. The use of pyrethroids should be restricted to delay resistance development. New-type insecticides should be developed to adjust the changes in the resistance spectrum. Our study provides abundant data on the Ae. albopictus kdr gene mutation in China; these findings will be useful for the correlation analysis of molecular mechanism of insecticide resistance.