Frequency of sodium channel genotypes and association with pyrethrum knockdown time in populations of Californian Aedes aegypti

Abstract

Background Since their detection in 2013, Aedes aegypti has become a widespread urban pest in California. The availability of cryptic larval breeding sites in residential areas and resistance to insecticides pose significant challenges to control efforts. Resistance to pyrethroids is largely attributed to mutations in the voltage gated sodium channels (VGSC), the pyrethroid site of action. However, past studies have indicated that VGSC mutations may not be entirely predictive of the observed resistance phenotype. Methods To investigate the frequencies of VGSC mutations and the relationship with pyrethroid insecticide resistance in California, we sampled Ae. aegypti from four locations in the Central Valley, and the Greater Los Angeles area. Mosquitoes from each location were subjected to an individual pyrethrum bottle bioassay to determine knockdown times. A subset of assayed mosquitoes from each location was then analyzed to determine the composition of 5 single nucleotide polymorphism (SNP) loci within the VGSC gene. Results The distribution of knockdown times for each of the five Californian populations sampled was non-parametric with potentially bimodal distributions. One group succumbs to insecticidal effects around 35–45 min and the second group lasts up to and beyond the termination of the assay (120+ min). We detected 5 polymorphic VGSC SNPs within the sampled California populations. One is potentially new and alternatively spliced (I915K), and four are documented and associated with resistance: F1534C, V1016I, V410L and S723T. The Central Valley populations (Clovis, Dinuba, Sanger and Kingsburg) are fairly homogenous with only 5% of the mosquitoes showing heterozygosity at any given position. In the Greater LA mosquitoes, 55% had at least one susceptible allele at any of the five SNP loci. The known resistance allele F1534C was detected in almost all sampled mosquitoes (99.4%). We also observe significant heterogeneity in the knockdown phenotypes of individuals with the identical VGSC haplotypes suggesting the presence of additional undefined resistance mechanisms. Conclusions Resistance associated VGSC SNPs are prevalent, particularly in the Central Valley. Interestingly, among mosquitoes carrying all 4 resistance associated SNPs, we observe significant heterogeneity in bottle bioassay profiles suggesting that other mechanisms are important to the individual resistance of Ae. aegypti in California. Keywords: Aedes aegypti, Resistance, Pyrethroid, IPLEX genotyping, Voltage gated sodium channel, California.

Evidence of Local Extinction and Reintroduction of Aedes aegypti in Exeter, California

“…aegypti had different frequencies of nonsynonymous SNPs or mutations in the VGSC gene than in Central CA populations, including the ubiquitous V410L mutation that is known to confer resistance to pyrethroids (Table 2). If all six of the mutations in the VGSC gene included in this study confer some levels of resistance to pyrethroids (kdr resistant alleles) then Southern CA populations have a higher proportion of susceptible alleles relative to central CA populations which is consistent with a previous report (13). Although Exeter 2014 and 2018 samples had different overall genetic profiles, they both contained a greater number of pyrethroid susceptible kdr alleles than the Clovis population.…”

“…We also analyzed 6 SNPs in the voltage-gated sodium channel gene, typically known as knockdown resistance gene (kdr) in mosquito literature (AAEL023266) located on Chromosome 3 using the method described in Mack et al (13). Five of these 6 SNPs were previously used in Mack et al (13).…”

“…We also analyzed 6 SNPs in the voltage-gated sodium channel gene, typically known as knockdown resistance gene (kdr) in mosquito literature (AAEL023266) located on Chromosome 3 using the method described in Mack et al (13). Five of these 6 SNPs were previously used in Mack et al (13). The SNP Q1853R (3:315931672) is a new addition to this study as it was only detected in the mosquitoes derived from the Exeter 2014 population.…”

“…Thirty-seven SNPs were selected from the published whole genome sequences (8), 29 of which were chosen due to their association with specific genetic clusters and eight within the Voltage Gated Sodium Channel gene. Data for 5 knockdown resistance (kdr) SNPs (F1534C, V410L, S723T, I915K, and V1016I) were included from (13). A full list of SNPs and primers can be found in Table S1.…”

“…Knockdown resistance (kdr) mutation genotypes per population.Clovis 2013 data is genome sequencing data. Clovis 2018 data and Greater Los Angeles (LA) 2018 data are from Mack et al(13). S stands for susceptible (reference) allele and R for resistant (alternate) allele.…”