Levels of Resistance to Pyrethroid among DistinctkdrAlleles inAedes aegyptiLaboratory Lines and Frequency ofkdrAlleles in 27 Natural Populations from Rio de Janeiro, Brazil

Abstract

Background. Several mutations in voltage gated sodium channel (NaV) have been identified inAedes aegyptipopulations worldwide. However, only few are related to knockdown resistance to pyrethroids, most of which with variations in the 1016 and 1534NaVsites. In Brazil, at least twoNaValleles are known:NaVR1, with a substitution in the 1534 (1016 Val++ 1534 Ilekdr) andNaVR2, with substitutions in both 1016 and sites (1016Ilekdr+ 1534Cyskdr). There is also the duplication in theNaVgene, with one copy carrying the substitution Ile1011Met, although its effects on pyrethroid resistance remain to be clarified. Our goals in this study were (1) to determine the role of eachkdrNaVallele and the duplication on pyrethroid resistance and (2) to screen the frequency of thekdralleles in 27 several naturalAe. aegyptipopulations from the metropolitan region of Rio de Janeiro.Methods. Pyrethroid resistance was evaluated by a knockdown time (KdT) assay, an adaptation of the WHO test tubes with paper impregnated with deltamethrin. We used laboratory-selectedAe. aegyptilineages: R1R1 and R2R2 (homozygous for thekdrNaVR1 andNaVR2 alleles, respectively), Dup (with duplication in theNaVgene), Rockefeller (the susceptibility reference control), and F1 hybrids among them. Genotyping of both 1016 and 1534NaVsites was performed in 811Ae. aegyptisampled from 27 localities from Rio de Janeiro (17), Niterói (6) and Nova Iguaçu (4) cities, Rio de Janeiro State, Brazil, with a TaqMan real time PCR approach.Results. The laboratory lineages R1R1, R2R2, and R1R2 were the only ones that needed more than 60 minutes toknock downall the insects exposed to the pyrethroid, being theKdT R2R2 > R1R2 > R1R1, corroborating the recessive nature of thekdrmutations. Frequency ofkdrallelesNaVR1 andNaVR2 in field-caught mosquitoes varied from 0 to 52% and 43 to 86%, respectively, evidencing high levels of “resistant genotypes” (R1R1, R1R2, and R2R2), which together summed 60 to 100% inAe. aegyptipopulations from Rio de Janeiro.Conclusions. TheNaVR1 andNaVR2kdralleles confer resistance to the pyrethroid deltamethrin in homozygotes and R1R2 heterozygotes, being the R2R2 most resistant genotype. The allele containing duplication in theNaVgene, with a mutation in the 1011 site, did not confer resistance under the tested conditions. The frequencies of the “resistant genotypes” are elevated inAe. aegyptinatural populations from Rio de Janeiro.