High voltage-gated sodium channel gene diversity in Aedes albopictus across Brazil

Abstract

The invasive Aedes albopictus, commonly known as the Asian tiger mosquito, has spread globally, posing public health risks because of its role as a secondary vector for arboviruses and capacity to transmit pathogens across sylvatic and urban cycles. In Brazil, where Ae. aegypti remains the primary vector of dengue, Zika, and chikungunya viruses; Ae. albopictus is being increasingly monitored because of its ecological plasticity and potential to develop insecticide resistance. Here, we analyzed the genetic diversity of voltage-gated sodium channel (Na_V) gene in Ae. albopictus populations across Brazil, in which knockdown resistance mutations (kdr) are associated with pyrethroid resistance. We collected Ae. albopictus from 46 Brazilian cities, extracted DNA from individual mosquitoes, and prepared pooled samples for next-generation sequencing. We targeted two Na_V segments, regions commonly associated with kdr in other mosquito species: IIS6 and IIIS6 segments. High-throughput sequencing and bioinformatics analysis were used to assess haplotype diversity, distribution, and phylogenetic relationships. We identified 20 IIS6 and 24 IIIS6 haplotypes, indicating high genetic diversity within the Na_V gene among Brazilian Ae. albopictus populations. No kdr mutations were detected despite the documented occurrence of these mutations in Ae. albopictus from other regions of the world. Nonetheless, we observed several synonymous polymorphisms, suggesting ancestral variation and potential for adaptive evolution. Our findings revealed substantial genetic diversity within the Na_V gene in Brazilian Ae. albopictus populations but no current evidence of pyrethroid resistance-associated kdr mutations. The observed diversity provides a foundation for tracking shifts in allele frequencies that may affect insecticide susceptibility and vector competence. Continuous monitoring of genetic variation is essential to preemptively address the development of resistance in Ae. albopictus and mitigate potential public health risks.

Keywords: Aedes albopictus; Asian tiger mosquito; Diversity; Molecular surveillance; Vector genetics.

Fig. 1

Nucleotide diversity of the voltage-gated sodium channel gene, IIS6 segment haplotypes, found in Aedes albopictus populations from Brazil. Geneious alignment among haplotypes found in this study (2s6.00-2s6.19), and consensus sequence generated by the software. Uppercase nucleotides correspond to the coding region (exons 20 and 21), while lowercase nucleotides refer to the intron, with alignment numbering at the top of each block. Invariable sites are indicated with dots, polymorphic sites with the alternative nucleotide, and gaps with (-). Codons in red represent codons 1016 where kdr mutations were previously described in Ae. albopictus. GenBank accession numbers of each haplotype are presented in Table 2.

Fig. 2

Phylogenetic relationship of the IIS6 haplotypes identified in Aedes albopictus populations from Brazil. Panel A–Maximum likelihood tree with bootstrap values exceeding 70%. Panel B–Alignment of intron 20, highlighting a region with greater sequence variation. Haplotypes were arranged in the same order as in the phylogenetic tree.

Fig. 3

Haplotype frequencies of IIS6 and IIIS6 segments in the Na_V gene of Aedes albopictus populations across Brazil. IIS6 (Panel A) and IIIS6 (Panel B) haplotype frequencies across populations and geographical regions. The populations are labelled with numbers matching the Brazilian map in Panel C.

Fig. 4

Nucleotide diversity in the voltage-gated sodium channel gene, IIIS6 segment, in Aedes albopictus populations from Brazil. Alignment among haplotypes found in this study (3s6.00-3s6.23) and consensus sequence generated by Geneious software. Uppercase nucleotides correspond to the coding region (Exons 30 and 31), while lowercase nucleotides refer to the intron, with alignment numbering at the top of each block, and primer position sequences underlined. Invariable sites are indicated with dots, polymorphic sites with the alternative nucleotide, and gaps with (-). Codons in red represent codons 1520, 1532, and 1534 where kdr mutations were found in Ae. albopictus from elsewhere. The Genbank accession numbers of each haplotype sequence are available in Table 2.

Fig. 5

Phylogenetic relationship of the IIIS6 haplotypes identified in Aedes albopictus populations from Brazil. Panel A–Maximum likelihood tree with bootstrap values exceeding 70%. Panel B–Alignment of intron 30, highlighting a region with greater sequence variation. Haplotypes were arranged in the same order as in the phylogenetic tree.

Fig. 6

Phased haplotypes of IIS6 and IIIS6 segments of the Na_V gene in Aedes albopictus populations from Brazil. Six distinct phasings were identified when at least one segment was monomorphic in a population. Bars represent alignments of the IIS6 and IIIS6 sequences, highlighting the polymorphisms. Synonymous codon changes are indicated.