Large-scale metagenomic surveillance study expands the known diversity of RNA viruses in mosquito populations from the Amazon Basin

The Amazon Basin is one of the most biologically diverse regions on Earth, yet its viral diversity remains poorly characterized. Mosquitoes are important vectors and reservoirs of RNA viruses, but little is known about the composition and structure of their viromes in remote areas of the Amazon. In this study, we performed a large-scale metagenomics survey of RNA viruses associated with mosquito populations collected from the Jurua River region in the Western Amazon Basin of Brazil. We analyzed 211 pooled samples of adult female mosquitoes collected across thirty-seven sites, representing one of the most comprehensive mosquito virome studies conducted in this region to date. Utilizing high-throughput sequencing and <i>de novo</i> assembly, we identified over 500 viral sequences from 18 families, including 21 complete or nearly complete genomes. Our analysis revealed 18 putative novel viral species spanning diverse families and strains of nine previously described viruses. Phylogenetic analyses also revealed undocumented diversity within several virus families, including <i>Iflaviridae, Mesoniviridae, Phasmaviridae, Phenuiviridae, Togaviridae,</i> and <i>Totiviridae,</i> encompassing both novel species and previously known viruses detected for the first time in this region. Our findings highlight the immense, yet largely unexplored, diversity of RNA viruses circulating in mosquito populations in this ecologically rich but understudied region and provide critical insights into the evolutionary dynamics of mosquito-associated viruses. By leveraging high-throughput sequencing to uncover novel viral strains, this research demonstrates the value of metagenomic approaches in expanding the known diversity, distribution, and evolutionary relationships of RNA viruses, contributing to a broader understanding of virus-mosquito interactions and genome evolution.