Exploration of Culex mosquitoes virome through small-RNA sequencing reveals host-specific composition and novel viruses in Antioquia, Colombia

Culex mosquitoes are vectors of clinically relevant arboviruses, including West Nile virus, St. Louis encephalitis virus, and Japanese encephalitis virus. In addition, mosquitoes harbor insect-specific viruses (ISV) that can influence their biology and affect their ability to transmit arboviruses through mechanisms like superinfection exclusion and immunomodulation. In this study, we characterized the virome of Culex nigripalpus, Cx. coronator, and Cx. pipiens from two localities in the Urabá region of Antioquia, Colombia, using small RNA sequencing for viral detection. Six ISVs belonging to the Flaviviridae, Orthomyxoviridae, Rhabdoviridae, and Narnaviridae families were identified, along with two novel viruses of the Partitiviridae and Totiviridae families, expanding the known diversity of Culex-associated viromes. The impact of these identified viruses on host biology remains unknown. However, we detected a high frequency of Culex flavivirus (CxFV), which may affect vector competence. Wolbachia screening revealed its presence exclusively in Cx. pipiens, suggesting potential host-specific interactions. Viral alpha and beta diversity varied across Culex species and locations, highlighting ecological differences in mosquito-virus associations. All identified viruses were validated through qPCR, and viral fragments were confirmed via Sanger sequencing. This study advances our understanding of Culex virome diversity, contributing valuable insights into mosquito-virus associations and highlighting the importance of virome characterization in regions where vector-borne diseases are prevalent. Furthermore, this study lays the groundwork for improved surveillance strategies for the discovery and timely detection of emerging pathogenic viruses, which is critical for public health and force health protection.