Metatranscriptomic insights into mosquitoes and sand flies from Colombia reveal extensive virome variability and detection of bacterial and viral-related resistomes

Mosquitoes and sand flies are primary vectors of a wide range of pathogenic microorganisms that affect human and animal health worldwide. Their microbial communities directly affect their vector capacity and have become a focus of research for the development of novel biocontrol strategies. Moreover, these microbial communities may harbor antimicrobial resistance genes (ARGs), a growing public health concern. Despite their relevance, the viral and bacterial compositions of several vector species in epidemiologically relevant regions in Colombia have not been characterized. In this study, the virome, metatranscriptomic bacteriome, and ARG profiles were characterized using metatranscriptomic analysis of mosquitoes and sand flies of the genera Aedes , Culex , Psorophora , Coquillettidia , Lutzomyia , and Psychodopygus. Specimens were collected from rural, peri-urban, and urban environments across seven locations in the Caribbean and Amazon regions of Colombia. RNA-seq libraries were generated and analyzed to identify viral sequences, transcriptionally active bacterial taxa, and ARGs associated with the insect microbiome. We identified 416 viral sequences across 23 RNA-seq libraries from mosquito and sandfly hosts. The viromes include sequences related to 13 families, including Metaviridae , Chuviridae , Xinmoviridae , Flaviviridae , and Rhabdoviridae . Metatranscriptomic analyses also revealed complex active bacterial microbiomes, with significant presence of Wolbachia in Culex , Coquillettidia and Aedes albopictus . The presence of other lower-abundance bacterial genera associated with humans, including Cutibacterium , Faecalibacterium , Prevotella , Hallela , and Escherichia , may reflect exposure to human-impacted environments. Additionally, the resistome included genes linked to resistance to fluoroquinolones, aminoglycosides, and tetracyclines, as well as instances of multidrug resistance associated with efflux pump systems. The presence of ARGs in sequences associated with mobile genetic elements suggests a potential role for horizontal gene transfer within the insect microbiome. Our results confirm that the viromes of mosquitoes and sand flies in Colombia are mainly composed of insect-specific viruses (ISVs), and their structures are not exclusively dependent on the host species. Notably, the bacterial fraction of the microbiome highlights the active role of both symbionts and other taxa from environmental sources. We identified transcriptionally active ARGs in hematophagous insects, highlighting their potential relevance for AMR surveillance. Further studies incorporating environmental matrices and functional validation are needed to assess this potential.