| Summary: | Insect-specific flaviviruses (ISFs), although not known to be pathogenic to humans and animals, can modulate the transmission of arboviruses by mosquitoes. In this study, we screened 6665 host-seeking, gravid and blood-fed mosquitoes for infection with flaviviruses and assessed the vertebrate hosts of the blood-fed mosquitoes sampled in Baringo and Kajiado counties; both dryland ecosystem counties in the Kenyan Rift Valley. Sequence fragments of two ISFs were detected. Cuacua virus (CuCuV) was found in three blood-fed <i>Mansonia</i> (<i>Ma.</i>) <i>africana</i>. The genome was sequenced by next-generation sequencing (NGS), confirming 95.8% nucleotide sequence identity to CuCuV detected in <i>Mansonia</i> sp. in Mozambique. Sequence fragments of a potential novel ISF showing nucleotide identity of 72% to Aedes flavivirus virus were detected in individual blood-fed <i>Aedes aegypti</i>, <i>Anopheles gambiae</i> s.l., <i>Ma. africana</i> and <i>Culex</i> (<i>Cx.</i>) <i>univittatus</i>, all having fed on human blood. Blood-meal analysis revealed that the collected mosquitoes fed on diverse hosts, primarily humans and livestock, with a minor representation of wild mammals, amphibians and birds. The potential impact of the detected ISFs on arbovirus transmission requires further research.
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