Dissecting the dynamics of virus-derived DNA of dengue virus 2 (DENV-2) in Aedes mosquitoes

Dengue is one of the neglected tropical diseases (NTDs) transmitted by Aedes mosquitoes and continues to spread globally. When mosquitoes are infected with dengue virus (DENV), virus-derived DNA (vDNA) is generated in mosquitoes, which subsequently contributes to their immune response. We traced the generation and presence of dengue virus type 2 (DENV-2) vDNA in experimentally infected cultured mosquito cells and Aedes mosquitoes, and notably, in wild mosquitoes collected in Burkina Faso. Detection of vDNA was achieved using a method incorporating loop-mediated isothermal amplification (LAMP), specifically, a LAMP-based vDNA detection method (vDNA-LAMP). The LAMP reaction, using primers targeting a segment of the NS5 region of DENV-2, detected vDNA from crude DNA extracted from experimentally infected cultured cells and Aedes mosquitoes. Detection revealed that the amount of DENV-2 vDNA generated in infected cells was relatively low; nevertheless, vDNA-LAMP enabled successful detection. The timing and quantity of vDNA generation in cultured cells were associated with the initial number of viral particles introduced during infection. Furthermore, vDNA-LAMP was applied to detect dengue virus vDNA in wild mosquitoes in dengue-endemic regions. This resulted in the successful detection of DENV-2 vDNA in field-collected mosquitoes, indicating that a proportion of wild mosquitoes in Burkina Faso harbored DENV-2 vDNA. Mapping these vDNA-positive mosquitoes allowed the identification of areas where infected mosquitoes and/or their progeny were likely present. These findings provide insights into the dynamics of DENV-2 vDNA in natural environments and underscore the potential of vDNA-LAMP as a tool for tracing vDNA in wild mosquitoes, which are responsible for transmitting viral infections.