LAUSR

Significance Wolbachia (wMel strain)-infected Aedes aegypti mosquitoes are refractory to disseminated arboviral infections. Yet previous studies into the mechanism behind Wolbachia-mediated virus blocking have not considered the involvement of lipids, apart from cholesterol, during superinfection. We used liquid chromatography mass spectrometry to study the lipidome in mosquito cells infected with virus, wMel, or superinfected with both virus and wMel. Interestingly, a class of lipids, acyl-carnitines increased during virus infection but remained low with wMel. These findings uncover a previously unknown role for acyl-carnitines in the interaction among virus, wMel, and cells, suggesting a mechanism underlying Wolbachia-mediated pathogen blocking. Importantly, this study supports Wolbachia introgression into A. Aegypti populations as a biocontrol method to reduce arboviral (e.g., DENV) transmission. Wolbachia-infected mosquitoes are refractory to flavivirus infections, but the role of lipids in Wolbachia-mediated virus blocking remains to be elucidated. Here, we use liquid chromatography mass spectrometry to provide a comprehensive picture of the lipidome of Aedes aegypti (Aag2) cells infected with Wolbachia only, either dengue or Zika virus only, and Wolbachia-infected Aag2 cells superinfected with either dengue or Zika virus. This approach identifies a class of lipids, acyl-carnitines, as being down-regulated during Wolbachia infection. Furthermore, treatment with an acyl-carnitine inhibitor assigns a crucial role for acyl-carnitines in the replication of dengue and Zika viruses. In contrast, depletion of acyl-carnitines increases Wolbachia density while addition of commercially available acyl-carnitines impairs Wolbachia production. Finally, we show an increase in flavivirus infection of Wolbachia-infected cells with the addition of acyl-carnitines. This study uncovers a previously unknown role for acyl-carnitines in this tripartite interaction that suggests an important and broad mechanism that underpins Wolbachia-mediated pathogen blocking.