Midgut cell atlas of four important mosquito species and the key factors associated with DENV infection

Mosquitoes serve as medically significant vectors for transmitting various arboviruses. The mosquito midgut is both the digestive organ and primary infection site via viral bloodmeals, and the midgut cells show heterogeneity on virus-vector interactions. Rapidly developed single-cell RNA sequencing (scRNA-seq) technologies allow to characterize midgut physiology and viral infection dynamics at cellular solution. At present, scRNA-seq studies about mosquito midgut cell atlas are still limited and lack established protocols for cellular isolation. Therefore, we developed and employed a compatible microwell-based scRNA-seq protocol on four important mosquito species including Aedes aegypti, Aedes albopictus, Culex pipiens pallens and Culex tritaeniorhynchus and successfully established their midgut cell atlases pre- and post-bloodmeal. Further, we performed scRNA-seq experiments on DENV-infected Ae. aegypti to characterize the viral infection dynamics at single-cell solution. Overall design: To establish midgut cell atlas of four medically important mosquitoes including Aedes aegypti, Aedes albopictus, Culex pipiens pallens and Culex tritaeniorhynchus, we developed and applied a compatible microwell-based scRNA-seq protocol to the female mosquitoes. We performed scRNA-seq experiments on the midgut samples of the four mosquito species at 7 days with only glucose solution to establish their midgut cell atlas. To decipher the transcriptional changes following the uninfectious bloodmeal, we utilized scRNA-seq experiments on both Aedes aegypti and Culex tritaeniorhynchus, respectively. Further, to characterize the dynamics of mosquito midgut post-infection, we performed scRNA-seq experiments on the DENV-infected Aedes aegypti.