Evolution and adaptation of dengue virus in response to high temperature passaging in mosquito cells

Here, we passaged replicate virus populations ten times at either 26°C (Low) or 37°C (High) in C6/36 <i>Aedes albopictus</i> to explore the differences in adaptation to these thermal environments. We then deep-sequenced the resulting passaged virus populations and tested their replicative fitness in an all-cross temperature regime. We also assessed the ability of the passaged viruses to replicate in the insect vector. While viruses from both thermal regimes accumulated substitutions, only those reared in the 37°C treatments exhibited nonsynonymous changes, including several in the E, or envelop protein, and multiple non-structural genes. Passaging at the higher temperature, also led to reduced replicative ability at 26°C in both cells and mosquitoes. One of the mutations in the E gene involved the loss of a glycosylation site previously shown to reduce infectivity in the vector. These findings suggest that viruses selected for growth at higher ambient temperatures may experience tradeoffs between thermostability and replication in the vector. Data files include: "Virus Evolution- Serial Passage and Daily Titer Averages"- Results from Serial Passaging of virus at 37°C and 26°C. Contains data for the highest titer across passages for multiple replicates at 37°C and 26°C, and data for average viral titer across samples at 37°C and 26°C from daily samples taken. "Vector Competence Data"- Results from the vector competence experiment across 6 treatment groups for 6, 9, and 12 days post infection and 3 sample sources (Saliva, Midgut, and Carcass). All data was analyzed in GraphPad Prism, Version 10.