Abstract
Arboviruses can intensify epidemics by acquiring single nucleotide variants, leading to clade replacement and severe outbreaks. We investigated dengue virus serotype 2 evolution in consecutive outbreaks from 2001 to 2003 in Taiwan, coinciding with overwintering and increased epidemic severity. The virus evolved from the early-epidemic strain (Ia) to the late-epidemic strains (Ib and II), featuring three amino acid differences. The later strains demonstrated increased replication at lower temperatures, and the NS5V357E mutation significantly boosts virus replication and virulence, regardless of the other two mutations (ET46I and NS5I271T). Crucially, the late NS5V357E signature swiftly emerged after infecting mosquitos with the early Ia strain, through thoracic injection or by feeding on Ia-infected mice. Thus, we discover the molecular events involved in overwintering and increased disease severity between consecutive dengue outbreaks. This study enhances our understanding of dengue epidemiology, aiding in predicting and monitoring the emergence of dengue strains with increased epidemic potential.