The NS4B A119T mutation of dengue virus 2 confers adaptation to mammalian cells and increased pathogenicity in AG129 mice

Mouse-adapted dengue virus type 2 (DENV2), designated the AG-P10 strain, was established through serial blind passages between AG129 mice and mosquito C6/36 cells. Compared with the parental strain INDIA/09-74, the AG-P10 strain exhibited increased virulence in AG129 mice, leading to rapid body weight loss and earlier mortality. In vitro characterization of the AG-P10 strain demonstrated significantly enhanced viral replication and larger focus formation in mammalian Vero and A549 cells but not in mosquito-derived C6/36 cells, suggesting host-specific adaptation. Whole-genome sequencing of the AG-P10 strain identified two nonsynonymous mutations, A119T in NS4B and E802Q in NS5. Through an analysis of recombinant viruses, we determined that the A119T mutation alone is sufficient to confer enhanced pathogenicity in AG129 mice and increased viral replication in Vero and A549 cells, suggesting its potential role in viral adaptation and pathogenesis. The DENV2 with the A119T mutation, as well as the AG-P10 strain, thus provides a robust animal model for developing antiviral therapeutics and vaccines against DENV and for investigating the molecular mechanisms governing DENV2 replication and virulence.