In-depth comparison of dengue virus laboratory strains and clinical isolates in terms of growth properties and NS1 production and detection in hepatic and non-hepatic cell lines.

Dengue virus (DV) is a major arboviral pathogen, with the liver being a primary target organ. The viral non-structural protein-1 (NS1) plays important roles in replication, autoimmunity, and pathogenesis, but strain-specific differences in NS1 secretion and cell responses remain incompletely understood. In this study, replication kinetics, NS1 production/detection, and apoptosis were examined in hepatocytes infected with laboratory strains and low-passage clinical isolates. All strains replicated efficiently, although laboratory strains showed slightly higher replication efficiency than clinical isolates. NS1 secretion patterns varied among strains, with predominant intracellular accumulation during infection, in contrast to NS1 plasmid-transfected cells where extracellular NS1 secretion was higher. Co-infection assays showed no viral interference, as individual serotypes maintained their replication profiles. Viral replication and NS1 secretion were reduced in non-hepatic cell lines, including Min6 and Vero cells, indicating cell type-dependent differences. Notably, NS1 from a DV1 laboratory strain was undetectable by NS1 antigen ELISAs despite robust replication. Site-directed mutagenesis identified a conserved leucine at position 237 as critical for NS1 detection, demonstrating that a single amino acid substitution (Leu237→Gln) can impair NS1 detectability. Both DV-infection and NS1-expression induced cleaved caspase-3 without apoptotic DNA fragmentation, allowing the virus to replicate longer using host cellular machinery. These findings highlight strain-dependent differences in NS1 biology with implications for dengue pathogenesis and diagnosis.