Hepatitis B virus (HBV) consists of 9 major genotypes (A to I), one minor strain (designated J) and multiple subtypes, which may have different natural history, disease progression and treatment response. As only cell lines expressing genotype D3 have been established, here we aim to establish stable cell lines producing high-titer cell culture-generated HBV (HBVcc) of different genotypes and to explore their infectivity, virological features and treatment response.
Stable cell lines producing high titer of HBV with genotypes A2, B2, C1, E, F1b and H were generated by transfecting replication-competent 1.3×length HBV genome in a plasmid containing an antibiotic marker in HepG2 cells that can support HBV replication. Clones with highest levels of HBV DNA and/or HBeAg were selected and expanded for large-scale purification of HBVcc. HBVcc of different genotypes were tested in cells and humanized chimeric mouse model.
All HBVcc infected mouse-passaged primary human hepatocytes (PXB cells) and genotypes exhibit different responses to human IFN-α with variable kinetics of reduction in HBV DNA, HBeAg and HBsAg. HBVcc of all genotypes were infectious in humanized chimeric mice but with variable kinetics of viremia and viral antigen production. Treatment of infected mice with human IFN-α resulted in modest and variable reductions of viremia and viral antigenemia. HBVcc passaged in humanized chimeric mice (HBVmp) infected PXB cells much more efficiently than that of the original HBVcc viral stock.
Here we generate stable cell lines producing HBV of various genotypes that are infectious in vitro and in vivo. We observe genotype-associated variations in viral antigen production, infection kinetics and responses to human IFN-α treatment in these models.
Stable cell lines producing high-titer cell culture-generated HBV of various genotypes were established. HBV genotypes showed stable infectivity in both in vitro and in vivo models which are valuable tools for antiviral development.

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