Lentiviral backbone-based hepatitis B virus replicon-mediated transfer favours the establishment of persistent hepatitis B virus infection in mice after hydrodynamic injection

• We herein describe an alternative HDI method for the study of HBV persistence in a murine model.
• Lentiviral backbone-based transduction favours the persistent hepatitis B virus infection in mice.
• Novel HBSS1-based vaccination in mice induces robust immunity.
• The clearance of the viremia and reduction of circulating antigen levels in mice was observed after HBSS1 vaccination.

Establishment of a non-transgenic mouse model of persistent hepatitis B virus (HBV) infection is urgently needed. In this study, we constructed novel lentiviral-transfer plasmids containing HBV replicon DNA (pCS-HBV1.3, containing a 1.3-fold-overlength genome of HBV) and employed hydrodynamic injection (HDI) to develop an HBV-persistent mouse model. We explored the impact of host (different mouse strains, BALB/c and C57BL/6), gender, and the plasmid backbone on persistent HBV in mice. Our data showed that HBV antigenaemia (HBsAg, HBeAg) and HBV DNA persisted for >56 days post-injection, while the appearance of anti-HBs antibody in the serum was only found among <30% of female C57BL/6 mice injected with pCS-HBV1.3. Moreover, HBcAg and HBV DNA were also detected in the liver of HDI mice. Compared with previous AAV-backbone based HBV replicon DNA transfer, we found that the HDI transfer with the lentiviral vector-based HBV replicon (pCS-HBV1.3) in this study resulted in a significantly higher level of HBV DNA transfer in the liver and longer persistence of HBV DNA and antigenaemia in the serum. Furthermore, we also showed that immunization of HBV replicon transfer mice with the novel HBSS1-based vaccines was able to overcome tolerance against HBV in mice and induces robust immunity (humoral as well as T-cell responses), followed by the clearance of the HBV viremia. We concluded that lentiviral backbone-based transfer vectors more readily establish persistent HBV infection in mouse models via HDI, providing a new tool useful for the study of HBV infection and immune-based therapies.