Evolution of naturally occurring Hepatitis B Virus Test and Immune Escape mutations facilitating persistence and symptomatic viral flare in an HIV-coinfected patient

Aims: Due to its high rate of naturally occuring mutations, Hepatitis B Virus (HBV) is able to adapt rapidly to selective pressures. Mutations in the immunodominant A-determinant domain of the HBsAg are usually associated with immunoglobulin or vaccine administration. We identified an HIV-coinfected, chronically HBV-infected patient who tested HBsAg negative and developed an acute hepatitis associated with an HBV flare.

Methods: Sequential HBsAg-ELISA test escape isolates obtained before and during a flare of hepatitis in an HIV coinfected patient who tested negative for the HBsAg were analysed for their genotypic and phenotypic properties. The identified novel patterns of mutations in the immunodominant A-determinant domain of the HBsAg envelope gene were cloned into replication-competent HBV vectors and tested by transient transfection of human hepatoma cells for their replication efficacy.

Results: Phylogenetic analyses indicated the evolutionary relationship between both isolates. Next to a common amino-acid exchange (K122R) within the HBsAg A-determinant domain, the virus obtained during the flare showed 5 mutations (T118M, P120K, V167A, S173N, P213L), creating a T128K-exchange in the polymerase. The mutant present 8 years before showed G112R, P120T, F134S, I212L exchanges in the HBsAg, introducing R120K, T128N, Y221F mutations in the polymerase. Phenotypic analysis demonstrated a significantly reduced replication fitness of both HBV isolates, as intracapsid progeny DNA and virion release were diminished. However, both isolates expressed the HBs envelope gene resulting in regular cytoplasmatic HBs protein expression and reduced HBsAg secretion.

Conclusion: Immune escape strategies and not an enhanced viral replication efficacy gave these HBV mutants a selective advantage to persist in their host and to cause a symptomatic viral flare. Even without exogenous selective pressure immune escape mutants can evolve and might hamper detection in diagnostic assays as well as clearance by the immune system.

HIV - Hepatitis B Virus - a-determinant - chronic infection - vaccination