RNA-Dependent RNA Polymerase of the Second Human Pegivirus Exhibits a High-Fidelity Feature

The virus-encoded RNA-dependent RNA polymerase (RdRp) is responsible for viral replication, and its fidelity is closely related to viral diversity, pathogenesis, virulence, and fitness. Hepatitis C virus (HCV) and the second human pegivirus (HPgV-2) belong to the family Flaviviridae and share some features, including similar viral genome structure. Unlike HCV, HPgV-2 preserves a highly conserved genome sequence and low intrahost variation. However, the underlying mechanism remains to be elucidated. In this study, we evaluated the fidelity of HPgV-2 and HCV RdRp in an in vitro RNA polymerase reaction system. The results showed higher fidelity of HPgV-2 RdRp than HCV NS5B with respect to the misincorporation rate due to their difference in recognizing nucleoside triphosphate (NTP) substrates. Furthermore, HPgV-2 RdRp showed lower sensitivity than HCV to sofosbuvir, a nucleotide inhibitor against HCV RdRp, which explained the insusceptibility of HPgV-2 to direct-acting antiviral (DAA) therapy against HCV infection. Our results indicate that HPgV-2 could be an excellent model for studying the mechanisms involved in viral polymerase fidelity as well as RNA virus diversity and evolution. IMPORTANCE RNA viruses represent the most important pathogens for humans and animals and exhibit rapid evolution and high adaptive capacity, which is due to the high mutation rates for using the error-prone RNA-dependent RNA polymerase (RdRp) during replication. The fidelity of RdRp is closely associated with viral diversity, fitness, and pathogenesis. Previous studies have shown that the second human pegivirus (HPgV-2) exhibits a highly conserved genome sequence and low intrahost variation, which might be due to the fidelity of HPgV-2 RdRp. In this work, we used a series of in vitro RNA polymerase assays to evaluate the in vitro fidelity of HPgV-2 RdRp and compared it with that of HCV RdRp. The results indicated that HPgV-2 RdRp preserves significantly higher fidelity than HCV RdRp, which might contribute to the conservation of the HPgV-2 genome. The unique feature of HPgV-2 RdRp fidelity provides a new model for investigation of viral RdRp fidelity.