Mutation Rate of the Hepadnavirus Genome

An essential factor for charting the evolution of hepadnaviruses is an estimation of the mutation rate of the virus genome during replication in the host. In order to determine the mutation rate of the hepadnavirus genome under defined experimental conditions, we transfected 10 neonatal woodchucks with an infectious molecular clone of woodchuck hepatitis virus (WHV). By 4 months post-transfection, all 10 animals showed serological evidence for WHV infection. Subsequently, 1 animal became chronically infected and was used for further study. At 16 months post-transfection WHV DNA from serum virions was cloned and the nucleotide sequence of three independent progeny genomes compared directly with that of the input recombinant DNA. Although the consensus nucleotide sequence remained unchanged, we found three differences in individual progeny genomes when compared to the parental genome sequence. Thus, we estimate the mutation rate of the WHV genome to be less than or equal to 2 X 10(-4) base substitutions/site/year. This figure is one to two orders of magnitude lower than the mutation rates previously calculated for the positive- and negative-strand RNA viruses, but is similar to the mutation rate of the gag gene which is the most slowly evolving gene of retroviruses. Therefore, we find that the hepadnavirus genome is relatively stable during replication in host tissues when compared to other viruses that lack polymerase-associated proofreading functions.