Hypervariable region 1 sequence stability during hepatitis C virus replication in chimpanzees

The putative envelope 2 (E2) gene of hepatitis C virus (HCV) contains a hig hly variable region referred to as hypervariable region 1 (HVR1). We hypoth esized that this genetic variability is driven by immune selection pressure , rather than representing the accumulation of random mutations in a region with relatively little functional constraint. To test this hypothesis, we examined the E2 sequence of a human inoculum that was passaged through eigh t chimpanzees, which appear to have a replicative rate (opportunity for cha nce mutation) similar to that of humans. Acute-phase plasma samples from a human (the inoculum) and six of eight serially infected chimpanzees were st udied. For each, 33 cloned cDNAs were examined by a combined hetero-duplex- single-stranded conformational polymorphism assay to assess quasispecies co mplexity and optimize selection of clones with unique gel shift patterns (c lonotypes) for sequencing. The sequence diversity of HCV was significantly lower in the chimpanzees than in the humans, and during eight serial passag es there was no change in the sequence of the majority clonotype from each animal examined. Similarly, the rates of protein sequence altering (nonsyno nymous) substitution were lower in the chimpanzees than in the humans. Thes e findings demonstrate that nonsynonymous mutations indicate selection pres sure rather than being an incidental result of HCV replication.