Genetic history of hepatitis C virus in East Asia

Abstract

The hepatitis C virus (HCV), which currently infects an estimated 3% of people worldwide, has been present in some human populations for several centuries, notably HCV genotypes 1 and 2 in West Africa and genotype 6 in Southeast Asia. Here we use newly developed methods of sequence analysis to conduct the first comprehensive investigation of the epidemic and evolutionary history of HCV in Asia. Our analysis includes new HCV core (n = 16) and NS5B (n = 14) gene sequences, obtained from serum samples of jaundiced patients from Laos. These exceptionally diverse isolates were analyzed in conjunction with all available reference strains using phylogenetic and Bayesian coalescent methods. We performed statistical tests of phylogeographic structure and applied a recently developed "relaxed molecular clock" approach to HCV for the first time, which indicated an unexpectedly high degree of rate variation. Our results reveal a >1,000-year-long development of genotype 6 in Asia, characterized by substantial phylogeographic structure and two distinct phases of epidemic history, before and during the 20th century. We conclude that HCV lineages representing preexisting and spatially restricted strains were involved in multiple, independent local epidemics during the 20th century. Our analysis explains the generation and maintenance of HCV diversity in Asia and could provide a template for further investigations of HCV spread in other regions.

FIG. 1.

A map of East Asia illustrating the geographic distribution of the NS5B gene data set (see also Fig. 3). Arrows indicate the countries of origin of the sequences and values in parentheses indicate the number of sequences from each country. Common subtypes present in each country (>20% of sequences) are shown in brackets. Note that the subtype distribution of NS5B sequences does not necessarily reflect the relative prevalence of infection by each subtype. Twelve isolates were sampled outside of Asia; available information indicates that at least seven of these were obtained from individuals of Asian origin.

FIG. 2.

Estimated maximum likelihood phylogeny for the core gene alignment. Bootstrap scores of >70% are shown next to well-supported nodes, and the phylogeny is midpoint rooted. Sequence names are given in the Los Alamos HCV Database format, as follows: subtype/country code/strain name/accession number. Where available, information is given in parentheses about the country of origin of emigrants from Southeast Asia to other countries. Gray bars indicate major subtypes of HCV genotype 6. Phylogeny branches and sequence names are colored according to the country of origin of the sampled individual. Country codes and colors for Asian strains are as follows: CN, China (red); VN, Vietnam (dark blue); HK, Hong Kong (red); TH, Thailand (green); IN, India (light blue); MM, Myanmar (gray); ID, Indonesia (brown); KH, Cambodia (orange); LA, Laos (magenta). Strains sampled outside of Asia are colored black (CA, Canada; FR, France; US, United States).

FIG. 3.

The estimated maximum likelihood phylogeny for the NS5B gene alignment. Bootstrap scores of >70% are shown next to well-supported nodes, and the phylogeny is midpoint rooted. See Fig. 2 for sequence naming details and the coloring scheme.

FIG. 4.

Estimated dates of origin of genotype 6, as obtained under model combinations A to F (see text for details). The composition and marginal posterior log likelihood of each model combination are provided. The error bars are the 95% highest posterior density credible intervals for each estimate.

FIG. 5.

The maximum clade support phylogeny of the concatenated (core plus NS5B) data set, obtained under the best-fitting model (combination E). Branch lengths represent time (see time scale at the bottom of the figure). Posterior probability scores (>0.9) are shown next to well-supported nodes. The shaded area corresponds to the 20th century, during which the lineages denoted by white circles showed rapid diversification. See Fig. 2 for sequence naming details.

FIG. 6.

Bayesian skyline plot, showing the epidemic history of genotype 6 estimated from the concatenated (core plus NS5B) data set (see text for details). The thick black line represents the estimated effective population size through time. The gray area represents the 95% highest posterior density confidence intervals for this estimate.