Quantifying the Diversification of Hepatitis C Virus (HCV) during Primary Infection: Estimates of the In Vivo Mutation Rate

University of Texas at Austin, United States of America
PLoS Pathogens (Impact Factor: 7.56). 08/2012; 8(8):e1002881. DOI: 10.1371/journal.ppat.1002881
Source: PubMed


Hepatitis C virus (HCV) is present in the host with multiple variants generated by its error prone RNA-dependent RNA polymerase. Little is known about the initial viral diversification and the viral life cycle processes that influence diversity. We studied the diversification of HCV during acute infection in 17 plasma donors, with frequent sampling early in infection. To analyze these data, we developed a new stochastic model of the HCV life cycle. We found that the accumulation of mutations is surprisingly slow: at 30 days, the viral population on average is still 46% identical to its transmitted viral genome. Fitting the model to the sequence data, we estimate the median in vivo viral mutation rate is 2.5×10⁻⁵ mutations per nucleotide per genome replication (range 1.6-6.2×10⁻⁵), about 5-fold lower than previous estimates. To confirm these results we analyzed the frequency of stop codons (N = 10) among all possible non-sense mutation targets (M = 898,335), and found a mutation rate of 2.8-3.2×10⁻⁵, consistent with the estimate from the dynamical model. The slow accumulation of mutations is consistent with slow turnover of infected cells and replication complexes within infected cells. This slow turnover is also inferred from the viral load kinetics. Our estimated mutation rate, which is similar to that of other RNA viruses (e.g., HIV and influenza), is also compatible with the accumulation of substitutions seen in HCV at the population level. Our model identifies the relevant processes (long-lived cells and slow turnover of replication complexes) and parameters involved in determining the rate of HCV diversification.

Download full-text


Available from: Tanmoy Bhattacharya
  • Source
    • "Introduction of point mutations by the RNA polymerase is the primary element contributing to the high genetic variability of HCV. The HCV mutation rate in vivo is ~ 2.5 × 10 − 5 per nucleotide per genome replication (Ribeiro et al. 2012); however, higher estimates have also been reported (Cuevas et al. 2009). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatitis C virus (HCV) is a major public health problem that affects more than 180 million people worldwide. Identification of HCV transmission networks is of critical importance for disease control. HCV related cases are often difficult to identify due to the characteristic long incubation period and lack of symptoms during the acute phase of the disease, making it challenging to link related cases to a common source of infection. Additionally, HCV transmissions chains are difficult to trace back since viral variants from epidemiologically linked cases are genetically related but rarely identical. Genetic relatedness studies primarily rely on information obtained from the rapidly evolving HCV hypervariable region 1 (HVR1). However, in some instances, the rapid divergence of this region can lead to loss of genetic links between related isolates, which represents an important challenge for outbreak investigations and genetic relatedness studies. Sequencing of multiple and longer subgenomic regions has been proposed as an alternative to overcome the limitations imposed by the rapid molecular evolution of the HCV HVR1. Additionally, conventional molecular approaches required to characterize the HCV intrahost genetic variation are laborious, time-consuming, and expensive while providing limited information about the composition of the viral population. Next generation sequencing (NGS) approaches enormously facilitate the characterization of the HCV intrahost population by detecting rare variants at much lower frequencies. Thus, NGS approaches using multiple subgenomic regions should improve the characterization of the HCV intrahost population. Here, we explore the usefulness of multiregion sequencing using a NGS platform for genetic relatedness studies among HCV cases.
    Full-text · Article · Dec 2015 · Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases
  • Source
    • "HCV is a positive-strand, 1 Â 10 4 -nucleotide RNA virus that replicates through a negative-strand intermediate using a viral RNA-dependent RNA polymerase NS5B. The polymerase lacks error-correction and has an in vivo mutation rate of 2.5 Â 10 À 5 mutations per nucleotide per replication (Ribeiro et al., 2012). As HCV has a high replication rate, producing as many as 10 12 virions per day, one could predict that every position in the virus would be mutated daily with the certainty of multiple mutations among the genomes (Neumann et al., 1998). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Identifying HCV drug resistance mutations (DRMs) is increasingly important as new direct acting antiviral therapies (DAA) become available. Tagged pooled pyrosequencing (TPP) was originally developed as cost-effective approach for detecting low abundance HIV DRMs. Using 127 HCV-positive samples from a Canadian injection drug user cohort, we demonstrated the suitability and efficiency of TPP for evaluating DRMs in HCV NS5B gene. At a mutation identification threshold of 1%, no nucleoside inhibitor DRMs were detected among these DAA naïve subjects. Clinical NS5B resistance to non-nucleoside inhibitors and interferon/ribavirin was predicted to be low within this cohort. S282T mutation, the primary mutation selected by sofosbuvir in vitro, was not identified while S282G/C/R variants were detected in 9 subjects. Further characterization on these new S282 variants using in silico molecular modeling implied their potential association with resistance. Combining TPP with in silico analysis detects NS5B polymorphisms that may explain differences in treatment outcomes.
    Full-text · Article · Mar 2015 · Virology
  • Source
    • "HCV RNA-dependent RNA polymerase (RdRp) lacks proof-reading mechanisms which result in high error rates during replication (Moradpour et al., 2007). The mutation rate has been estimated to be $2.5 Â 10 À5 mutations per nucleotide per genome replication (Ribeiro et al., 2012). As a consequence, HCV is genetically heterogeneous and is represented by seven HCV genotypes and multiple subtypes (Smith et al., 2014). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatitis C virus (HCV) genotype 3a accounts for ∼80% of HCV infections in Pakistan, where ∼ 10 million people are HCV-infected. Here, we report analysis of the genetic heterogeneity of HCV NS3 and NS5b subgenomic regions from genotype 3a variants obtained from Pakistan. Phylogenetic analyses showed that Pakistani genotype 3a variants were as genetically diverse as global variants, with extensive intermixing. Bayesian estimates showed that the most recent ancestor for genotype 3a in Pakistan was last extant in ∼1896-1914 C.E. (range: 1851-1932). This genotype experienced a population expansion starting from ∼1905 until ∼1970 after which the effective population leveled. Death/birth models suggest that HCV 3a has reached saturating diversity with decreasing turnover rate and positive extinction. Taken together, these observations are consistent with a long and complex history of HCV 3a infection in Pakistan.
    Full-text · Article · Oct 2014 · Infection Genetics and Evolution
Show more