Hepatitis C Virus Quasispecies Diversity Research Articles

Quasispecies Diversity Is a Major Risk Factor for Vertical Hepatitis C Virus Transmission.

Vertical transmission is the major cause of pediatric hepatitis C virus (HCV) infection. The objective of this study was to better understand HCV pathogenesis in pregnant women and provide insights into risk factors and mechanisms involved in vertical transmission. Evolutionary dynamics of HCV variant spectra and HCV-specific neutralizing antibody responses were examined using high-throughput sequencing and pseudoparticle-based assays in pregnant women monoinfected with HCV (n = 17) or coinfected with HCV and human immunodeficiency virus (HIV)-1 (n = 15). Overall, statistically significant associations were found between HCV quasispecies diversity, selective pressure exerted on the HCV E2 envelope protein, and neutralizing activity of maternal immunoglobulins. Women with low quasispecies diversity displayed significantly higher mean aspartate aminotransferase and alanine aminotransferase levels throughout pregnancy, but this difference was restricted to monoinfected participants. Low quasispecies diversity and inefficient neutralizing activity were also significantly associated with vertical transmission, but only in the monoinfected group. These results indicate that maternal neutralizing antibody responses play a role in the prevention of vertical HCV transmission, but not in presence of HIV-1 coinfection, and suggest that the mechanism of vertical transmission may be different between monoinfected and coinfected women. These findings could inform management strategies for the prevention of vertical HCV transmission.

Divergent quasispecies evolution in de novo hepatitis C virus infection associated with bone marrow transplantation

Quasispecies is a remarkable characteristic of hepatitis C virus (HCV) and has profound roles in HCV biology and clinical practice. The understanding of HCV quasispecies behavior, in particular in acute HCV infection, is valuable for vaccine development and therapeutic interference. However, acute HCV infection is seldom encountered in clinic practice due to its silent onset. In the present study, we reported a unique case of de novo HCV infection associated with the transplantation of bone marrow from a HCV-positive donor. HCV quasispecies diversity was determined in both the donor and the recipient over a 4-year follow-up, accompanied with simultaneous measurement of HCV neutralizing antibody. Detailed genetic and phylogenetic analyses revealed a divergent quasispecies evolution, which was not related to dynamic changes of HCV neutralizing antibody. Instead, our data suggested an essential role of the fitness adaptation of founder viral population in driving such an evolutionary pattern.

Changes of HCV quasispecies during combination therapy with interferon and ribavirin

Treatment of chronic hepatitis C virus (HCV) infection with interferon (IFN) and ribavirin improves the rate of eradication of HCV, but only about 13–14% of non-responders (NR) with HCV of genotype 1b previously treated with IFN achieve a sustained virological response (SVR). To determine whether HCV quasispecies diversity correlates with the outcome of therapy with IFN and ribavirin, we studied 13 patients undergoing combination therapy with IFN-α2b and ribavirin after failure of IFN monotherapy. HCV quasispecies diversity was assessed by cloning and sequencing before and during combination therapy. During therapy, quasispecies diversity diminished in NR patients, both in the hypervariable region (HVR) 1 of the envelope 2 (E2) domain and in the interferon sensitivity-determining region (ISDR) in the NS5A. Pre-treatment nucleotide quasispecies diversity was lower in SVR and end-of-therapy viral response (ETR) patients than in NR patients. Resistance to ribavirin was associated with high pre-treatment heterogeneity and the selection of quasispecies of the HCV genome. HVR quasispecies may be a predictor of efficacy of combination therapy with IFN and ribavirin.

Changes of HCV quasispecies during combination therapy with interferon and ribavirin.

Treatment of chronic hepatitis C virus (HCV) infection with interferon (IFN) and ribavirin improves the rate of eradication of HCV, but only about 13-14% of non-responders (NR) with HCV of genotype 1b previously treated with IFN achieve a sustained virological response (SVR). To determine whether HCV quasispecies diversity correlates with the outcome of therapy with IFN and ribavirin, we studied 13 patients undergoing combination therapy with IFN-alpha2b and ribavirin after failure of IFN monotherapy. HCV quasispecies diversity was assessed by cloning and sequencing before and during combination therapy. During therapy, quasispecies diversity diminished in NR patients, both in the hypervariable region (HVR) 1 of the envelope 2 (E2) domain and in the interferon sensitivity-determining region (ISDR) in the NS5A. Pre-treatment nucleotide quasispecies diversity was lower in SVR and end-of-therapy viral response (ETR) patients than in NR patients. Resistance to ribavirin was associated with high pre-treatment heterogeneity and the selection of quasispecies of the HCV genome. HVR quasispecies may be a predictor of efficacy of combination therapy with IFN and ribavirin.

Impact of highly active antiretroviral therapy and immunologic status on hepatitis C virus quasispecies diversity in human immunodeficiency virus/hepatitis C virus-coinfected patients.

This study analyzes the effect of highly active antiretroviral therapy (HAART), and thus immunologic status, on hepatitis C virus (HCV) load and quasispecies diversity in patients coinfected with the human immunodeficiency virus (HIV) and HCV. Three cohorts of coinfected patients were analyzed retrospectively over a period of 7 to 10 months: group A was antiretroviral drug naïve at baseline and then on HAART for the remainder of the study, group B did not receive antiretroviral therapy at any point, and group C was on HAART for the entire study. HCV quasispecies diversity was analyzed by sequencing hypervariable region 1. In a longitudinal analysis, there was no significant change from baseline in any immunologic, virologic, or quasispecies parameter in any of the three groups. However, in comparison to groups A and B, group C had significantly higher CD4+- and CD8+-cell counts, a trend toward a higher HCV load, and significantly increased number of HCV clones, entropy, genetic distance, and ratio of nonsynonymous substitutions per nonsynonymous site to synonymous substitutions per synonymous site (Ka/Ks). In addition, CD4+-cell count was positively correlated with HCV load, genetic distance, and Ka. Interestingly, patients infected with HCV genotype 2 or 3 had a significantly higher CD4+-cell count, HCV load, genetic distance, and Ka/Ks than those infected with genotype 1. These results suggest that there is no immediate effect of HAART on HCV but that, with prolonged HAART, immune restoration results in an increase in HCV load and quasispecies diversity.

The NS5a gene of hepatitis C virus in patients treated with interferon-?

Patients infected with hepatitis C virus (HCV) genotype 3 have a better response to interferon-alpha (IFN-alpha) therapy than those infected with genotype 1. There are extensive sequence differences between genotypes in the 3' half of the NS5a gene. An association between IFN-alpha response and the interferon sensitivity-determining region (ISDR) (amino acids 2209-2248) of HCV genotype 1b has been described [Enomoto et al. (1996) New England Journal of Medicine 334:771-776]. A prospective study was conducted to determine whether the derived NS5A amino acid sequence or quasi-species diversity could predict response to IFN-alpha therapy. Serum samples were obtained before, during, and after treatment from 35 IFN-alpha-treated patients chronically infected with HCV (eight with type1b,13 with type1a, and 14 with type3a). Nucleotide sequences were determined, and amino acid sequences corresponding to residues 2178-2390 of the polyprotein were derived. Quasi-species complexity was analysed by amplification of the ISDR region (2270-2403), followed by single-stranded conformation polymorphism (SSCP). No amino acid sequence that could be used to predict response to treatment was found, and there was no selection of specific amino acid residues during treatment. A striking lack of variability was seen in HCV genotype 3a, but the small degree of variation could suggest an effect on response. SSCP showed that variation in the predominant NS5a sequence occurred in the presence and absence of therapeutically administered IFN-alpha. HCV quasi-species diversity pretreatment did not predict IFN-alpha treatment outcome. The conclusion of the study is that the amino acid sequence of NS5a cannot be used to predict the efficacy of treatment with IFN-alpha in HCV-infected patients in Scotland. No evidence was found to support the selection of IFN-alpha-resistant strains in the NS5a gene.

INCREASED PREOPERATIVE QUASISPECIES DIVERSITY MAY BE PREDICTIVE OF LONG-TERM OUTCOME AFTER ORTHOTOPIC LIVER TRANSPLANTATION FOR HEPATITIS C VIRUS

277 Purpose: Recurrence of hepatitis C virus (HCV), the leading indication for orthotopic liver transplantation (OLT), accounts for a significant proportion of graft dysfunction. At present, predicting recurrence is difficult. Viral factors may be predictive of outcome. Therefore, we correlated HCV amino acid sequence diversity at the first hypervariable region (HVR1) of the second envelope protein (E2) with long-term outcome. Methods: Twenty-four patients with HCV-induced end-stage liver disease who underwent liver transplantation between 10/95 to 12/98 were prospectively evaluated. Recurrence of significant hepatitis was defined by pathology and biochemical parameters. Twelve representative patients underwent detailed pre- and post-operative serum quasispecies analysis by nucleotide sequencing the HVR1 of cloned PCR products. Results: A summary of results is below (mean ± SEM): (Table)TableMean follow up was 612±25 days. All patients had viremia within 2 months after OLT. Biochemical recurrence occurred at 169±55 days. There was no difference between groups in ischemic time, HLA matching, donor and recipient age, HCV quantity, use of CSA or FK506, gender, or CMV status. One death occurred (17%, P=NS) in the recurrent hepatitis group. Eighty-three percent of patients with asymptomatic viremia had a predominant major variant (accounting for >50% of clones) compared to 17% in those with recurrence (P<0.025). In summary, patients with significant recurrent hepatitis had a greater preoperative diversity of HCV quasispecies in the HVR1 that did not persist in the post-operative period. Conclusions: Increased HCV quasispecies diversity may be predictive of recurrent HCV-induced hepatitis after OLT for HCV. This diversity did not persist postoperatively.