Investigation of putative multisubtype hepatitis C virus infections in vivo by heteroduplex mobility analysis of core/envelope subgenomes.

The frequency that multiple different subtypes of hepatitis C virus (HCV) simultaneously infect a given individual is controversial. To address this question, heteroduplex mobility analysis (HMA) of portions of the HCV core and envelope 1 region was optimized for sensitive and specific detection of mixtures of HCV genomes of different genotype or subtype. Using the standard HCV genotyping approach of 5'-untranslated region (UTR) analysis, 28 of 374 (7.5%) chronic hepatitis C research subjects were classified as having either multiple-subtype HCV infections (n = 21) or switching HCV subtypes over time (n = 7), the latter pattern implying viral superinfection. Upon retesting of specimens by HMA, 25 of 28 multiple-subtype results could not be reproduced. All three patients with positive results were injection drug users with potential multiple HCV exposures. To address the hypothesis of tissue sequestration of multiple-subtype HCV infections, liver (n = 22), peripheral blood mononuclear cell (n = 13), perihepatic lymph node (n = 16), and serum (n = 19) specimens from 23 subjects with end-stage hepatitis C were collected and analyzed by the HMA technique. Whereas 5'-UTR results implicated mixed-subtype HCV infections in 2 subjects, HMA testing revealed no evidence of a second HCV subtype in any tissue compartment (0 of 70 compartments [0%]) or within any given subject (0 of 23 subjects [0%]). In summary, a large proportion of mixed-genotype and switching-genotype patterns generated by 5'-UTR analysis were not reproducible using the HMA approach, emphasizing the need for additional study.

Hepatitis C virus dynamics during natural infection are associated with long-term histological outcome of chronic hepatitis C disease.

BACKGROUND: The long-term dynamics of hepatitis C virus (HCV) infection and their association with hepatitis C disease are unknown. METHODS: Fifty-two treatment-naive subjects with chronic HCV genotype 1 infection were selected from the Alaska Natives and American Indians cohort. Viral RNA levels were measured in 223 specimens (mean, 4.3 specimens/subject) over 457 patient-years. Viral quasispecies diversity was analyzed in 187 specimens (mean, 3.6 specimens/subject) over 365 patient-years. RESULTS: Thirty-three subjects had minimal hepatic fibrosis, and 19 developed bridging fibrosis or cirrhosis. There was no significant difference in host variables, including alcohol consumption, between disease groups. Subjects with mild disease had higher serum RNA levels after 2 decades of infection (P=.013), greater fluctuations in RNA levels over time (P=.04), higher intraspecimen quasispecies diversity (P=.001), and higher rates of quasispecies diversification (P=.004) than did subjects with severe disease. On multivariate analysis, the odds of having severe disease were 15.3 (95% confidence interval, 2.3-99.6) times higher among persons with low quasispecies diversification rates compared with the odds among persons with high diversification rates. CONCLUSIONS: Histological progression of hepatitis C is tightly associated with homogenization of HCV quasispecies, perhaps reflecting immune failure and/or selective outgrowth of aggressive viral variants.

High serum hepatitis C virus (HCV) RNA load predicts the presence of HCV RNA in saliva from individuals with chronic and acute HCV infection.

The detection of hepatitis C virus (HCV) in saliva was studied. Twenty-three subjects with chronic HCV infection and 1 subject with acute HCV infection were enrolled in a 21-day study. Roche COBAS Amplicor and Bayer VERSANT HCV RNA qualitative assays were used. For the 23 subjects with chronic HCV infection, 72% of 474 saliva samples were positive (or were imputed to be positive) for HCV RNA. Serum HCV RNA load predicted the detection of HCV RNA in saliva (odds ratio of 378.7 [95% confidence interval, 18.9-9996.6] for each additional log10 value). This association was also observed in 1 subject with acute HCV infection. Thus, our data demonstrate that salivary HCV RNA detection was associated with serum HCV RNA load in individuals who were chronically or acutely infected with HCV.

Strengths and limitations of commercial tests for hepatitis C virus RNA quantification.

The sensitivity of the Roche COBAS Monitor v.2.0 test was slightly better than that of the Bayer bDNA 3.0 test, but the Monitor test underquantified specimens by 2.5- to 10.6-fold even at relatively low hepatitis C virus RNA concentrations. Dilution prior to assay minimally decreased the reproducibility of the Monitor assay, leading to the recommendation for all specimens to be diluted 1:100 prior to testing by this assay.

Early hepatitis C viral kinetics correlate with long-term outcome in patients receiving high dose induction followed by combination interferon and ribavirin therapy.

BACKGROUND/AIMS: The majority of patients with genotype 1 do not respond to interferon (IFN) plus ribavirin. Limited data exist on the use of induction followed by combination therapy. METHODS: In this prospective study of 28 patients infected with genotype 1, randomization involved either daily or twice daily high dose IFN for 6 weeks, followed by standard therapy of 3 million units three times a week in combination with ribavirin for an additional 42 weeks. Hepatitis C virus (HCV) RNA was quantitated before and frequently during treatment. RESULTS: The best correlate of response was delta (the infected cell loss rate). Sixteen patients continued on the study because they had at least a 2 log drop in their HCV RNA levels by week 12; all but one were PCR negative for HCV RNA at 48 weeks, and 14 of these 16 patients continued to be PCR negative at 72 weeks. Both African-Americans in our trial failed to respond to therapy, and differences were evident during the induction phase. CONCLUSIONS: This randomized study of induction IFN therapy followed by combination IFN plus ribavirin yielded the highest rate of sustained response (50%) reported to date in chronically HCV-infected patients with genotype 1. The predictive value of the infected cell loss rate needs to be evaluated prospectively in larger studies, particularly in patients receiving pegylated IFN.