Prevalence and impact of occult hepatitis B infection in chronic hepatitis C patients treated with pegylated interferon and ribavirin.

The prevalence of occult hepatitis B, defined by absence of HBsAg and HBV DNA, ranges widely in patients with hepatitis C. This may influence the treatment of hepatitis C and the severity of liver disease. Sensitive and specific real-time PCR techniques are available commercially and can detect more reliably low HBV DNA levels. The aim of this study was to determine the prevalence of occult hepatitis B virus infection using the COBAS Taqman assay (Roche Diagnostics, Meylan, France) in the serum and liver of HBsAg negative patients with chronic hepatitis C and to evaluate its clinical consequences on liver pathology and its impact on the response to treatment with peg-IFNalpha and Ribavirin. HBV DNA detection was assessed retrospectively on 140 sera and 113 liver biopsies of HCV positive/HBsAg negative patients before treatment. A 4.4% (5/113) prevalence of occult hepatitis B was recorded in liver samples and in none of the sera. Anti-HBc was not detected in one, three of whom were sustained virological responders to treatment, one was relapsed responder and one was non-responder. Furthermore, in this cohort composed of 12% anti-HBs negative/anti-HBc positive and 20% anti-HBs positive/anti-HBc positive patients, anti-HBc was not associated with pre-therapeutic viral load, ALT serum levels, and histological activity or fibrosis. Using a commercial real-time PCR assay, we observed a low prevalence of occult B hepatitis. This, just as anti-HBC status, had no clinical impact in a large cohort of hepatitis C patients. It therefore does not appear useful to screen for occult hepatitis B in these patients with this test before beginning HCV treatment.

Inhibition of Epstein-Barr virus replication by small interfering RNA targeting the Epstein-Barr virus protease gene.

BACKGROUND: The Epstein-Barr virus (EBV) protease (PR), coded by the BVRF2 gene, is essential for the maturation of the viral capsid and viral DNA packaging during the late stage of the EBV lytic cycle. Like the other herpesvirus serine PRs, EBV PR could be a target for the inhibition of EBV replication. To date, no data have been reported on the inhibition of EBV PR messenger RNA (mRNA) by small interfering RNA (siRNA). METHODS: In this study, siRNAs targeting EBV PR were delivered to the epithelial 293 cell line stably transfected with the complete B95-8 EBV episome. EBV DNA and PR mRNA were quantified by real-time PCR in cells and supernatant, protein expression was assessed by immunoblotting, and production of EBV infectious particles in the culture medium was measured by Raji cell superinfection. RESULTS: The EBV PR mRNA within the cells was reduced by 73%, the PR protein by 35% and the amount of virus in the cell supernatant was drastically decreased by 86% or 95%, depending on the method. CONCLUSIONS: The strong effect of the siRNA targeting EBV PR on EBV replication attests to the crucial role played by EBV PR in the production of infectious particles and suggests that targeting this enzyme can be a new strategy against EBV-associated diseases where virus replication occurs.

Assessment of selective real-time PCR for quantitation of lamivudine and adefovir hepatitis B virus-resistant strains and comparison with direct sequencing and line probe assays.

A selective real-time PCR (sPCR) assay has been developed to detect the rtM204V/I and rtN236T mutations of hepatitis B virus (HBV) associated with resistance to lamivudine and adefovir. Using mixtures of mutant and wild-type plasmids, this sPCR was able to detect 0.1% of mutated strain in a total plasmid population of 10(5) copies and was more sensitive in detecting resistant strains than the line probe INNO-LiPA-DR-v2 assay and a direct sequencing assay. The comparison of these methods on 20 clinical specimens from treated patients confirmed the plasmid results: the three methods were concordant for the detection of the mutant strains in 72% of the cases and the discrepant results were caused mainly by the sequencing assay's lack of sensitivity. The line probe assay was more sensitive for detecting mutations than sPCR when the viral load was less than 10(4) copies/ml; conversely, the sPCR provided a more sensitive detection when the viral load was greater than 10(4) copies/ml. Although difficult to perform in clinical practice, sPCR appears to be a reliable technique for detecting and quantifying quasi-species resistant to lamivudine (LAM) and adefovir (ADV) and can be useful to gain a better understanding of the natural history of antiviral resistance during the treatment of chronic hepatitis B (CHB).

Sustained virological and biochemical responses to lamivudine and adefovir dipivoxil combination in a chronic hepatitis B infection despite mutations conferring resistance to both drugs.

BACKGROUND: Sequential monotherapies of nucleotide analogs used in chronic hepatitis B treatment can lead to the selection of a resistance mutation to each antiviral drug. CASE PRESENTATION: A patient with chronic hepatitis B was successively treated with lamivudine monotherapy, lamivudine-adefovir dual therapy, adefovir monotherapy and again with an adefovir-lamivudine dual therapy. Lamivudine-associated mutations (rtL180M and rtM204V/I) followed by adefovir-associated mutations (rtN236T and rtA181V) emerged during the two monotherapy regimens. Despite the presence of rtM204V/I, rtA181V, and rtN236T mutations at the beginning of the second dual therapy, sustained biochemical and virological responses have been observed thus far after 23 months. CONCLUSION: This case illustrates that rtM204V/I, rtA181V, and rtN236T resistance mutations can coexist in a patient but do not preclude the recycling of lamivudine and adefovir in combination therapy, when no other therapeutic choices are available.

A prospective follow-up of Epstein-Barr virus LMP1 genotypes in saliva and blood during infectious mononucleosis.

To monitor multiple Epstein-Barr virus (EBV) infections during the early and convalescent stages of infectious mononucleosis (IM), a cloning and sequencing study of the LMP1 gene was conducted in saliva and peripheral blood mononuclear cells (PBMCs) from 23 patients with IM at day 0 (D0) and day 180 (D180) after the onset of the disease. Multiple EBV strains were detected in 9 (39%) of the patients during follow-up, with 7 of 9 cases detected as early as D0. Six of the nine patients harbored the same dominant strain in saliva and PBMCs during follow-up, with a trend toward a restriction of the number of EBV strains in saliva but not in PBMCs at D180. Furthermore, transmission of a minor strain was observed between partners in a heterosexual couple. There was no correlation between multiple infections and EBV DNA load in either compartment.

Long-term shedding of infectious epstein-barr virus after infectious mononucleosis.

Epstein-Barr virus (EBV) DNA loads in peripheral blood mononuclear cells (PBMCs), plasma, and saliva, as well as infectivity of the virus in saliva, were evaluated in 20 patients for 6 months after the onset of infectious mononucleosis (IM). All patients displayed sustained high EBV DNA loads in the saliva, associated with a persistent infectivity of saliva at day 180. EBV DNA load in PBMCs decreased significantly from day 0 to day 180 (in spite of a viral rebound between day 30 and day 90 in 90% of the patients), and EBV DNA rapidly disappeared from plasma. These data show that patients with IM remain highly infectious during convalescence.