Cellular immune responses associated with occult hepatitis C virus infection of the liver

Occult hepatitis C virus (HCV) infection is a type of recently identified chronic infection that is evidenced only by detection of HCV RNA in liver; patients consistently test negative for antibodies to HCV and HCV RNA in serum. Using ex vivo and in vitro measures of T-cell responses, we have identified functional virus-specific memory CD4(+) and CD8(+) T cells in the peripheral blood of patients with occult HCV infection. The features of the virus-specific T cells were consistent with immune surveillance functions, supporting previous exposure to HCV. In addition, the magnitudes of CD4(+) and CD8(+) T-cell responses were in parallel and correlated inversely with the extent of liver HCV infection. The detection of HCV-specific T cells in individuals in whom HCV RNA can persist in the liver despite the absence of viremia and antibodies indicates that HCV replication is prolonged in the face of virus-specific CD4(+) and CD8(+) T-cell responses. These findings demonstrate that HCV-specific cellular immune responses are markers not only of previous exposure to and recovery from HCV but also of ongoing occult HCV infection.     

Inhibition of hepatitis C virus-like particle binding to target cells by antiviral antibodies in acute and chronic hepatitis C

Hepatitis C virus (HCV) is a leading cause of chronic viral hepatitis worldwide. The study of antibody-mediated virus neutralization has been hampered by the lack of an efficient and high-throughput cell culture system for the study of virus neutralization. The HCV structural proteins have been shown to assemble into noninfectious HCV-like particles (HCV-LPs). Similar to serum-derived virions, HCV-LPs bind and enter human hepatocytes and hepatoma cell lines. In this study, we developed an HCV-LP-based model system for a systematic functional analysis of antiviral antibodies from patients with acute or chronic hepatitis C. We demonstrate that cellular HCV-LP binding was specifically inhibited by antiviral antibodies from patients with acute or chronic hepatitis C in a dose-dependent manner. Using a library of homologous overlapping envelope peptides covering the entire HCV envelope, we identified an epitope in the N-terminal E2 region (SQKIQLVNTNGSWHI; amino acid positions 408 to 422) as one target of human antiviral antibodies inhibiting cellular particle binding. Using a large panel of serum samples from patients with acute and chronic hepatitis C, we demonstrated that the presence of antibodies with inhibition of binding activity was not associated with viral clearance. In conclusion, antibody-mediated inhibition of cellular HCV-LP binding represents a convenient system for the functional characterization of human anti-HCV antibodies, allowing the mapping of envelope neutralization epitopes targeted by naturally occurring antiviral antibodies.     

Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge

A major problem in hepatitis C virus (HCV) immunotherapy or vaccine design is the extreme variability of the virus. We identified human monoclonal antibodies (mAbs) that neutralize genetically diverse HCV isolates and protect against heterologous HCV quasispecies challenge in a human liver-chimeric mouse model. The results provide evidence that broadly neutralizing antibodies to HCV protect against heterologous viral infection and suggest that a prophylactic vaccine against HCV may be achievable.     

Seroepidemiology of viral infections among intravenous drug users in northern California.

Intravenous drug users are frequently exposed to parenterally transmitted viral infections, and these infections can spread to the general population through sexual activity. We investigated the prevalence of serologic markers for human immunodeficiency virus type 1 (HIV-1), human T-cell lymphotropic virus type I/II (HTLV-I/II), hepatitis B virus (HBV), and hepatitis C virus (HCV) in intravenous drug users and their sexual contacts. Of 585 drug users from northern California tested for these serologic markers, 72% were reactive for the antibody to HCV, 71% for the antibody to hepatitis B core antigen, 12% for HTLV-I/II antibodies, and 1% for the HIV-1 antibody. The prevalence of serologic markers for these four viruses correlated with the duration of intravenous drug use, the ethnic group, and the drug of choice. More than 85% of subjects infected with either HCV or HBV were coinfected with the other virus. All persons reactive to HTLV-I/II antibodies had antibodies for either HBV or HCV. Of 81 sexual contacts tested, 17% had evidence of HBV infection while only 6% were reactive for HTLV-I/II antibodies and 4% for the antibody to HCV. None of this group was infected with HIV-1. We conclude that HTLV-I/II and HCV are inefficiently transmitted to sexual contacts while HBV is spread more readily. Programs designed to discourage the sharing of drug paraphernalia, such as needle and syringe exchanges, should decrease the risk of parenterally spread viral infections in intravenous drug users and thus slow the spread of these infections to the general population.     

Antibodies Targeting Novel Neutralizing Epitopes of Hepatitis C Virus Glycoprotein Preclude Genotype 2 Virus Infection

Currently, there is no effective vaccine to prevent hepatitis C virus (HCV) infection, partly due to our insufficient understanding of the virus glycoprotein immunology. Most neutralizing antibodies (nAbs) were identified using glycoprotein immunogens, such as recombinant E1E2, HCV pseudoparticles or cell culture derived HCV. However, the fact that in the HCV acute infection phase, only a small proportion of patients are self-resolved accompanied with the emergence of nAbs, indicates the limited immunogenicity of glycoprotein itself to induce effective antibodies against a highly evolved virus. Secondly, in previous reports, the immunogen sequence was mostly the genotype of the 1a H77 strain. Rarely, other genotypes/subtypes have been studied, although theoretically one genotype/subtype immunogen is able to induce cross-genotype neutralizing antibodies. To overcome these drawbacks and find potential novel neutralizing epitopes, 57 overlapping peptides encompassing the full-length glycoprotein E1E2 of subtype 1b were synthesized to immunize BALB/c mice, and the neutralizing reactive of the induced antisera against HCVpp genotypes 1–6 was determined. We defined a domain comprising amino acids (aa) 192–221, 232–251, 262–281 and 292–331 of E1, and 421–543, 564–583, 594–618 and 634–673 of E2, as the neutralizing regions of HCV glycoprotein. Peptides PUHI26 (aa 444–463) and PUHI45 (aa 604–618)-induced antisera displayed the most potent broad neutralizing reactive. Two monoclonal antibodies recognizing the PUHI26 and PUHI45 epitopes efficiently precluded genotype 2 viral (HCVcc JFH and J6 strains) infection, but they did not neutralize other genotypes. Our study mapped a neutralizing epitope region of HCV glycoprotein using a novel immunization strategy, and identified two monoclonal antibodies effective in preventing genotype 2 virus infection.     

Epidermal Growth Factor Receptor-Dependent Mutual Amplification between Netrin-1 and the Hepatitis C Virus

Hepatitis C virus (HCV) is an oncogenic virus associated with the onset of hepatocellular carcinoma (HCC). The present study investigated the possible link between HCV infection and Netrin-1, a ligand for dependence receptors that sustains tumorigenesis, in particular in inflammation-associated tumors. We show that Netrin-1 expression is significantly elevated in HCV+ liver biopsies compared to hepatitis B virus (HBV+) and uninfected samples. Furthermore, Netrin-1 was upregulated in all histological stages of HCV+ hepatic lesions, from minimal liver fibrosis to cirrhosis and HCC, compared to histologically matched HCV- tissues. Both cirrhosis and HCV contributed to the induction of Netrin-1 expression, whereas anti-HCV treatment resulted in a reduction of Netrin-1 expression. In vitro, HCV increased the level and translation of Netrin-1 in a NS5A-La-related protein 1 (LARP1)-dependent fashion. Knockdown and forced expression experiments identified the receptor uncoordinated receptor-5 (UNC5A) as an antagonist of the Netrin-1 signal, though it did not affect the death of HCV-infected cells. Netrin-1 enhanced infectivity of HCV particles and promoted viral entry by increasing the activation and decreasing the recycling of the epidermal growth factor receptor (EGFR), a protein that is dysregulated in HCC. Netrin-1 and HCV are, therefore, reciprocal inducers in vitro and in patients, as seen from the increase in viral morphogenesis and viral entry, both phenomena converging toward an increase in the level of infectivity of HCV virions. This functional association involving a cancer-related virus and Netrin-1 argues for evaluating the implication of UNC5 receptor ligands in other oncogenic microbial species.     

Towards a solution for hepatitis C virus hypervariability: mimotopes of the hypervariable region 1 can induce antibodies cross-reacting with a large number of viral variants.

The hypervariable region 1 (HVR1) of the putative envelope protein E2 of hepatitis C virus (HCV) is the most variable antigenic fragment in the whole viral genome and is mainly responsible for the large inter-and intra-individual heterogeneity of the infecting virus. It contains a principal neutralization epitope and has been proposed as the major player in the mechanism of escape from host immune response. Since anti-HVR1 antibodies are the only species shown to possess protective activity up to date, developing an effective prevention therapy is a very difficult task. We have approached the problem of HVR1 variability by deriving a consensus profile from >200 HVR1 sequences from different viral isolates and used it as a template to generate a vast repertoire of synthetic HVR1 surrogates displayed on M13 bacteriophage. This library was affinity selected using many different sera from infected patients. Phages were identified which react very frequently with patients' sera and bind serum antibodies that cross-react with a large panel of HVR1 peptides derived from natural HCV variants. When injected into experimental animals, the 'mimotopes' with the highest cross-reactivity induced antibodies which recognized the same panel of natural HVR1 variants. In these mimotopes we identified a sequence pattern responsible for the observed cross-reactivity. These data may hold the key for future development of a prophylactic vaccine against HCV.     

Amino Acid Residue-Specific Neutralization and Nonneutralization of Hepatitis C Virus by Monoclonal Antibodies to the E2 Protein

Antibodies to epitopes in the E2 protein of hepatitis C virus (HCV) reduce the viral infectivity in vivo and in vitro. However, the virus can persist in patients in the presence of neutralizing antibodies. In this study, we generated a panel of monoclonal antibodies that bound specifically to the region between residues 427 and 446 of the E2 protein of HCV genotype 1a, and we examined their capacity to neutralize HCV in a cell culture system. Of the four monoclonal antibodies described here, two were able to neutralize the virus in a genotype 1a-specific manner. The other two failed to neutralize the virus. Moreover, one of the nonneutralizing antibodies could interfere with the neutralizing activity of a chimpanzee polyclonal antibody at E2 residues 412 to 426, as it did with an HCV-specific immune globulin preparation, which was derived from the pooled plasma of chronic hepatitis C patients. Mapping the epitope-paratope contact interfaces revealed that these functionally distinct antibodies shared binding specificity for key amino acid residues, including W⁴³⁷, L⁴³⁸, L⁴⁴¹, and F⁴⁴², within the same epitope of the E2 protein. These data suggest that the effectiveness of antibody-mediated neutralization of HCV could be deduced from the interplay between an antibody and a specific set of amino acid residues. Further understanding of the molecular mechanisms of antibody-mediated neutralization and nonneutralization should provide insights for designing a vaccine to control HCV infection in vivo.     

High-avidity monoclonal antibodies against the human scavenger class B type I receptor efficiently block hepatitis C virus infection in the presence of high-density lipoprotein

The human scavenger class B type 1 receptor (SR-B1/Cla1) was identified as a putative receptor for hepatitis C virus (HCV) because it binds to soluble recombinant HCV envelope glycoprotein E2 (sE2). High-density lipoprotein (HDL), a natural SR-B1 ligand, was shown to increase the in vitro infectivity of retroviral pseudoparticles bearing HCV envelope glycoproteins and of cell culture-derived HCV (HCVcc), suggesting that SR-B1 promotes viral entry in an HDL-dependent manner. To determine whether SR-B1 participates directly in HCV infection or facilitates HCV entry through lipoprotein uptake, we generated a panel of monoclonal antibodies (MAbs) against native human SR-B1. Two of them, 3D5 and C167, bound to conformation-dependent SR-B1 determinants and inhibited the interaction of sE2 with SR-B1. These antibodies efficiently blocked HCVcc infection of Huh-7.5 hepatoma cells in a dose-dependent manner. To examine the role of HDL in SR-B1-mediated HCVcc infection, we set up conditions for HCVcc production and infection in serum-free medium. HCVcc efficiently infected Huh-7.5 cells in the absence of serum lipoproteins, and addition of HDL led to a twofold increase in infectivity. However, the HDL-induced enhancement of infection had no impact on the neutralization potency of MAb C167, despite its ability to inhibit both HDL binding to cells and SR-B1-mediated lipid transfer. Of note, MAb C167 also potently blocked Huh-7.5 infection by an HCV strain recovered from HCVcc-infected chimpanzees. These results demonstrate that SR-B1 is essential for infection with HCV produced in vitro and in vivo and suggest the possible use of anti-SR-B1 antibodies as therapeutic agents.     

Adaptation of Hepatitis C Virus to Mouse CD81 Permits Infection of Mouse Cells in the Absence of Human Entry Factors

Hepatitis C virus (HCV) naturally infects only humans and chimpanzees. The determinants responsible for this narrow species tropism are not well defined. Virus cell entry involves human scavenger receptor class B type I (SR-BI), CD81, claudin-1 and occludin. Among these, at least CD81 and occludin are utilized in a highly species-specific fashion, thus contributing to the narrow host range of HCV. We adapted HCV to mouse CD81 and identified three envelope glycoprotein mutations which together enhance infection of cells with mouse or other rodent receptors approximately 100-fold. These mutations enhanced interaction with human CD81 and increased exposure of the binding site for CD81 on the surface of virus particles. These changes were accompanied by augmented susceptibility of adapted HCV to neutralization by E2-specific antibodies indicative of major conformational changes of virus-resident E1/E2-complexes. Neutralization with CD81, SR-BI- and claudin-1-specific antibodies and knock down of occludin expression by siRNAs indicate that the adapted virus remains dependent on these host factors but apparently utilizes CD81, SR-BI and occludin with increased efficiency. Importantly, adapted E1/E2 complexes mediate HCV cell entry into mouse cells in the absence of human entry factors. These results further our knowledge of HCV receptor interactions and indicate that three glycoprotein mutations are sufficient to overcome the species-specific restriction of HCV cell entry into mouse cells. Moreover, these findings should contribute to the development of an immunocompetent small animal model fully permissive to HCV.     

Occult HCV Infection: An Unexpected Finding in a Population Unselected for Hepatic Disease

Background

Occult Hepatitis C virus (HCV) infection is a new pathological entity characterized by presence of liver disease and absence or very low levels of detectable HCV-RNA in serum. Abnormal values of liver enzymes and presence of replicative HCV-RNA in peripheral blood mononuclear cells are also observed. Aim of the study was to evaluate occult HCV occurrence in a population unselected for hepatic disease.

Methodology/Principal Findings

We chose from previous epidemiological studies three series of subjects (n = 276, age range 40–65 years) unselected for hepatic disease. These subjects were tested for the presence of HCV antibodies and HCV-RNA in plasma and in the peripheral blood mononuclear cells (PBMCs) by using commercial systems. All subjects tested negative for HCV antibodies and plasma HCV-RNA and showed normal levels of liver enzymes; 9/276 patients (3.3%) were positive for HCV-RNA in PBMCs, identifying a subset of subjects with potential occult HCV infection. We could determine the HCV type for 8 of the 9 patients finding type 1a (3 patients), type 1b (2 patients), and type 2a (3 patients).

Conclusions

The results of this study show evidence that occult HCV infection may occur in a population unselected for hepatic disease. A potential risk of HCV infection spread by subjects harbouring occult HCV infection should be considered. Design of prospective studies focusing on the frequency of infection in the general population and on the clinical evolution of occult HCV infection will be needed to verify this unexpected finding.     

Detection of hepatitis C virus RNA in the hearts of patients with hepatogenic cardiomyopathy

We examined the Hepatitis C virus (HCV) genome in the myocardium and liver obtained at autopsy from seven patients with HCV-positive liver cirrhosis and hepatocellular carcinoma (HCC) by in situ hybridization and histopathological studies. The HCV virus genome was detected in the myocardium of one patient as well as in the liver in three out of seven patients. However, Epstein-Barr (EB) virus genome could not be detected in liver or myocardium. In the patient who showed positive reaction to HCV in myocardium, both serum HCV and Hepatitis B virus (HBV) antibodies were positive. It is unknown whether this was related to an immunological abnormality of the host or to an interaction between RNA and DNA viruses. In conclusion, we could identify the HCV genome in the myocardium of a patient with hepatogenic myocardosis.     

Mapping of a conformational epitope shared between E1 and E2 on the serum-derived human hepatitis C virus envelope

Monoclonal antibody D32.10 produced by immunizing mice with a hepatitis C virus (HCV)-enriched pellet obtained from plasmapheresis of a chronically HCV1b-infected patient binds HCV particles derived from serum of different HCV1a- and HCV1b-infected patients. Moreover, this monoclonal has been shown to recognize both HCV envelope proteins E1 and E2. In an attempt to provide novel insight into the membrane topology of HCV envelope glycoproteins E1 and E2, we localized the epitope recognized by D32.10 on the E1 and/or E2 sequence using Ph.D.-12 phage display peptide library technology. Mimotopes selected from the phage display dodecapeptide library by D32.10 shared partial similarities with 297RHWTTQGCNC306 of the HCV E1 glycoprotein and with both 613YRLWHYPCT621 and 480PDQRPYCWHYPPKPC494 of the HCV E2 glycoprotein. Immunoreactivity of D32.10 with overlapping peptides corresponding to these three HCV regions confirmed these localizations and suggested that the three regions identified are likely closely juxtaposed on the surface of serum-derived particles as predicted by the secondary model structure of HCV E2 derived from the tick-borne encephalitis virus E protein. This assertion was supported by the detection of specific antibodies directed against these three E1E2 regions in sera from HCV-infected patients.     

Human apolipoprotein e is required for infectivity and production of hepatitis C virus in cell culture

Recent advances in reverse genetics of hepatitis C virus (HCV) made it possible to determine the properties and biochemical compositions of HCV virions. Sedimentation analysis and characterization of HCV RNA-containing particles produced in the cultured cells revealed that HCV virions cover a large range of heterogeneous densities in sucrose gradient. The fractions of low densities are infectious, while the higher-density fractions containing the majority of HCV virion RNA are not. HCV core protein and apolipoprotein B and apolipoprotein E (apoE) were detected in the infectious HCV virions. The level of apoE correlates very well with HCV infectivity. Both apoE- and HCV E2-specific monoclonal antibodies precipitated HCV, demonstrating that HCV virions contain apoE and E2 proteins. apoE-specific monoclonal antibodies efficiently neutralized HCV infectivity in a dose-dependent manner, resulting in a reduction of infectious HCV by nearly 4 orders of magnitude. The knockdown of apoE expression by specific small interfering RNAs (siRNAs) remarkably reduced the levels of intracellular as well as secreted HCV virions. The apoE siRNA suppressed HCV production by more than 100-fold at 50 nM. These findings demonstrate that apoE is required for HCV virion infectivity and production, suggesting that HCV virions are assembled as apoE-enriched lipoprotein particles. Our findings also identified apoE as a novel target for discovery and development of antiviral drugs and monoclonal antibodies to suppress HCV virion formation and infection.     

Preclinical evaluation of two neutralizing human monoclonal antibodies against hepatitis C virus (HCV): a potential treatment to prevent HCV reinfection in liver transplant patients

Passive immunotherapy is potentially effective in preventing reinfection of liver grafts in hepatitis C virus (HCV)-associated liver transplant patients. A combination of monoclonal antibodies directed against different epitopes may be advantageous against a highly mutating virus such as HCV. Two human monoclonal antibodies (HumAbs) against the E2 envelope protein of HCV were developed and tested for the ability to neutralize the virus and prevent human liver infection. These antibodies, designated HCV-AB 68 and HCV-AB 65, recognize different conformational epitopes on E2. They were characterized in vitro biochemically and functionally. Both HumAbs are immunoglobulin G1 and have affinity constants to recombinant E2 constructs in the range of 10(-10) M. They are able to immunoprecipitate HCV particles from infected patients' sera from diverse genotypes and to stain HCV-infected human liver tissue. Both antibodies can fix complement and form immune complexes, but they do not activate complement-dependent or antibody-dependent cytotoxicity. Upon complement fixation, the monoclonal antibodies induce phagocytosis of the immune complexes by neutrophils, suggesting that the mechanism of viral clearance includes endocytosis. In vivo, in the HCV-Trimera model, both HumAbs were capable of inhibiting HCV infection of human liver fragments and of reducing the mean viral load in HCV-positive animals. The demonstrated neutralizing activities of HCV-AB 68 and HCV-AB 65 suggest that they have the potential to prevent reinfection in liver transplant patients and to serve as prophylactic treatment in postexposure events.     

Hepatitis C virus (HCV)-induced immunoglobulin hypermutation reduces the affinity and neutralizing activities of antibodies against HCV envelope protein

Hepatitis C virus (HCV) often causes persistent infection despite the presence of neutralizing antibodies against the virus in the sera of hepatitis C patients. HCV infects both hepatocytes and B cells through the binding of its envelope glycoprotein E2 to CD81, the putative viral receptor. Previously, we have shown that E2-CD81 interaction induces hypermutation of heavy-chain immunoglobulin (V(H)) in B cells. We hypothesize that if HCV infects antibody-producing B cells, the resultant hypermutation of V(H) may lower the affinity and specificity of the HCV-specific antibodies, enabling HCV to escape from immune surveillance. To test this hypothesis, we infected human hybridoma clones producing either neutralizing or non-neutralizing anti-E2 or anti-E1 antibodies with a lymphotropic HCV (SB strain). All of the hybridoma clones, except for a neutralizing antibody-producing hybridoma, could be infected with HCV and support virus replication for at least 8 weeks after infection. The V(H) sequences in the infected hybridomas had a significantly higher mutation frequency than those in the uninfected hybridomas, with mutations concentrating in complementarity-determining region 3. These mutations lowered the antibody affinity against the targeting protein and also lowered the virus-neutralizing activity of anti-E2 antibodies. Furthermore, antibody-mediated complement-dependent cytotoxicity with the antibodies secreted from the HCV-infected hybridomas was impaired. These results suggest that HCV infection could cause some anti-HCV-antibody-producing hybridoma B cells to make less-protective antibodies.     

Hepatitis C virus and Epstein-Barr virus: dual infection or atypical antibody response

Positive serological reactions against hepatitis C virus (HCV) appeared in the course of Epstein-Barr virus (EBV) infectious mononucleosis. In 429 consecutive patients with high levels of transaminases, 28 patients with EBV primary infection were found. The presence of anti-HCV antibodies and HCV RNA was studied in these subjects. In seven patients anti-HCV antibodies (C33 and C22c RIBA bands) were detected, but all were polymerase chain reaction (PCR) negative. These results may have been due solely to a HCV infection or were an atypical response to HCV in the course of infectious mononucleosis.     

Hepatitis C virus induces CD81 and claudin-1 endocytosis

Hepatitis C virus (HCV) leads to progressive liver disease and hepatocellular carcinoma. Current treatments are only partially effective, and new therapies targeting viral and host pathways are required. Virus entry into a host cell provides a conserved target for therapeutic intervention. Tetraspanin CD81, scavenger receptor class B member I, and the tight-junction proteins claudin-1 and occludin have been identified as essential entry receptors. Limited information is available on the role of receptor trafficking in HCV entry. We demonstrate here that anti-CD81 antibodies inhibit HCV infection at late times after virus internalization, suggesting a role for intracellular CD81 in HCV infection. Several tetraspanins have been reported to internalize via motifs in their C-terminal cytoplasmic domains; however, CD81 lacks such motifs, leading several laboratories to suggest a limited role for CD81 endocytosis in HCV entry. We demonstrate CD81 internalization via a clathrin- and dynamin-dependent process, independent of its cytoplasmic domain, suggesting a role for associated partner proteins in regulating CD81 trafficking. Live cell imaging demonstrates CD81 and claudin-1 coendocytosis and fusion with Rab5 expressing endosomes, supporting a role for this receptor complex in HCV internalization. Receptor-specific antibodies and HCV particles increase CD81 and claudin-1 endocytosis, supporting a model wherein HCV stimulates receptor trafficking to promote particle internalization.     

Hepatitis C Virus Induces Epidermal Growth Factor Receptor Activation via CD81 Binding for Viral Internalization and Entry

While epidermal growth factor receptor (EGFR) has been shown to be important in the entry process for multiple viruses, including hepatitis C virus (HCV), the molecular mechanisms by which EGFR facilitates HCV entry are not well understood. Using the infectious cell culture HCV model (HCVcc), we demonstrate that the binding of HCVcc particles to human hepatocyte cells induces EGFR activation that is dependent on interactions between HCV and CD81 but not claudin 1. EGFR activation can also be induced by antibody mediated cross-linking of CD81. In addition, EGFR ligands that enhance the kinetics of HCV entry induce EGFR internalization and colocalization with CD81. While EGFR kinase inhibitors inhibit HCV infection primarily by preventing EGFR endocytosis, antibodies that block EGFR ligand binding or inhibitors of EGFR downstream signaling have no effect on HCV entry. These data demonstrate that EGFR internalization is critical for HCV entry and identify a hitherto-unknown association between CD81 and EGFR.     

Hepatitis B and C in the hemodialysis unit of Tocantins,Brazil: serological and molecular profiles

A survey was conducted in the hemodialysis population of the state of Tocantins, Brazil, aiming to assess the prevalence of hepatitis B virus (HBV) and hepatitis C virus (HCV) infections, to analyze associated risk factors, and also to investigate these viruses genotypes distribution. During January and March 2001, all patients (n = 100) were interviewed at the unique dialysis unit in Tocantins. Blood samples were collected and serum samples were screened for HBV serological markers. Hepatitis B surface antigen positive samples were tested for HBV DNA. All samples were also tested for anti-HCV antibodies and HCV RNA. An overall prevalence of 45% was found for HBV infection (4% were HBsAg/anti-HBc positive, 2% were anti-HBc only and 39% had anti-HBc/anti-HBs markers). Concerning HCV infection, anti-HCV and HCV RNA were detected in 13% and 14% of the subjects, respectively. Three patients were HCV RNA positive and anti-HCV negative, resulting in an overall HCV prevalence of 16%. Univariate analysis of risk factors showed that only shift and length of tile on hemodialysis were associated with HBV and HCV positivity respectively. Among the four HBsAg-positive samples, HBV DNA was detected in three of them, which were identified as genotype A by restriction fragment length polymorphism (RFLP) analysis. All 14HCV RNA-positive samples were genotyped by INNO-LiPA. Genotypes la and 3a were found in 85% and 15%, respectively. The present data show low HBsAg and HCV prevalence rates. The risk factors associated with HBV and HCV positivity suggest that nosocomial transmission may influence in spreading these viruses in the dialysis unit studied.