GBV-C: State of the Art and Future Prospects
- First Online:
- Cite this article as:
- Giret, M.T.M. & Kallas, E.G. Curr HIV/AIDS Rep (2012) 9: 26. doi:10.1007/s11904-011-0109-1
The GB virus C is a common non-pathogenic virus, member of the Flaviviridae family with worldwide distribution. Favorable clinical course and reduced mortality among HIV-infected patients was demonstrated by several studies with patients co-infected with the GB virus C (GBV-C). This potential benefit of GBV-C has been demonstrated in the pre-HAART and post-HAART eras; however, this effect was not observed in all studies and the discrepancy may be due to changes during the course of HIV infection, characteristic of the cohort, and the degree of therapeutic response. The GBV-C has been found to decrease HIV replication in in vitro models, highlighting the interference of persistent GBV-C viremia. The mechanism of the beneficial effect of GBV-C appears to be mediated by changes in the cellular immune response, and elucidation of putative protective effects of GBV-C in HIV co-infection could potentially identify novel targets for anti-HIV agents.
KeywordsHuman immunodeficiency virus (HIV)GB virus C (GBV-C)Co-infectionActivationInhibition
During the past several years, our understanding of the concepts of HIV-1 interaction with other agents has made us think about how one virus can affect the reaction to another virus in humans.
Since the discovery of the GB virus C (GBV-C) and hepatitis G (HGV) by the Virus Discovery groups at Genelabs, Inc., and Abbott Laboratories [1, 2], the study of this virus has become a new challenge in the attempt to retrospectively track down the causative agent of an acute hepatitis. However, a few years later, no link to infectious hepatitis has been found, but a surprising association between this virus and improved survival in HIV-infected individuals has been described. This is the reason why it motivates new research approaches and critical studies, which are still in progress.
In this review we first briefly discuss the GB virus C basic biology, the epidemiology, and the transmission, focusing on the implication and significance of GBV-C in the pathogenesis of HIV infection. We then outline some of the approaches in the understanding of the mechanisms involved in the interaction of two such viruses, with emphasis on the immunological and clinical insights provided by these studies. Finally, we highlight the GBV-C proteins that have been identified, which specifically appears to inhibit HIV replication.
The GB Virus C: Epidemiology and Transmission
The GB virus type C (GBV-C, also previously described as hepatitis G virus) is an enveloped, positive-sense, single-stranded RNA virus belonging to the family Flaviviridae, and is closely related to the hepatitis C virus (HCV). It has been proposed that it should be classified as a member of a fourth genus in the Flaviviridae family, named Peguivirus . GBV-C is a lymphotropic virus and the GBV-C quasispecies nature of GBV-C in lymphocyte subsets within peripheral blood mononuclear cells is apparently related to the viral population infecting CD8(+) T cells and B cells .
GB virus C is a common, non-pathogenic human blood-borne virus discovered in 1995, of ancient ancestry. A GBV-C virus variant was identified in chimpanzees and the prevalence and natural history of chimpanzee GBV-C variant (GBV-C, cpz) appears to be similar to the human GBV-C infection .
The geographical distribution is related to the co-evolution of the viruses with humans during the migrations throughout history [6, 7] and phylogenetics analyses of GBV-C isolates have demonstrated the presence of multiple genotypes with consistent geographical clustering .
Co-infection with GBV-C is frequent in patients suffering from HIV type-1 (HIV-1) infection due to common routes of transmission.
The exposure to GBV-C can be assessed by measuring both GBV-C RNA and E2 antibodies. The presence of GBV-C RNA is based on nucleic acid amplification and quantification, determined by real-time reverse transcription polymerase chain reaction (RT-PCR). Some immunocompetent individuals can eliminate GBV-C virus within the few first years after infection; however, the scenario may be different in HIV-infected individuals, in whom the GBV-C clearance may not always be linked with the development of E2 antibodies. Likewise, the role of E2 antibody development in GBV-C RNA clearance remains unclear. Some recent studies have attempted to address this question and in a cohort of recently HIV-1–infected subjects, the incidence density of GBV-C clearance was found to be 10 cases per 100 person-year, whereas among those HIV-positive individuals susceptible to GBV-C RNA infection, the incidence density of new GBV-C infections was threefold lower . Clearance of viremia is generally associated with the resolution of GBV-C infection, and the development of antibodies to the envelope glycoprotein E2, detected by immunoassays, may indicate prior infection. Similarly, the lack of E2 antibody detection may represent, in the first place, the absence of sensitivity of the nucleic acid tests to detect low levels of viral replication by the conventional methods. Second, the length of window period between the clearance of viremia and the detection of antibodies is unknown. Third, the lack of specificity of the EIA for E2 antibodies can lead to undetectable level of antibodies. On the other hand, concomitant detection of GBV-C RNA and E2 antibodies in blood is rare.
Since the exact role of anti–GBV-C antibodies in HIV-infected patients also remains unknown and commercial systems to detect specific markers of GBV-C infection are not available, the construction of chimeric multiple antigenic peptides (MAPs) formed by two domains of different GBV-C proteins, and containing epitopes from E2, NS4, and NS5 GBV-C, has been recently proposed. Thus, sequences from envelope and nonstructural GBV-C proteins with good sensitivity/specificity balances in the detection of anti–GBV-C antibodies can provide the basis for future prevalence studies of the GBV-C infection based on serodiagnosis [9, 10].
Summary of immunological evidence for the possible role of GBV-C and the beneficial interaction with HIV-1
HIV-positive persons with a well-defined duration of infection
HIV RNA load, CD4 T-cell count, survival, CDC stage
Higher CD4T cell count, lower HIV viral load, slower progression to AIDS, lower mortality
Lefrere et al. 
Patients with hemophilia who became HIV-infected
CCR5 genotype, HIV and HCV viral loads, CD4 and CD8 lymphocyte counts
Higher CD4T cell count and better AIDS-free survival rates
Yeo et al. 
GB virus C co-infection in HIV-1 women, homosexual and IVDU
GBV-C and HIV viral load, CD4 cell count and disease progression
Higher CD4 and CD8T cell count, slower progression to AIDS, longer survival
Tillmann et al. 
GB virus C co-infection in HIV-1-infected women and IVDU individuals
HIV RNA load, CD4 T-cell count, survival
Improved survival and higher CD4 cell count
Xiang et al. 
Asymptomatic HIV-1–seropositive patients
GBV-C RNA level; plasma HIV-1 viral load; CD4 cell counts, IL-2, IL-4, IL-10, and IL-12
Higher CD4T cell count and intact T-helper 1 cytokine profile
Nunnari et al. 
GBV-C RNA, E2 antibody, HIV disease progression
Williams et al. 
Untreated HIV-1 patients with GBV-C coinfection
Cell surface Fas expression; apoptosis
Expression of Fas directly correlated with sensitivity to Fas-mediated apoptosis; reduced cell surface Fas expression in GBV-C viremic patients
Moenkemeyer et al. 
GBV-C co-infection in HIV-1-infected patients with HAART
Activation of the interferon system and circulating (DC) in vivo
Increasing frequency of circulating CD80+ plasmocytoid DC (pDC)
Lalle et al. 
Jurkat cell lines expressing NS5A protein and peptides
HIV replication in cell lines
Expression of GBV-C NS5A amino acids 152–165 is sufficient to inhibit HIV replication in vitro
Xiang et al. 
GB virus C co-infection in recently HIV type-1-infected subjects
Activation markers: CD38, CCR5, CD69, CD25, HLA-DR
Reduction in the CD38 on CD4 and CD8T cells and CCR5 on CD8T cells; lower expression of CD69 and CD25
Maidana-Giret et al. 
GB virus C co-infection in advanced HIV type-1 disease
CCR5 and CXCR4 co-receptors
Reduction in the expression of CCR5 and CXCR4 co-receptors on CD4T cells
Schwarze-Zander et al. 
Mounting evidence has emerged to demonstrate that the parenteral route is, beside the well-described sexual contact, an important method of viral transmission. Despite the variable prevalence found in several HIV-1 cohort studies, GBV-C acquisition is consistently more frequent when comparing to other groups of individuals.
The prevalence of GBV-C plasma viremia is significantly higher in asymptomatic (16.7%) and symptomatic (16.2%) HIV-infected individuals [7, 17–19] than in blood donors (bellow 4%) [6, 20, 21]. No statistically significant difference has been reported in the prevalence of GBV-C infection among HIV-infected subjects comparing intravenous drug users (IDUs) (13.5%) to heterosexuals (6.7%) . Another report has shown a similar prevalence (13.6%) in a group of non-infected HIV-infected hemodialysis patients, all of them from the same genotype 2, supporting the hypothesis of parenteral transmission . Moreover, GBV-C infection was associated with HIV infection and both sexual and parenteral risk behaviors, and findings from cohorts of pregnant women in Thailand demonstrated that it was independently associated with an increasing number of lifetime sexual partners .
Although the viral load is an important factor associated with the rate of vertical HIV transmission, the report by Supapol et al.  adds new and interesting information in the transmission from mother to child through pregnancy and/or delivery. In agreement with previously published data, the authors found no association between maternal GBV-C status and the HIV transmission. More importantly, the reduced mother-to-child transmission (MTCT) of HIV was significantly associated with infant acquisition of GBV-C, but not with maternal GBV-C infection.
The frequency of GBV-C exposure in children with chronic renal failure has been shown to be as high as 51% compared to the healthy group with 8%, and may be the result of frequent blood transfusions and longer disease course. However, GBV-C infection, in these children, was not associated with significant changes in hepatic biochemical parameters .
Implication and Significance of GBV-C in the Pathogenesis of HIV Infection
The discovery of the GB virus C provided a new insight into a novel virus-virus interaction. GBV-C virus was initially studied as part of the viral hepatitis research groups; nevertheless, as mentioned earlier, it remains a virus that has not been linked to any disease in humans. Most of the attention paid to GBV-C has focused on its potential progression-influencing effect in HIV infection and over the past several years a number of studies have found GBV-C to exert a favorable impact on the course of HIV-1 by deferring the setting of immunodeficiency. GBV-C co-infected patients had reduced HIV RNA levels, and in recent years, compelling evidence for down regulation on CD4 molecule expression or HIV CD4 modulatory effect by structural conserved elements, among flaviviruses, has been demonstrated .
Whether a beneficial effect on the immune system of the HIV-infected patient is exerted by GBV-C, a correlation of GBV-C load with higher CD4+ T-cell counts or lower HIV RNA copies/mL should be expected, reflecting inhibition of HIV replication. Perhaps, some studies failed to demonstrate such relationship  and others have found a trend toward a negative correlation between the GBV-C load and the HIV-1 viral load, but no correlation between the GBV-C load and the number of CD4+ T cells at the earlier visit . This specific result may be explained as it is composed of recently HIV-1–infected individuals. Additionally, the median viral load of GBV-C at 1 year after enrollment was not statistically significantly higher compared to the baseline levels. Therefore, it is not possible to conclude whether the increase in GBV-C viral load can be due to viral transactivation exerted by HIV-1 replication, or to immunodeficiency progression. Nevertheless, these previous data are consistent with a recent study showing that GBV-C viremia appears to reactivate in HIV-infected individuals, resulting in a significant increase of GBV-C viral load .
Down regulation of CD4 molecule by flaviviruses may also attenuate HIV pathogenesis by decreasing either the TCR signal required for activation or by lowering the threshold of the TCR signal. In addition, modulation of T-cell activation was recently described and GBV-C co-infection was associated with reduced T-cell activation (CD38 + CD4+, CD38 + CD8+, CCR5 + CD8+) . Another very robust cohort study, followed in the AIDS Clinical Trials Group 328 study, characterized other important factors in response to IL-2 treatment. In this study, GBV-C–infected individuals have blunted CD4 proliferation in response to IL-2 therapy . Perhaps, the measurement of GBV-C viremia status prior to administration of IL-2 therapy has not yet been validated.
The persistence of GB virus C viremia in patients with chronic HIV-1 infection associated with increased survival has been documented. But others factors rather than human genetics must be involved because the frequency of GB virus C viremia in HIV-infected elite suppressors and chronically HIV-infected patients with progressive disease was not significantly different, and GB virus C does not appear to explain the favorable course seen in the group of elite suppressors .
GBV-C co-infection in HIV-1 disease leads to reduced expression of the two major HIV-1 co-receptors, CCR5 and CXCR4, on CD4+ T cells in patients at an advanced stage of immunodeficiency, providing a possible molecular explanation for the clinical benefit of GBV-C co-infection at an advanced stage of immunodeficiency .
Whether GBV-C infection has a significance in other viral infections remains to be characterized, and this may have rendered new data such as the lower degree of hepatic lesions in HCV infections observed in the triple-infected patients, in comparison to HIV-HCV co-infected who were negative for GBV-C RNA , as well as an increased rate of GBV-C clearance among young, HIV-seronegative injection drug users chronically infected by HCV . The co-infection with GBV-C in patients with chronic hepatitis C does not worsen the clinical course of chronic hepatitis C or diminish response of HCV to viral therapy, as perhaps the interferon/ribavirin combination therapy may clear GBV-C viremia .
In the context of GBV-C diversity, the extent of our knowledge is still limited. Consequently, little information is available for the geographical distribution of the genotypes and its influence on the clinical outcome. It is difficult to conclude whether the GBV-C genotype influences the clinical outcome due to the limited number of cases in the study cohorts after such stratification. Larger studies may demonstrate whether a particular genotype, if any, has a different impact in the clinical course of HIV-1 infection or in the surrogate markers of progression. Moreover, it is still not known if the spontaneous resolution of GBV-C viremia is necessarily followed by the appearance of E2 antibodies or only reflects a fluctuation of GBV-C RNA levels. Therefore, it is also important to know the GBV-C genotype if we consider that this aspect can impact the progression of HIV disease.
New Mechanisms of Interaction Proposed
The variability in both the clinical progression and transmission of HIV infection has prompted a search for co-factors influencing the virus replication. Although it is clear that host immune and genetic factors influence the progression of HIV disease, as well as the replication kinetics of particular viral strains, a variety of acquired infectious agents also appear to influence the HIV replication, the activation of CD4+ and CD8+ T cells, and the widespread cytokine secretion.
As in many viral infections, chronic immune activation is a hallmark of progressive HIV disease. A new mechanism was proposed in a study cohort of recently HIV infected individuals, showing that GB virus type C exerts a favorable influence by modulating T-cell activation independently of HIV-1 viral load . Other authors addressed the evidence of GBV-C association with an improvement in cirrhosis-free survival in HCV/HIV co-infected subjects and following the data from recent years, raising a question about how GBV-C might influence this effect. The observations, from the same group, focused on the identification of genes and proteins involved in TCR signaling and T-cell activation such as the lymphocyte-specific protein tyrosine kinase (LCK), most commonly found in T cells, to be significantly reduced in associated T-cell signaling pathway in the liver from GBV-C–positive HCV/HIV individuals . This finding could be the reason, at least in part, for a reduction in HCV-related liver disease.
Since innate antiviral response may be involved in the protection, it has been speculated that the possible role of GBV-C is as an activator of innate immunity in HIV-positive patients. A study addressing the extent of activation of the interferon (IFN) system and of circulating dendritic cells revealed that GBV-C co-infection promotes the activation of IFN-γ, as well as the activation and maturation of circulating plasmacytoid DC, boosting the innate antiviral response to HIV infection. In addition, the frequency of circulating pDC expressing the CD80 activation marker was increased in GBV-C–positive patients, and was correlated with GBV-C viral load . The importance of the response to HIV disease progression is correlated with depletion of Th1 cytokines, which can be restored after initiating HAART.
In untreated HIV-1 patients GBV-C co-infection was associated with a significantly lower percentage of Fas-expressing cells as compared to GBV-C non–co-infected individuals. Expression of Fas was directly correlated with sensitivity to Fas-mediated apoptosis . These findings raise the possibility that GBV-C could interfere in the T-cell apoptosis in HIV-infected patients, which could explain the preserved levels of CD4+ T cells by the presence of GBV-C viremia in some cases.
The mechanistic studies analyzing the GBV-C structure are also in progress. In this aspect, GBV-C NS5A is a phosphoprotein required for infectivity and RNA replication and the group of Xiang et al. , following previous results, characterized the protein requirement within the GBV-C NS5A protein that is responsible for inhibiting HIV infection in vitro. Through the expression of 10 different NS5A polypeptides and after careful analyses, they suggested that either phosphorylation or a structural motif of the peptide is required for HIV inhibition.
The GBV-C NS5A mediates HIV inhibition in part by decreasing the surface expression of CXCR4, the other major HIV co-receptor, and by inducing release of stromal cell-derived factor 1 (SDF-1), the chemokine ligand for CXCR4. The same group expressed the DENV NS5 protein in a CD4 T-cell line (Jurkat) to determine whether this protein, like the related GBV-C NS5A protein, interferes with HIV replication. It appears that the HIV inhibition mediated by the DENV NS5 protein results from intracellular protein-protein interactions that lead to modification of cellular susceptibility to HIV infection, suggesting that this protein is involved in immune-evasion and/or replication . Furthermore, when the NS5 proteins of other flaviviruses were expressed in CD4+ T cells, the same inhibitory effect on HIV replication was observed, suggesting that regulation of CD4 expression by flaviviruses may interfere with innate and adaptive immunity and contributes to inhibition of in vitro HIV replication . Studies on the structural protein characteristics may provide further insight in the composition of the GBV-C viral structure and its interference in the HIV life cycle.
Although broadly neutralizing antibodies to HIV-1 are well described, the identification of GBV-C–specific antibodies that neutralize HIV replication has proven difficult. Data published last year have indicated that the anti-E2 Abs from the GBV-C neutralize and precipitate HIV-1 particles via interaction with a cellular Ag present in HIV-1 Gag particle, inhibiting HIV attachment to cells but not inhibiting entry steps following attachment. These data led the authors to conclude that GBV-C E2 protein has a structural motif that elicits antibody production that cross-reacts with a cellular antigen present on retrovirus particles, independent of HIV-1 envelope glycoproteins. The data have provided evidence that a heterologous viral protein can induce HIV-1–neutralizing antibodies [41••].
In the meantime, several studies in the physiochemical and biochemistry fields have been developed, some of them analyzing the envelope proteins of flaviviruses, whereas others characterizing putative fusion peptides within GBV-C E1 and E2 involved in fusion events. A better characterization of the envelope proteins and its interaction identified as a class II fusion proteins that are not cleaved during biosynthesis and fusion peptides were found within both GBV-C E1 [42, 43] and E2 [44, 45] proteins. One of the E2 peptides studied has shown to inhibit specifically the membrane leakage induced by the HIV-1 gp41 fusion peptide.
Recent findings are addressing the question of which region of the E2 protein is involved in suppression of HIV-1 replication. A peptide sequence of the E2 envelope protein of GBV-C has been proved to decrease cellular membrane fusion and interfere with the HIV-1 infectivity in a dose-dependent manner, indicating an E2 GBV-C peptide–gp41 HIV-1 peptide interaction. All these results support the hypothesis that gp41-fusion protein from the HIV is inhibited by the peptide P59 from the E2 protein of GB virus C .
The demonstration of the E2-derived peptides preventing HIV-1 binding or fusion, presumably via interaction with the HIV-1 particle, reveals a new mechanism of viral interference, suggesting that the envelope protein E2 of GBV-C targets directly HIV-1 particles to avoid entry of the latter 
The identification of other E1 proteins that could inhibit the activity of the peptide sequence of the surface protein gp41 of HIV, corresponding to the fusion peptide of the virus (HIV-1 FP), was the aim of another study. The results highlight a specific E1 synthetic peptide that could be involved in preventing the entry of HIV-1 by binding to the HIV-1 FP. Therefore, exploring GBV-C peptides and HIV-1 FP interaction could lead to the development of new therapeutic agents for the treatment of HIV/AIDS . The peptide described by the author is derived from the E1 structural protein of GBV-C that was previously shown to inhibit leakage of vesicular contents caused by the HIV-1 fusion peptide (HIV-1 FP).
In order to gain information on the fusogenic potential of virus-derived synthetic peptides, the same group examined their interfacial properties and studied them in monolayers and bilayers. The new results indicate that the peptide E1 (64–81) interacts with the fusion protein of HIV-1 (FP) to form a new structure, and that this may be the cause of the previously observed inhibition of the activity of HIV-1 FP by E1 (64–81) . After a construction of a large number of peptides, the group selected specific sequences as the best candidates for future studies. Those peptides were capable of inhibiting membrane fusion and the interaction of HIV-1 FP with bilayers . This new information could be potentially used in future anti–HIV-1 research.
Questions to be further explored
What insight to the dynamics of HIV-1/GBV-C co-infection?
What is the mechanism by which inhibition occurs?
What is the cellular antigen recognized by the anti–GBV-C E2 antibody?
Can GBV-C immune response be inferred using traditional methods?
What is the activity of described GBV-C–specific peptides against HIV-1 fusion peptide once incubated with cells?
Are there confounding factors that contribute to the divergent results such as introduction of HAART and GBV-C RNA levels?
Are there protection correlates and viral control attributes of exposed seronegative individuals?
Is GBV-C susceptible to any available antiviral drugs?
What is the role of GBV-C infection in monocytic-derived cells?
Can GBV-C infect CD4+ monocytic cells such as macrophages or monocyte-derived dendritic cells?
Is there a specific GBV-C genotype conferring protection against HIV disease progression?
The interaction of certain GBV-C peptide sequences with the HIV-1 fusion peptide has been proven through the use of biophysical techniques. Structure-activity and relationship studies have played a critical role in the ongoing debate about the GBV-C and HIV peptides interaction . Moreover, it has been shown how GBV-C E2 domains notably decrease cellular membrane fusion and interfere with the HIV-1 infectivity in a dose-dependent manner .
Two GBV-C proteins have been identified that specifically inhibit HIV replication, the E2 and NS5A. The E2 protein seems to inhibit the HIV gp 41 fusion and therefore an initial step in the HIV infection. Therefore, the continued study into the interaction between GBV-C peptides and HIV-1 fusion protein could lead to the development of new therapeutic agents for HIV/AIDS , and the recent study in natural history of non-human GBV-C in captive chimpanzees  has suggested that they could serve as a good host to explore HIV–GBV-C co-infection.
GBV-C and HIV-1 co-infection leads to reduced expression of the two major HIV-1 co-receptors, CCR5 and CXCR4, on CD4+ T cells in patients at an advanced stage of immunodeficiency , and the GBV-C viremia has shown to diminish CD4+ T-cell proliferation in response to IL-2 . Likewise, new groups of subjects are required to infer the patterns and pathways of viral spread within complex transmission networks. An especially interesting area of investigation would be the protection correlates and viral control attributes of exposed seronegative individuals, long-term non-progressors, and elite suppressors, and the association of GBV-C in these subsets of HIV-infected individuals. Furthermore, we recognize that the complex set of processes that drive the beneficial effect of GBV-C viremia in the context of HIV infection can be addressed most effectively by examining its constituent steps; this includes the immunological characteristic of the GBV-C/HIV-infected cells, the acquisition of antigen-binding receptor, and the regulatory processes that may take place in the intracellular compartment.
Additional work also is needed to identify the GBV-C E2 cellular receptor, characterize GBV-C E1 and E2 domains responsible for HIV inhibition, and to examine GBV-C E2-mediated fusion in the context of the entire envelope protein or viral particles .
Finally, it is essential to investigate the evolutionary pattern in the context of other pathogens that simultaneously circulate within human population. We are now in a better position to understand these essential mechanisms of interaction between these two viruses and whether the recognition of a specific GBV-C structure is responsible for eliciting anti-HIV activity that could serve as a therapeutic option in the future.
No potential conflicts of interest relevant to this article were reported.