Infection of Glia by Human Pegivirus Suppresses Peroxisomal and Antiviral Signaling Pathways.

Human pegivirus (HPgV) infects peripheral leukocytes but was recently shown to be a neurotropic virus associated with leukoencephalitis in humans. In the present study, we investigated the neural cell tropism of HPgV as well as its effects on host immune responses. HPgV wild type (WT) and a mutant virus with a deletion in the HPgV NS2 gene (ΔNS2) were able to productively infect human astrocytes and microglia but not neurons or an oligodendrocyte-derived cell line. Of note, the ΔNS2 virus replicated better than WT pegivirus in astrocytes, with both viruses being able to subsequently infect and spread in fresh human astrocyte cultures. Infection of human glia by HPgV WT and ΔNS2 viruses resulted in suppression of peroxisome-associated genes, including PEX11B, ABCD1, PEX7, ABCD3, PEX3, and PEX5L, during peak viral production, which was accompanied by reduced expression of IFNB, IRF3, IRF1, and MAVS, particularly in ΔNS2-infected cells. These data were consistent with analyses of brain tissue from patients infected with HPgV in which we observed suppression of peroxisome and type I interferon gene transcripts, including PEX11B, ABCD3, IRF1, and IRF3, with concurrent loss of PMP70 immunoreactivity in glia. Our data indicate that human astrocytes and microglia are permissive to HPgV infection, resulting in peroxisome injury and suppressed antiviral signaling that is influenced by viral diversity. IMPORTANCE Human pegiviruses are detected in 1 to 5% of the general population, principally infecting leukocytes, although their effects on human health remain uncertain. Here, we show that human pegivirus infects specific neural cell types in culture and human brain and, like other neurotropic flaviviruses, causes suppression of peroxisome and antiviral signaling pathways, which could favor ongoing viral infection and perhaps confer susceptibility to the development of neurological disease.

Transmission of GB virus type C via transfusion in a cohort of HIV-infected patients.

BACKGROUND: GB virus C (GBV-C) infection is transmitted by blood exposure and associated with lower human immunodeficiency virus (HIV) load and slower HIV disease progression. Few studies describe predictors of acute GBV-C infection following transfusion in HIV-infected patients. METHODS: We used a limited-access database from the National Heart Lung and Blood Institute's Viral Activation Transfusion Study, a randomized controlled trial of leukoreduced versus nonleukoreduced transfusions received by HIV-infected, transfusion-naive patients. Blood samples from 489 subjects were tested for GBV-C markers in pretransfusion and posttransfusion samples. We estimated the risk of acquiring GBV-C RNA and predictors of GBV-C acquisition, using pooled logistic regression. RESULTS: GBV-C RNA was detected ≤120 days following the first transfusion in 22 (7.5%) of 294 subjects who were GBV-C negative before transfusion. The risk of GBV-C RNA acquisition increased with each unit transfused (odds ratio, 1.09; 95% confidence interval, 1.06-1.11). Lower baseline HIV load and use of antiretroviral therapy were associated with subsequent GBV-C RNA acquisition, after control for units of blood transfused. Leukoreduced status of transfused units was not associated with GBV-C transmission. CONCLUSIONS: Blood transfusion is associated with a significant risk of GBV-C acquisition among HIV-infected patients. Transmission of GBV-C by blood transfusion was inversely related to HIV load.

Human pegivirus viremia in HCV/HIV co-infected patients: Direct acting antivirals exert anti-pegivirus effects.

BACKGROUND: Human pegivirus (HPgV) is a single-stranded RNA virus​ that is closely related to hepatitis C virus (HCV)​. HPgV has also been shown to infect patients with human immunodeficiency virus (HIV). The mechanisms and disease outcomes of HPgV infections are largely unknown, although it has been implicated in both cancer and neurological diseases. There are no established therapies for HPgV. OBJECTIVES: To estimate the prevalence of HPgV in a cohort of HCV/HIV co-infected patients undergoing treatment for HCV with direct acting antivirals (DAA) and investigate the effect of DAA therapy on HPgV infection. STUDY DESIGN: RNA was extracted from plasma samples collected at time points before, during, and after DAA. HPgV RNA abundance was quantified by droplet digital PCR assays targeting the NS5A and 5'UTR domains and confirmed by RT-qPCR. Clinical, demographic and treatment data were analysed. RESULTS: HPgV RNA was detected and quantified in 26 of 100 patients' plasma (26%) before starting DAA. Patients with detectable HPgV were more likely to be male, had higher peak HIV plasma levels, and a history of injection drug use. Patients receiving sofosbuvir/ledipasvir (n = 9) displayed significantly lower HPgV levels at time of DAA completion and had lower post-DAA HPgV rebound​ levels compared to patients receiving sofosbuvir/velpatasvir (n = 11) although both regimens significantly reduced viremia directly following DAA completion. Sustained suppression of HPgV was â€‹also observed among patients (n = 2) receiving pegylated-interferon. CONCLUSIONS: HPgV RNA ​was frequently detected in HCV/HIV co-infected patients and ​was​ supressed by DAA and pegylated interferon therapies with sofosbuvir-ledipasvir showing greatest antiviral activity. These findings suggest potential treatment strategies for HPgV infections​.

GB virus C infection among young, HIV-negative injection drug users with and without hepatitis C virus infection.

Our study examined the association between GB virus C (GBV-C) and (i) hepatitis C virus (HCV) infection status, (ii) biomedical indicators of liver disease (alanine and aspartate aminotransferases) and (iii) HCV RNA level among young injection drug users (IDUs) recruited using street outreach and respondent-driven methods. Cross-sectional and longitudinal analyses were completed. GBV-C (active or resolved) infection was significantly (P < 0.05) more prevalent among HCV antibody-positive (anti-HCV+) (65.1%) than antibody-negative (anti-HCV-) (32.3%) (OR = 3.9, 95% CI: 2.3-6.9) IDUs. The prevalence of resolved GBV-C infection was highest among those with chronic HCV infection (41.9%), followed by those with resolved HCV infection (34.4%) and significantly lower (P < 0.05) among anti-HCV participants (16.9%). Although not statistically significant (P = 0.13), a similar pattern was observed for active GBV-C infection. No association between GBV-C infection status and biomedical indicators of liver disease or HCV RNA level over time was observed. In conclusion, GBV-C infection prevalence was higher among anti-HCV+ compared to anti-HCV- young IDUs, similar to prior studies among older populations. In particular, chronically HCV-infected young IDUs had an increased rate of GBV-C clearance.

Effect of antiretroviral therapy and hepatitis c co-infection on changes in lipid levels in HIV-Infected patients 48 weeks after initiation of therapy.

Hepatitis C virus (HCV) commonly co-infects HIV-infected individuals. Antiretroviral therapy (ART) is associated with elevated serum lipid levels, and HCV infection is associated with low serum lipid levels. Fasting lipid levels were investigated in 1,434 ART-naïve HIV-infected people participating in the AIDS Clinical Trials Group (ACTG) Longitudinal Linked Randomized Trials (ALLRT) protocol who prospectively initiated ART with 3 agents. Subjects with elevated liver-associated enzymes (>5 x ULN) were excluded. Demographics, body mass index, HCV status, CD4 cell count, HIV RNA, liver enzymes, lipid levels, and glucose were assessed before and following 48 weeks of ART. HCV-positive subjects (n = 160; 11%) were older, more likely to be Black, have a history of intravenous drug use (IDU), have higher baseline liver-associated enzyme levels than the HCV-negative group (p < .001 for each), and to have diabetes at baseline (5% vs. 2%, p = .07). Lipid levels rose in both groups following ART, and the differences were not significant except that HDL levels increased significantly more in the HCV-positive group (p = .006). In summary, HCV infection did not appear to provide significant protection against ART-induced hyperlipidemia in this cohort of HIV-infected subjects prospectively enrolled in ART trials, although HDL levels rose to a greater degree.

T cells and HIV-induced T cell syncytia exhibit the same motility cycle.

Ameboid cells ranging in complexity from Dictyostelium amebas to human polymorphonuclear leukocytes (PMNs) translocate in a cyclical fashion. Using computer-assisted motion analysis, we have analyzed the motility of human lymphocytes of the immortal SupT1 cell line and of a peripheral blood mononuclear cell population highly enriched for CD4-positive cells (CD4-enriched PBMCs) on four substrates--plastic, dehydrated rat tail collagen, hydrated rat tail collagen, and bovine aortic endothelium. In addition, we have analyzed the motility on these substrates of syncytia induced by human immunodeficiency virus (HIV) in cultures of both cell types. It is demonstrated that both SupT1 cells and CD4-enriched PBMCs exhibit a motility cycle with a period of 1.6 min that is independent of substrate, independent of average cell velocity, and similar to the periods of translocating Dictyostelium amebas and PMNs. More surprisingly, it is demonstrated that HIV-induced SupT1 and PBMC syncytia with volumes 10 to 100 times those of single cells exhibit the same motility cycle as their single-cell progenitors. These observations support the generality of the motility cycle in animal cells and, for the first time, demonstrate that the cycle is independent of cell size.

Physicians' effectiveness in assessing risk for human immunodeficiency virus infection.

An American Medical Association committee recently recommended that physicians routinely screen patients for behaviors that put patients at risk for human immunodeficiency virus infection, yet there is evidence that this screening does not occur routinely. Faculty, fellows, and residents at a teaching hospital in a midwestern state with a low prevalence of acquired immunodeficiency syndrome were surveyed regarding their experience in screening for human immunodeficiency virus, their training related to substance abuse and human sexuality, and their confidence and ease in addressing such topics with their patients. Results indicated that only 11% routinely screened patients for high-risk behaviors. While most physicians had received training in human sexuality, most had not received training in substance abuse screening. Those trained felt more confident in addressing substance abuse and human sexuality and felt more comfortable in caring for patients known to be infected with human immunodeficiency virus. A concerted effort to encourage human immunodeficiency virus risk assessment by physicians is needed. This should include training opportunities in screening and counseling patients about sexual activities and substance abuse.

Comparison of a neural network approach with five traditional methods for predicting creatinine clearance in patients with human immunodeficiency virus infection.

STUDY OBJECTIVE: To compare the results of an artificial neural network approach with those of five published creatinine clearance (Cl(cr)) prediction equations and with the measured (true) Cl(cr) in patients infected with the human immunodeficiency virus (HIV). DESIGN: Six-month prospective study. SETTINGS: Two university medical centers. PATIENTS: Sixty-five HIV-infected patients: 18 relatively healthy outpatients and 47 inpatients. INTERVENTIONS: All subjects had urine collected for 24 hours to determine Cl(cr). MEASUREMENTS AND MAIN RESULTS: The 16 input variables were age, ideal body weight, actual body weight, body surface area, height, and the following blood chemistries: sodium, potassium, aspartate aminotransferase, alanine aminotransferase, red blood cell count, platelet count, white blood cell count, glucose, serum creatinine, blood urea nitrogen, and albumin. The only output variable was Cl(cr). A training set of 55 subjects was used to develop the relationship between input variables and the output variable. The trained neural network was then used to predict Cl(cr) of a validation set of 10 subjects. Mean differences between predicted Cl(cr) and actual Cl(cr) (bias) were 4.1, 28.7, 29.4, 26.0, 31.8, and 55.8 ml/min/1.73 m2 for the artificial neural network, Cockcroft and Gault, Jelliffe 1, Jelliffe 2, Mawer et al, and Hull et al methods, respectively. CONCLUSION: The accuracy of predicting Cl(cr) in subjects with HIV infection by the artificial neural network is superior to that of the five equations that are currently used in clinical settings.

Effect of a single ethanol exposure on HIV replication in human lymphocytes.

BACKGROUND: Alcoholism is known to cause perturbations in cellular and humoral immunity, and some data suggest that acute alcohol ingestion enhances HIV replication in the lymphocytes of drinkers. METHODS: To study the acute effects of alcohol ingestion on HIV replication, oral ethanol (1 g/kg) was administered to 12 healthy volunteers in a controlled clinical setting. In vitro replication of HIV in the subjects' cultured lymphocytes and changes in lymphocyte phenotypes were evaluated. RESULTS: Statistically significant increases in peripheral lymphocytes and natural killer cell numbers were identified after the initial ethanol trial. HIV replication also increased in the isolated lymphocytes of some subjects after ethanol ingestion, but most subjects in the second trial showed essentially no changes in any of these parameters. CONCLUSIONS: Our results are consistent with either a subtle, study-induced stress-related enhancement in HIV replication or significant individual variation in response to ethanol. The results do not provide evidence for a general increase in HIV replication in the lymphocytes of subjects following a single in vivo ethanol dose of 1 g/kg.

Risk factors for hepatitis C fibrosis: a prospective study of United States veterans compared with nonveterans.

Chronic hepatitis C virus (HCV) infection causes cirrhosis in many infected patients; however, a better understanding of the risk factors for fibrosis progression in high HCV prevalence groups such as US veterans is needed. We wished to compare the demographic, clinical characteristics, and independent variables that influence fibrosis in US veterans vs nonveterans with chronic HCV. HCV-seropositive US veterans (n = 459) and nonveterans (n = 395) prospectively completed a detailed medical, social and occupational questionnaire. Clinical factors for progressive liver disease were compared between veterans and nonveterans and fibrosis stage assessed on liver biopsies (168 veterans and 208 nonveterans). Using polychotomous logistic regression, fibrosis was analysed as both a progressive and categorical outcome to determine independent risk factors for both patient groups. Although veterans were significantly older and had higher lifetime alcohol consumption than nonveterans, their median fibrosis scores did not differ from nonveterans. By univariate analysis, alanine aminotransferase, necroinflammatory activity (NIA), and cryoglobulin positivity were associated with fibrosis in veterans and nonveterans (P < 0.05, all comparisons), whereas steatosis was associated with fibrosis only in nonveterans (P < 0.0001). By multivariate analysis, NIA was an independent risk factor for fibrosis in both groups (P < 0.01). However, fibrosis in nonveterans was also independently associated with steatosis, significant alcohol consumption and age (P < 0.04, all comparisons). Independent risk factors for fibrosis vary among high HCV prevalence groups such as veterans when compared with nonveterans. Understanding specific patient cohort effects is important for determining independent risk factors for disease progression in chronic HCV infection.

Impact of GB virus type C infection on mother-to-child HIV transmission in the Women and Infants Transmission Study Cohort.

BACKGROUND: GB virus type C (GBV-C) viraemia is associated with a beneficial outcome in HIV-infected individuals in several though not all studies. GBV-C viraemia was examined in a matched case-control study of 133 HIV-infected pregnant women who transmitted HIV to their infants ('cases') and 266 non-transmitting controls. METHODS: HIV-infected children and controls were pair-matched for high-risk delivery, race and year of delivery. GBV-C status was determined in maternal plasma samples obtained at or within 3 months of delivery. RESULTS: Pregnant women with GBV-C viraemia (11% of those studied) had lower HIV RNA levels (P=0.01) and higher CD4 percentages (P=0.0006) [corrected] than women without GBV-C. A trend towards decreased mother-to-child transmission in the multivariate analysis was observed among GBV-C viraemic women delivering after highly active antiretroviral therapy (HAART) became available [odds ratio (OR) 0.30, 95% confidence interval (CI) 0.08-1.05; P=0.06], but not in women delivering prior to the widespread use of HAART. CONCLUSIONS: GBV-C viraemia was associated with a beneficial effect on CD4 percentage and HIV RNA level in these pregnant women, and was also associated with a trend towards reduced risk of mother-to-child HIV transmission among women after HAART became available. Further studies with larger or multiple cohorts are necessary to assess possible benefits in this population.

Effect of GB virus C on response to antiretroviral therapy in HIV-infected Brazilians.

OBJECTIVES: GB virus C (GBV-C) infection is associated with delayed mortality in HIV-infected people in most, but not all, studies. Previous investigations of the effect of GBV-C viraemia on response to antiretroviral therapy (ART) were inconclusive. To determine the effect of GBV-C on ART, we retrospectively analysed plasma samples taken from patients in a prospective randomized clinical trial of ART in HIV-positive Brazilians. METHODS: GBV-C viraemia was characterized by testing stored serum samples from 175 participants by reverse transcriptase-polymerase chain reaction (RT-PCR). Subjects were randomized to receive indinavir (n=59), zidovudine and lamivudine (n=58), or zidovudine, lamivudine and indinavir (n=58). The effect of GBV-C viraemia on the average change in HIV viral load and CD4 count following initiation of therapy was evaluated in a multiple regression analysis. RESULTS: The prevalence of GBV-C viraemia was similar to that observed in previous studies (24%). HIV viral load decreased following ART to a significantly greater extent in patients with GBV-C viraemia (by 0.48 log(10) HIV-1 RNA copies/mL, P=0.009, adjusting for age, ART group, and baseline CD4 count). Although there was no significant difference in change in CD4 count between individuals with and without GBV-C viraemia overall, CD4 counts were higher following 48 weeks of therapy in GBV-C viraemic individuals receiving the least potent ART regimen (zidovudine and lamivudine) compared with those without GBV-C infection. CONCLUSIONS: GBV-C viraemia is associated with an enhanced reduction of HIV viral load in response to ART. In this study of treatment-naive individuals during 48 weeks of follow up, patients with GBV-C viraemia had reductions in HIV viral load that were approximately 0.5 log copies/mL greater than those found in patients without GBV-C viraemia. This is similar to reductions observed with nucleoside reverse transcriptase inhibitors.

Effect of early and late GB virus C viraemia on survival of HIV-infected individuals: a meta-analysis.

OBJECTIVES: To conduct a meta-analysis to synthesize the evidence regarding the effect of co-infection with GB virus C (GBV-C) on survival of HIV-infected individuals, and to estimate the effect. METHODS: A Bayesian meta-analysis was conducted to synthesize evidence from eligible studies. Prospective survival studies of HIV-1-infected individuals, with outcome defined as time from baseline to all-cause death, were included and classified by whether GBV-C status was determined in early or late HIV disease. The primary measure was the hazard ratio (HR) of death for HIV-infected individuals with GBV-C infection versus those without GBV-C infection. RESULTS: Eleven studies from eight publications met the inclusion criteria. For studies with GBV-C status measured 2 years or less after HIV seroconversion (912 subjects), the combined HR was 0.88 [95% credible interval (CI) 0.30, 1.50]. For studies with GBV-C status measured more than 2 years after HIV seroconversion (1294 subjects), the combined HR was 0.41 (95% CI 0.23, 0.69). CONCLUSIONS: No conclusive evidence was found of an association between survival and GBV-C infection early in HIV disease. However, when GBV-C infection was present later in HIV disease, a significant reduction in the hazard for mortality was observed for those with co-infection. Potential explanations for this difference include a non-proportional benefit of GBV-C over time, possibly related to clearance of GBV-C infection early in HIV disease. The timing of GBV-C infection appears to account for the contradictory results of studies on the effect of GBV-C coinfection on survival of HIV-infected people.

Effect of primer selection on estimates of GB virus C (GBV-C) prevalence and response to antiretroviral therapy for optimal testing for GBV-C viremia.

GB virus C (GBV-C; also called hepatitis G virus) is a common cause of infection associated with prolonged survival among HIV-infected individuals. The prevalences of GBV-C viremia vary widely in different studies, and there has been poor agreement among different laboratories performing GBV-C RNA detection in quality control studies. To determine the optimal method of measuring GBV-C RNA in clinical samples, samples obtained from 939 HIV-infected subjects were studied using reverse transcription (RT)-PCR methods amplifying four separate regions of the GBV-C genome. Primers amplifying the E2 coding region were 100% specific; however, their sensitivity was only 76.6%. In contrast, primers amplifying three additional conserved regions of the GBV-C genome (the 5' nontranslated region and the nonstructural protein-coding regions 3 and 5A) were more sensitive but produced higher rates of false-positive results. Using low-specificity primer sets influenced the significance of association between GBV-C viremia and response to antiretroviral therapy. Using a quantitative GBV-C RNA method, the GBV-C RNA concentration did not correlate with baseline or set point HIV RNA levels; however, a correlation between negative, low, and high GBV-C RNA levels and increasing reduction in HIV RNA following antiretroviral therapy was observed. Subjects with both GBV-C E2 antibody and viremia had significantly lower GBV-C RNA levels than did viremic subjects without E2 antibody. These studies demonstrate that accurate detection of GBV-C RNA by nested RT-PCR requires the use of primers representing multiple genome regions. Analyses based on testing with single primers do not lead to reliable conclusions about the association between GBV-C infection and clinical outcomes.

Host immune response to hepatitis A virus.

Hepatitis A virus (HAV) is transmitted by the fecal-oral route. The virus crosses through the gastrointestinal tract by an uncharacterized mechanism and travels to the liver, where it replicates in hepatocytes. It is released into the bloodstream and is simultaneously present in the bile and shed in the feces. Fecal shedding and viremia are maximal at the onset of liver function abnormalities and terminate about the time humoral immunity is detected, approximately 28 days after exposure. IgM, IgA, and IgG anti-HAV antibodies are usually present at onset of symptoms. Although the IgM response becomes undetectable usually within 6 months, IgG responses frequently persist for life, providing protection against reinfection. Pre- and postexposure immunization with pooled human serum immunoglobulin (ISG) is approximately 90% effective in preventing hepatitis A. Recipients of ISG have very low levels of detectable anti-HAV antibodies, and vaccines that elicit anti-HAV levels comparable with those produced by ISG should confer similar protection.

Passive immunization against hepatitis A.

Administration of human serum immune globulin (Ig) is an effective means of protecting individuals against hepatitis A virus (HAV) infection and disease. Several large field studies have demonstrated that if given before exposure, Ig will prevent infection with HAV. Furthermore, if Ig is given during the incubation period of hepatitis A, the severity of infection may be reduced and potentially clinical infections may be converted into subclinical ones. Although uncommon, infection which occurs in the presence of circulating antibody may occasionally lead to passive-active immunity. Unfortunately, the duration of Ig protection is dose dependent, and high dose administration provides less than six months protection. Ig preparations contain HAV antibodies at levels detectable by commercial immunoassays; however, recipients of Ig do not have detectable levels of HAV antibodies when tested by the same method. Using more sensitive immunoassays and neutralization assays, low titres of HAV antibody can be detected in Ig recipients. Since Ig provides approximately 90% efficacy in preventing hepatitis A, it would appear that very low levels of HAV antibody are needed to prevent infection. Consequently, measurement of HAV antibodies elicited by HAV vaccines should provide a reasonable method to evaluate their immunogenicity and predict their efficacy.