Seroprevalence of hepatitis B, C, and D and associated factors in the semi-isolated Yanomami Amazonian indigenous community.

BACKGROUND: Viral hepatitis is a significant health concern among indigenous population in the Americas. In Brazil, reports find high endemicity of HBV and HDV infections has been reported in several indigenous groups. However, few studies have documented the prevalence of HBV, HCV and HDV in the Yanomami. In this study, the prevalence of hepatitis B, C, and D serological markers and potential risk factors were investigated to provide guidance for the development of strategies aimed at reducing viral transmission in the Yanomami indigenous villages. METHODS: This cross-sectional study was carried out in March 2015 and included 430 individuals from four Yanomami villages: Alapusi (n = 78), Castanha/Ahima (n = 126), Gasolina (n = 105), and Taibrapa (n = 121). A rapid test was used for detection of HBsAg and anti-HCV and chemiluminescent immunoassay for anti-HBs, anti-HBc, and anti-HDV antibodies. RESULTS: HBsAg, anti-HBc, and anti-HBs were detected in 8.8, 45.5, and 49.4% of the participants, respectively. The estimated HBV status: current infection 9.6% (38/395); resolved infection 43.3% (171/395); vaccine immunity 20.5% (81/395), and susceptible to HBV 26.6% (105/395). Gasolina presented the lowest prevalence of HBV infection (6.5%) and the highest prevalence of vaccine immunity (26.9%). Children < 15 years old were highly susceptible to infection, as 53.1% did not have antibodies to HBV, while more than 80% of individuals over 45 years of age had been exposed to HBV. The markers for HDV were founded among 12.5% (4/32) of the HBsAg carriers. Anti-HCV was identified in all villages, with the highest prevalence in Alapusi (5.1%). Possible risk factors such as the use of piercings, tattoos, and contact with prospectors showed no statistical difference between the groups. CONCLUSIONS: Viral hepatitis B and serological markers for HCV and HDV were found to be widely distributed among the Yanomami indigenous community, while the prevalence of vaccine immunity to HBV was low. This finding reinforces the importance of promoting systematized diagnostic and vaccination strategies in indigenous communities. Our data confirm that isolated and difficult-to-reach indigenous communities lack appropriate access to diagnosis, treatment, and vaccination.

Feasibility of dried blood spot for hepatitis C diagnosis in vulnerable subjects and people living in remote areas from Brazil.

BACKGROUND: Agile, accessible and cheap diagnosis of hepatitis C virus (HCV) infection is essential to achieve the elimination of this infection, worldwide, as mandated by the World Health Organzation as part of its strategy for 2030. Dried blood spots (DBS) can be an attractive alternative for sample collection among people living in remote areas and vulnerable populations due to the less invasive collection, its biosafety, and storage & transportation of samples at room temperature. DESIGN: This study aims to estimate the usefulness of dried blood spot samples for the diagnosis and the assessment of HCV infection rates in three different settings in Brazil. Cross-sectional analysis of a sample collection from different populations, aiming to assess the performance of the testing algorithms and respective procedures among different populations with diverse background infection rates. METHODS: We reported the evaluation of DBS as alternative samples for detecting anti-HCV in different groups in real life conditions: (I) Vulnerable subjects living in remote areas of Southeast, North and Northeast Brazil (n = 1464); (II) Beauticians (n = 288); (III) People who use non-injectable drugs (n = 201); (IV) patients referred to outpatient care (n = 275). RESULTS: General assay accuracy was 99%, with a weighted kappa value of 0.9, showing an excellent performance. Sensitivities ranged from 87.5% to 100.0% between groups and specificities were above 99.2%. A total of 194 individuals had HCV RNA in serum and concordance of anti-HCV detection in DBS was 98.4%. CONCLUSIONS: DBS samples could be used for anti-HCV detection in different populations recruited in real life conditions and ambulatory settings, with a high overall sensitivity and specificity.

Association of Pre-S/S and Polymerase Mutations with Acute and Chronic Hepatitis B Virus Infections in Patients from Rio de Janeiro, Brazil.

Several hepatitis B virus (HBV)-related factors, including the viral load, genotype, and genomic mutations, have been linked to the development of liver diseases. Therefore, in this study we aimed to investigate the influence of HBV genetic variability during acute and chronic infection phases. A real-time nested PCR was used to detect HBV DNA in all samples (acute, n = 22; chronic, n = 49). All samples were sequenced for phylogenetic and mutation analyses. Genotype A, sub-genotype A1, was the most common genotype in the study population. A total of 190 mutations were found in the pre-S/S gene area and the acute profile revealed a greater number of nucleotide mutations (p < 0.05). However, both profiles contained nucleotide mutations linked to immune escape and an increased risk of hepatocellular carcinomas (acute, A7T; chronic, A7Q). Furthermore, 17 amino acid substitutions were identified in the viral polymerase region, including the drug resistance mutations lamivudine and entecavir (rtL180M), with statistically significant differences between the mutant and wild type strains. Owing to the natural occurrence of these mutations, it is important to screen for resistance mutations before beginning therapy.

Serological and molecular characterization of hepatitis B virus infection in chronic kidney disease patients from Rio de Janeiro, Brazil.

Chronic kidney disease (CKD) patients undergoing hemodialysis (HD) are more vulnerable to blood-borne viral infections due to frequent invasive procedures. Hepatitis B virus (HBV) infection in this cohort of patients has been a matter of concern worldwide. The objective of this cross-sectional study was to evaluate the frequency of serological markers for hepatitis B, and the occurrence of overt and occult HBV infection (OBI) and its molecular characterization in serum samples from 644 CKD patients in HD units located in Rio de Janeiro, Brazil, from 2013 to 2017. HBV DNA was investigated in HBsAg reactive and "anti-HBc alone" samples to determine infecting genotypes and genetic relatedness between sequences. The prevalence of serological markers HBsAg+, anti-HBc alone, anti-HBc+/anti-HBs+, anti-HBs+, anti-HBc/anti-HBs/HBsAg were 5.9%, 2.8%, 30.7%, 26.6%, 34.0%, respectively. HBV DNA was detected in 39.5% (15/38) of the HBsAg+ and in 5/18 (27.8%) of the "anti-HBc alone" individuals, indicating a high prevalence of OBI within this group. We found a higher prevalence of HBV/A1 (65%), followed by HBV/D3 (20%), and HBV/A2 (15%). Bayesian MCC tree with a highly supported clade, genetic distance comparison, and identical nucleotide sequences suggested a nosocomial spread of HBV in some units. The high prevalence of HBV infection and low number of individuals immune to infection reinforces the need for vaccination in this group. The presence of closely related strains in the same HD unit reinforces the importance of continuous improvement of safety control measures and laboratory surveillance of serological markers to prevent the risk of infection and transmission of HBV.

Enduring Challenges despite Progress in Preventing Mother-to-Child Transmission of Hepatitis B Virus in Angola.

Sub-Saharan Africa has one of the highest rates of hepatitis B virus (HBV) infection globally, with an incidence of 1.5 million and 0.8 million yearly deaths, which drives synergistic efforts towards its elimination. To assess the risk of mother-to-child transmission of HBV infection, a cross-sectional study was conducted on 1012 pregnant women in Angola to investigate HBV serological and molecular profiles. The prevalence of HBV was 8.7% (n = 88), with hepatitis B core IgM antibody (anti-HBc IgM) positivity identified in 12.8%, hepatitis B "e" antigen (HBeAg) positivity in 30%, and HBV DNA ≥ 200,000 IU/mL in 28.2%. Family tracking studied 44 children, of which 11 (25%) received at least two doses of the hepatitis B vaccine. HBV was detected in 10/44 (22.7%) children, with vaccination reported in one infected child. Further testing identified anti-HBc IgM positivity in 3/10 (30%), HBeAg positivity in 55%, and both seromarkers in 20%. The results revealed the importance of antenatal HBV screening, antiviral prophylaxis for mothers with high viral loads or HBeAg positivity, and timely first-dose hepatitis B vaccines in newborns. Anti-HBc IgM positivity among pregnant women and children highlights prophylactic measures worth considering, including antenatal hepatitis B vaccination and catch-up vaccination to young children.

Optimization of a real time PCR methodology for HCV RNA quantification in saliva samples.

Saliva may be an alternative biological specimen to expand HCV detection. This study aims to evaluate an in-house quantitative RT-PCR for HCV RNA quantification in saliva. A total of 80 individuals (56 anti-HCV/HCV RNA + and 24 negative controls) donated serum and saliva, that were tested using an in-house quantitative PCR for HCV RNA. The median viral load was 4.77 log10 copies/mL (1.04-7.0 log10 copies/mL) in serum and 2.31 log10 copies/mL (1.0-3.84 log10 copies/mL) in saliva. A sensitivity of 80 % and specificity of 100 % were observed for HCV detection in saliva, which demonstrates the usefulness of in-house real-time PCR to quantify HCV RNA in saliva samples, which might increase the access of molecular diagnosis of HCV in laboratories that lack complex infrastructures for molecular testing and in individuals with poor venous access.

Immunological and virological aspects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and hepatitis C virus.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can generate a systemic inflammatory response, characterized by a cytokine storm and associated with an exaggerated release of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-17, all of which can affect the liver. Here, we aimed to evaluate the cytokine profiles of patients suffering from coronavirus disease (COVID)-19 and/or hepatitis. We subjected 87 patients to serology and/or polymerase chain reaction analysis for the hepatitis C virus. They were also tested for TNF-α, IL-6, and IL-17 using commercial immunoassay kits. The test results of the COVID-19/hepatitis C patients (n = 8) were compared with that of the negative controls (n = 28), hepatitis C patients (n = 29), and COVID-19 patients (n = 22). All COVID-19 patients (mono- and coinfected) expressed high levels of cytokines. The COVID-19/hepatitis patients exhibited higher levels of IL-6 (6.33 ± 3.9 pg/ml) and IL-17 (102.23 ± 2.7 pg/ml); however, TNF-α values were lower (68.08 ± 15.88 pg/ml), as compared with that of the hepatitis patients (p < 0.001), and lower than that of the COVID-19 patients and exceptionally for TNF-α (p < 0.05). These data highlight the importance of monitoring patients with hepatitis and COVID-19.

Peripheral blood iNKT cell activation correlates with liver damage during acute hepatitis C.

Invariant NK T (iNKT) cells are implicated in viral clearance; however, their role in hepatitis C virus (HCV) infection remains controversial. Here, iNKT cells were studied during different stages of HCV infection. iNKT cells from patients with acute HCV infection and people who inject drugs (PWID) with chronic or spontaneously resolved HCV infection were characterized by flow cytometry. In a longitudinal analysis during acute HCV infection, frequencies of activated CD38+ iNKT cells reproducibly declined in spontaneously resolving patients, whereas they were persistently elevated in patients progressing to chronic infection. During the first year of infection, the frequency of activated CD38+ or CD69+ iNKT cells strongly correlated with alanine transaminase levels with particularly pronounced correlations in spontaneously resolving patients. Increased frequencies of activated iNKT cells in chronic HCV infection were confirmed in cross-sectional analyses of PWID with chronic or spontaneously resolved HCV infection; however, no apparent functional differences were observed with various stimulation protocols. Our data suggest that iNKT cells are activated during acute hepatitis C and that activation is sustained in chronic infection. The correlation between the frequency of activated iNKT cells and alanine transaminase may point toward a role of iNKT cells in liver damage.

Cytokines profile in patients with acute and chronic hepatitis B infection.

Hepatitis B virus (HBV) is one of the leading causes of acute and chronic hepatitis and represents a serious public health threat. Cytokines are important chemical mediators that regulate the differentiation, proliferation, and function of immune cells, with accumulating evidence indicating that the inadequate immune responses are responsible for the elimination or persistence of HBV. This study aimed to determine the cytokine profiles (IFN-γ, TNF-α, IL-2, IL-4, IL-6, IL-10, and IL-17A) during HBV infection and investigate their association with genotypes. A total of 66 plasma samples, 19 from patients with acute and 47 with chronic hepatitis B infection, were subjected to biochemical tests, nested-PCR, and real-time PCR, with cytokines evaluated using a commercial BD Cytometric Bead Array Human Th1/Th2/Th17 Cytokine Kit. Healthy controls (10 individuals) were selected from blood donors with no history of liver diseases. No correlation was found between genotypes, viral load, and cytokines analyzed. All cytokines showed higher levels of production among infected individuals when compared with the control group. A positive correlation classified as moderate to strong was found between cytokines IFN-γ, TNF, IL-10, IL-6, IL-4, and IL-2 through the Spearman correlation coefficient. TNF (P = 0.009), IL-10 (P < 0.001), and IL-6 (P < 0.001) levels were higher in acute individuals compared with chronic and control groups. Theses cytokines could be involved in the elimination of virus and protection against chronicity.

Serological and molecular characterization of hepatitis B virus infection in chronic kidney disease patients from Rio de Janeiro, Brazil

ABSTRACT Chronic kidney disease (CKD) patients undergoing hemodialysis (HD) are more vulnerable to blood-borne viral infections due to frequent invasive procedures. Hepatitis B virus (HBV) infection in this cohort of patients has been a matter of concern worldwide. The objective of this cross-sectional study was to evaluate the frequency of serological markers for hepatitis B, and the occurrence of overt and occult HBV infection (OBI) and its molecular characterization in serum samples from 644 CKD patients in HD units located in Rio de Janeiro, Brazil, from 2013 to 2017. HBV DNA was investigated in HBsAg reactive and "anti-HBc alone" samples to determine infecting genotypes and genetic relatedness between sequences. The prevalence of serological markers HBsAg+, anti-HBc alone, anti-HBc+/anti-HBs+, anti-HBs+, anti-HBc/anti-HBs/HBsAg were 5.9%, 2.8%, 30.7%, 26.6%, 34.0%, respectively. HBV DNA was detected in 39.5% (15/38) of the HBsAg+ and in 5/18 (27.8%) of the "anti-HBc alone" individuals, indicating a high prevalence of OBI within this group. We found a higher prevalence of HBV/A1 (65%), followed by HBV/D3 (20%), and HBV/A2 (15%). Bayesian MCC tree with a highly supported clade, genetic distance comparison, and identical nucleotide sequences suggested a nosocomial spread of HBV in some units. The high prevalence of HBV infection and low number of individuals immune to infection reinforces the need for vaccination in this group. The presence of closely related strains in the same HD unit reinforces the importance of continuous improvement of safety control measures and laboratory surveillance of serological markers to prevent the risk of infection and transmission of HBV.

Transfusion Transmissible Infections in Blood Donors in the Province of Bié, Angola, during a 15-Year Follow-Up, Imply the Need for Pathogen Reduction Technologies.

Transfusion transmissible infections (TTIs), caused by hepatitis B virus (HBV), human immunode-ficiency virus (HIV), hepatitis C virus (HCV), and syphilis, have a high global impact, especially in sub-Saharan Africa. We evaluated the trend of these infections over time in blood donors in Angola. A retrospective cross-sectional study was conducted among blood donors in Angola from 2005 to 2020. Additionally, frozen samples obtained from blood donors in 2007 were investigated to identify chronic HCV carriers and possible occult HBV infection (OBI). The overall prevalence of HBV, HCV, HIV, and syphilis was 8.5, 3, 2.1, and 4.4%, respectively, among 57,979 blood donors. HBV was predominant among male donors, while the remaining TTIs were predominant among women. Donors >50 years had a significantly high prevalence for all TTIs. Chronic HCV infection was ab-sent in 500 samples tested and OBI was present in 3%. Our results show the continued high prev-alence of TTIs among blood donors in Angola. Most infections showed a significantly low preva-lence in years with campaigns seeking voluntary blood donors, thus, reinforcing the importance of this type of donor to ensure safe blood. Africa, with a high prevalence of diverse pathogens, should consider cost-effective pathogen reduction technologies, once they are commercially accessible, to increase the availability of safe blood.

Clinical and laboratory characteristics of hepatitis C and COVID-19 coinfection: Prolonged RNA shedding in nonhospitalized case.

Nonhospitalized COVID-19 and hepatitis C-coinfected patient presented prolonged RNA shedding and mild course of infection. This finding demonstrated the importance of long follow-up of these patients.

Applicability of oral fluid samples for tracking hepatitis B virus mutations, genotyping, and phylogenetic analysis.

Little is known about the usefulness of saliva samples for hepatitis B virus (HBV) genotyping and mutation analysis. The aim of this study was to evaluate the usefulness of oral fluid samples to determine HBV genotype distribution, S/polymerase mutations, and HBV subpopulation diversity among chronically HBV-infected individuals. Serum and oral fluid samples were obtained from 18 individuals for PCR and nucleotide sequencing of the HBV surface antigen gene. Biochemical analysis of liver enzymes (ALT, AST, GGT) and HBV, HCV, and HIV serological tests were also performed. All serum samples were HBsAg (+), anti-HBc (+), and anti-HBs (-); 55.6% were HBeAg (+)/anti-HBe (-), and 11.1% were anti-HIV (+). The mean HBV DNA viral load was 6.1 ± 2.3 log IU/mL. The HBV genotype distribution was as follows: A, 72.2%; D, 11.1%; E, 5.6%; F, 11.1%. A concordance of 100% in genotype classification and 99.8% in sequence similarity between paired oral fluid and serum samples was observed. HBsAg mutations were detected in all samples, but no resistance mutations were found in the polymerase gene. This study demonstrates that oral fluid samples can be used reliably for tracking HBV mutations, genotyping, and phylogenetic analysis. This could be important for molecular epidemiology studies with hard-to-reach populations.

Dried blood spot sampling as an alternative for the improvement of hepatitis B and C diagnosis in key populations.

BACKGROUND: To achieve the elimination of hepatitis B and C, there is an urgent need to develop alternative strategies to increase the access of diagnosis, particularly among key populations such as people living with human immunodeficiency virus (HIV), individuals with coagulopathies and chronic kidney disease (CKD) patients. AIM: To evaluate the use of dried blood spot (DBS) in the detection of hepatitis B virus (HBV) and hepatitis C virus (HCV) markers. METHODS: A total of 430 individuals comprised of people living with HIV, coagulopathies and CKD provided paired serum and DBS samples. HBsAg, anti-HBc and anti-HCV were tested in those samples using a commercial electrochemiluminescence. Demographic and selected behavioral variables were evaluated to assess possible association with HBV and HCV positivity. RESULTS: Using DBS, HBsAg prevalence varied from 3.9% to 22.1%, anti-HBc rates varied from 25.5% to 45.6% and anti-HCV positivity ranged from 15.9% to 41.2% in key populations. Specificities of HBV and HCV tests using DBS varied from 88.9% to 100%. The HBsAg assay demonstrated the best performance in CKD and coagulopathy individuals and the anti-HCV test had a sensitivity and specificity of 100% in people living with HIV. Accuracy of HBV and HCV detection in DBS varied from 90.2% to 100%. In the CKD group, HBsAg positivity was associated with infrequent use of condoms, and anti-HBc positivity was associated with sharing nail cutters/razors/toothbrushes. Anti-HCV reactivity was positively associated with a history of transplantation and length of time using hemodialysis in both specimens. In people living with HIV, only the male gender was associated with anti-HBc positivity in serum and DBS. CONCLUSION: DBS with electrochemiluminescence are useful tools for the diagnosis and prevalence studies of hepatitis B and C among key populations and may increase the opportunity to foster prevention and treatment.

Cytokine, Genotype, and Viral Load Profile in the Acute and Chronic Hepatitis B.

Several hepatitis B virus (HBV) factors, including viral load, genotype, genome mutations, and cytokine production, have been reported to be associated with different risks of progression of liver disease. The aim of this study was to verify if there is an association among the levels of cytokines (interleukin [IL]-35, IL-6, IL-17A, interferon [IFN]-γ) in the plasma, viral load, and the different genotypes of HBV in patients with acute or chronic hepatitis B. Methods: 49 serum samples, 20 from acute and 29 from chronic cases, were submitted to a real-time and nested-polymerase chain reaction to quantify, detect, and genotype HBV DNA. The cytokines IL-35, IL-6, IL-17A, and IFN-γ were detected by an enzyme-linked immunosorbent assay (ELISA). The median viral load was 3.15 log10 IU DNA/mL and 2.90 log10 IU DNA/mL for acute and chronic patients, respectively. Genotype A, D, E, and F were identified in chronic carriers of HBV infection, while only genotype A and F were identified in individuals with acute infection. IFN-γ (p = 0.024) and IL-17A (p = 0.046) levels were significantly increased in chronic patients and IL-6 and IL-35 were higher in patients with acute infection, however, without statistical difference. IL-17A and IFN-γ can be modulating proinflammatory effects and inducing hepatocellular damage, in chronic patients, and IL-6 and IL-35 may be involved in viral elimination and protection against chronicity during the acute phase of infection. These results can contribute to understanding of the complex regulatory mechanisms of the host antiviral response related to cytokine production during acute and chronic HBV infection.

Evaluation of accuracy of hepatitis B virus antigen and antibody detection and relationship between epidemiological factors using dried blood spot.

Dried blood spots (DBS) testing might increase the access for Hepatitis B virus (HBV) diagnosis, but little is known about the performance of these assays in real life conditions. This study aims to evaluate the diagnostic accuracy of HBsAg, anti-HBc and anti-HBs detection in DBS in clinical settings and field studies and to evaluate demographic and risk behaviour according the presence of HBsAg and anti-HBc. Paired sera and DBS samples were obtained from 2309 individuals from 3 groups, defined as follows: G1: clinical setting (n = 5-19), G2: general population (n = 1305) and G3: vulnerable individuals that could be more exposed to blood contact (n = 485). Sera and DBS were tested using commercial enzyme immunoassay (EIA), with some modifications added. Using DBS samples, the specificity values were above 90 % for HBsAg and anti-HBc in all groups and for anti-HBs range from 58.6%-85%. HBsAg testing had the best performance in GI (sensitivity = 84.4 %) and among those samples that the paired serum also presented anti-HBc marker (sensitivity = 91.6 %). High sensitivity of anti-HBc testing in DBS samples was observed in GI (80.8 %) and among HBV active cases (HBsAg+/anti-HBc+) (98.4 %). Testing of anti-HBs in DBS showed the highest sensitivity in GIII (65.5 %), in previous HBV exposed and cured individuals and when serum titers were above 100 IU/mL (86.7 %). DBS samples could be used for screening and prevalence studies for HBsAg and anti-HBc, particularly in clinical settings and among HBV active cases in field studies.

Analysis of Humoral Immune Responses in Chikungunya Virus (CHIKV)-Infected Patients and Individuals Vaccinated With a Candidate CHIKV Vaccine.

BACKGROUND: Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes severe flu-like symptoms. The acute symptoms disappear after 1 week, but chronic arthralgia can persist for years. In this study, humoral immune responses in CHIKV-infected patients and vaccinees were analyzed. METHODS: Alphavirus neutralization activity was analyzed with pseudotyped lentiviral vectors, and antibody epitope mapping was performed with a peptide array. RESULTS: The greatest CHIKV neutralization activity was observed 60-92 days after onset of symptoms. The amount of CHIKV-specific antibodies and their binding avidity and cross-reactivity with other alphaviruses increased over time. Chikungunya virus and o'nyong-nyong virus (ONNV) were both neutralized to a similar extent. Linear antibody binding epitopes were mainly found in E2 domain B and the acid-sensitive regions (ASRs). In addition, serum samples from healthy volunteers vaccinated with a measles-vectored chikungunya vaccine candidate, MV-CHIK, were analyzed. Neutralization activity in the samples from the vaccine cohort was 2- to 6-fold lower than in samples from CHIKV-infected patients. In contrast to infection, vaccination only induced cross-neutralization with ONNV, and the E2 ASR1 was the major antibody target. CONCLUSIONS: These data could assist vaccine design and enable the identification of correlates of protection necessary for vaccine efficacy.