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.

Epidemiology of hepatitis B and C virus infection in Central West Argentina.

Little information is available regarding the prevalence of viral hepatitis in Central West Argentina. This study aims to give new information regarding HBV and HCV prevalence, genotypes, and risk factors in Central West Argentina and the suitability of dried blood spot (DBS) sampling for HBV and HCV screening. METHODS: A total of 622 individuals were included; the mean age was 36.6 ± 14.3 years and 55.4% were females. HBV and HCV markers were detected using serological and molecular analysis, and risk factors were evaluated using statistical analysis. RESULTS: Using serum samples, the HBsAg prevalence was 1.8%, the rate of HBV exposure (anti-HBc positivity) was 5.3%, and the rate of HBV immunity was 34.9%. HBV DNA was found in four out of 11 HBsAg+ samples, and the viruses in three of these samples were classified as genotypes A1, A2 and F2a. Multivariate analysis showed that anti-HBs positivity was associated with the level of schooling and history of HBV vaccination. The anti-HCV prevalence was 2.6%, and HCV RNA was found in 11 samples, seven of which contained viruses of genotypes 1a (n = 2), 1b (n = 3) and 2 (n = 2). The sensitivity of the DBS assay for HBsAg, anti-HBc, and anti-HCV was 100%, 66.6%, and 75%, respectively, and the specificity was above 98% for all markers when compared to serum. CONCLUSION: A low rate of HBV immunity was observed, demonstrating the importance of HBV vaccination. High HCV prevalence was found, and HCV 1b was closely related to other Argentinian isolates. Finally, the performance of DBS testing in this population needs more optimization to increase its sensitivity and specificity.

Genetic characterization of the parvovirus full-length VP2 gene in domestic cats in Brazil.

Feline parvovirus (FPV) and canine parvovirus (CPV) are over 98% identical in their DNA sequences, and the new variants of CPV (2a/2b/2c) have gained the ability to infect and replicate in cats. The aim of this study was to determine the genetic diversity in the VP2 gene of parvovirus strains circulating in domestic cats in Brazil during a 10-year period (2008-2017). For parvovirus screening, specific PCR was performed, and 25 (34.7%) of 72 cats tested positive. The PCR-positive samples were further subjected to full-length VP2 sequencing (1755 bp), and eight sequences (36%) were characterized as FPV, seven (28%) as CPV-2a and (32%) nine (36%) as CPV-2b. One sequence (RJ1085/11) showing typical CPV amino acid (aa) at residues 80 R, 93 N, 103 A, 232 I, and 323 N could not be characterized at this time. The sequences in this study displayed aa changes previously described for FPV (A14T, A91S, I101T, N564S, and A568G) from cats and CPV-2a/2b (S297N and Y324L) from dogs. However, the Y324L mutation has not yet been reported in any CPV-2a/2b strains from cats. Phylogenetic analysis supported the division of these sequences into two well-defined clades, clade 1 for FPV and clade 2 for CPV2a/2b. Unusually, the sequence RJ1085/11 was grouped separately. Two recombination breakpoints were detected by Bootscan and 3Seq methods implemented in the RDP4. This study is the first report of CPV-2a/2b in cats in Brazil. The detection of FPV strains with mutations characteristic of CPV indicates that Brazilian FPV strains have undergone genetic changes.

Could Herpesviridae be the cause of severe acute hepatitis of unknown origin in children?

INTRODUCTION: Severe acute hepatitis (SAH) is defined by a severe inflammation of hepatocytes in the liver parenchyma which can lead to an acute liver failure, a clinical condition with high mortality rate that can be triggered by several factors but is usually associated to hepatotropic viruses' infection. In 2022, cases of children with severe acute hepatitis of unknown origin hospitalized in Glasgow, Scotland, were reported. Possible causes of this condition include, but are not limited to, undiagnosed viral (and non-viral) infections, autoimmune hepatitis, drug and/or chemical toxicity, mitochondrial chain respiratory and metabolic disorders. AREAS COVERED: Herpesviruses can cause severe acute hepatitis, but little is known about the role and the mechanisms of herpesviruses as a causative agent of this type of hepatitis. We review the role of herpesviruses as causative agent of SAH in children and other possible mechanisms involved in this disease. EXPERT OPINION: Differential diagnosis for herpesvirus in SAH should be implemented in all settings. Alternative fluids, such as saliva and dried blood, could be used in the diagnosis to overwhelm the availability of biological specimens at sufficient volume. In the future, genetic studies could also be added to increase the knowledge about severe acute hepatitis in children.

Differences in risky sexual behaviors and HIV prevalence between men who have sex with men and transgender women in the Midwest Brazil.

Men who have sex with men (MSM) and transgender women (TW) are disproportionally affected by HIV infection. This cross-sectional study evaluated the HIV-1/2 prevalence, risk factors and HIV molecular features of MSM and TW from Midwest Brazil. Four hundred and thirty participants (278 MSM and 152 TW) from Mato Grosso do Sul, Brazil, were interviewed and tested for HIV-1/2 infection between November 2011 and September 2013. Participants who were assigned male at birth, older than 18 years old and self-declared as MSM or TW were recruited from LGBT+ associations, as well as public (parks, square, streets, etc) and private [nightclubs, saunas, brothels, etc] places. The prevalence of HIV-1 was 14.4% (9.0% among MSM and 24% among TW; p<0.001). The factor independently associated with HIV-1 infection among MSM was being 30 years-old or older. Among TW, having suffered sexual coercion, lifetime syphilis infection and hepatitis C virus exposure were associated with HIV-1 infection. Phylogenetic analyses classified 65% sequences as subtype B and 35% as possible recombinants. All but one recombinant sample were from TW individuals. High HIV-1 prevalences were observed in both groups, highlighting the urgent need to devise specific HIV interventions targeting these key populations. Notably, TWs are more vulnerable to HIV infection, which was associated with sexual violence and co-infection with other STIs. With regard to MSM, being 30 years old or older was significanty associated to HIV, reinforcing the idea that MSM are less exposed [or exposed later] to STIs than TWs, although MSM are clearly more vulnerable than the general population.

Genetic diversity of hepatitis B virus quasispecies in different biological compartments reveals distinct genotypes.

The selection pressure imposed by the host immune system impacts hepatitis B virus (HBV) quasispecies variability. This study evaluates HBV genetic diversity in different biological fluids. Twenty paired serum, oral fluid, and DBS samples from chronic HBV carriers were analyzed using both Sanger and next generation sequencing (NGS). The mean HBV viral load in serum was 5.19 ± 4.3 log IU/mL (median 5.29, IQR 3.01-7.93). Genotype distribution was: HBV/A1 55% (11/20), A2 15% (3/20), D3 10% (2/20), F2 15% (3/20), and F4 5% (1/20). Genotype agreement between serum and oral fluid was 100% (genetic distances 0.0-0.006), while that between serum and DBS was 80% (genetic distances 0.0-0.115). Two individuals presented discordant genotypes in serum and DBS. Minor population analysis revealed a mixed population. All samples displayed mutations in polymerase and/or surface genes. Major population analysis of the polymerase pointed to positions H122 and M129 as the most polymorphic (≥ 75% variability), followed by V163 (55%) and I253 (50%). Neither Sanger nor NGS detected any antiviral primary resistance mutations in the major populations. Minor population analysis, however, demonstrated the rtM204I resistance mutation in all individuals, ranging from 2.8 to 7.5% in serum, 2.5 to 6.3% in oral fluid, and 3.6 to 7.2% in DBS. This study demonstrated that different fluids can be used to assess HBV diversity, nonetheless, genotypic differences according to biological compartments can be observed.

Dried blood spot sampling for hepatitis B virus quantification, sequencing and mutation detection.

Hepatitis B virus (HBV) diagnosis is performed on serum samples, but the access to this diagnosis is difficult in low-income regions. The use of dried blood spot (DBS) samples does not require special structure for collection, storage or transport. This study evaluates the use of DBS for detection, quantification and sequencing of HBV DNA using in-house techniques. Two study groups were included: 92 HBsAg + individuals and 49 negative controls. Serum and DBS samples were submitted to quantitative and qualitative in-house PCR for S/pol genes, sequencing and phylogenetic analyses. Total of 84 serum samples were successfully amplified. Of them, 63 paired DBS were also positive in qualitative PCR. Qualitative PCR in DBS presented a sensitivity of 75% and specificity of 100% (Kappa = 0.689). Quantitative PCR in DBS presented a detection limit of 852.5 copies/mL (250 IU/mL), sensitivity of 77.63% and specificity of 100% (Kappa = 0.731). A total of 63 serum samples and 36 DBS samples were submitted to sequencing, revealing the circulation of genotypes A (65.08%), D (4.8%), E (3.2%) and F (27%) with 100% of correspondence between serum and DBS. All sequenced samples displayed polymorphisms in HBsAg gene. An HIV-coinfected patient presented the rtM204V/I-rtL180M double resistance mutation in serum and DBS. In conclusion, DBS is an alternative to detect, quantify and characterize HBV DNA, being a possibility of increasing diagnosis in low-income settings, closing gaps in HBV control.

HAV infection in Brazilian men who have sex with men: The importance of surveillance to avoid outbreaks.

BACKGROUND: Hepatitis A is a fecal-oral infection caused by hepatitis A virus (HAV). Men who have sex with men (MSM) and transgender women (TW) have been reported as target groups for HAV infection. This study aimed to determine the seroprevalence, risk factors, and circulating strains associated with HAV infection among MSM and TW in Central Brazil. METHODS: A cross-sectional study was conducted from November 2011 to September 2013. Serum samples were collected from 425 individuals for anti-HAV antibody testing and HAV molecular characterization. Of them, 149 (35.1%) participants were self-identified as transgender women. Statistical analysis was performed to evaluate the risk factors of HAV seropositivity. RESULTS: The seroprevalence of HAV exposure was 69.7% (95% Confidence Interval: 65.3-74.0%). Serological evidence of HAV was significantly higher in participants who self-identified as transgender women (83.2%) than MSM (62.3%). Increasing age, non-white race, and lower monthly household income were independently associated with HAV exposure among MSM. Only lower monthly household income was independently associated with HAV exposure among TW. One anti-HAV IgM positive sample, from a transgender woman (0.2%), was detected and classified as subgenotype IA. CONCLUSIONS: High HAV prevalence was observed, markedly among TW. Considering the risky sexual behaviors this population is exposed to, HAV vaccination and prevention programs targeting this population should be considered to prevent outbreaks and the burden of the disease.

Author Correction: Hepatitis B: changes in epidemiological features of Afro-descendant communities in Central Brazil.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

High prevalence of Hepatitis C Virus infection among people who use crack cocaine in an important international drug trafficking route in Central-West Region Brazil.

In this study, the prevalence rate, associated risk factors and genetic diversity of hepatitis C virus (HCV) infection were determined among people who use crack from an international drug trafficking route in Central-West, Brazil. Blood samples were collected from 700 users of crack from Campo Grande and two border cities of Mato Grosso do Sul State and tested for HCV infection using serological and molecular testing methodologies. Anti-HCV was detected in 31/700 (4.5%, 95% CI: 2.9-6.0%) and HCV RNA in 26/31 (83.9%) of anti-HCV positive samples. Phylogenetic analysis of three HCV sub-genomic regions (5'UTR, NS5B and HVR-1) revealed the circulation of 1a (73.9%), 1b (8.7%) and 3a (17.4%) genotypes. Next-generation sequencing and phylogenetic analysis of intra-host viral populations of HCV HVR-1 showed a significant variation in intra-host genetic diversity among infected individuals, with 58.8% composed of more than one sub-population. Bayesian analysis estimated that the most recent common HCV ancestor for strains identified here was introduced to this region after 1975 following expansion of intravenous drug use in Brazil. Multivariate analyses showed that only 'ever having injected drugs' was independently associated with HCV infection. These results indicate an increasing spread of multiple HCV strains requiring public health intervention, such as harm reduction, testing services and treatment among crack users in this important border region of Central Brazil.