BackgroundHepatitis B virus (HBV) genotype F (HBV/F) is considered to be indigenous to the Americas, but its emergence and spread in the continent remain unknown. Previously, only two HBV/F complete genome sequences from Brazil were available, limiting the contribution of Brazilian isolates to the phylogenetic studies of HBV/F. The present study was carried out to assess the proportion and geographic distributions of HBV/F subgenotypes in Brazil, to determine the full-length genomic sequences of HBV/F isolates from different Brazilian geographic regions, and to investigate the detailed evolutionary history and phylogeography of HBV/F in Brazil.MethodsComplete HBV/F genomes isolated from 12 Brazilian patients, representing the HBV/F subgenotypes circulating in Brazil, were sequenced and analyzed together with sequences retrieved from GenBank, using the Bayesian coalescent and phylogeographic framework.ResultsPhylogenetic analysis using all Brazilian HBV/F S-gene sequences available in GenBank showed that HBV/F2a is found at higher frequencies countrywide and corresponds to all sequences isolated in the Brazilian Amazon Basin. In addition, the evolutionary analysis using complete genome sequences estimated an older median ancestral age for the Brazilian HBV/F2a compared to the Brazilian HBV/F1b and HBV/F4 subgenotypes, suggesting that HBV/F2a represents the original native HBV of Brazil. The phylogeographic patterns suggested a north-to-south flow of HBV/F2a from Venezuela to Brazil, whereas HBV/F1b and HBV/F4 strains appeared to have spread from Argentina to Brazil.ConclusionsThis study suggests a plausible route of introduction of HBV/F subgenotypes in Brazil and demonstrates the usefulness of recently developed computational tools for investigating the evolutionary history of HBV.

in both patients, with .99 % of colonies hybridizing to a probe specific for this insertion. Phylogenetic analyses of full-length genomes and precore/core fragments revealed that F4 and F1b were the co-infecting subgenotypes in the Brazilian and Argentinian patients, respectively. Bootscanning analysis provided evidence of recombination in several clones from both patients, with recombination breakpoints located mainly at the precore/core region. These data should encourage further investigations on the clinical implications of HBV/G recombinants in HBV/HIV co-infected patients.

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Most cases of hepatocellular carcinoma (HCC) are due to chronic hepatitis B virus (HBV) or hepatitis C virus (HCV) infection worldwide. The aim of this study was to determine the viral genotypes and frequency of 17 mutations (15 for HBV and 2 for HCV), described previously as able to influence the course of chronic liver disease, in patients with and without HCC. This transversal study included 157 Brazilian patients with chronic hepatitis B (n = 51) and C (n = 106). Of these, 12 and 40 patients had HBV- and HCV-related HCC, respectively. Nucleotide sequencing of core promoter, pre-core, and pre-S/S regions of HBV and core region of HCV strains was performed to determine their genotypes and the frequency of the respective mutations. Among the HBV isolates, subgenotype A1 was the most prevalent in both patients with (90%) and without (61%) HCC. Fourteen out of the 15 mutations under study, as well as five different pre-S deletions, were identified. Core promoter T1753V, A1762T, and G1764A mutations were more frequent in patients with HCC than in those without, although with no statistical difference. However, a significant correlation was observed between T1753V mutation and elevation of transaminases levels (P < 0.05). As for HCV, mutation at residue 70 in the core protein of genotype 1b strains was significantly more frequent in patients with cirrhosis (56.3%) than in those without (9.1%) (P = 0.018). The detection of some key mutations in the genomes of HBV and HCV might be helpful to predict the clinical outcome of patients with chronic liver disease.

Hepatocellular carcinoma (HCC) is the second most common cause of cancer mortality worldwide. Most cases of HCC are associated with cirrhosis related to chronic hepatitis B virus or hepatitis C virus infections. Hypermethylation of promoter regions is the main epigenetic mechanism of gene silencing and has been involved in HCC development. The aim of this study was to determine whether aberrant methylation of RASSF1A and DOK1 gene promoters is associated with the progression of liver disease in Brazilian patients. Methylation levels were measured by pyrosequencing in 41 (20 HCC, 9 cirrhotic, and 12 non-cirrhotic) liver tissue samples. Mean rates of methylation in RASSF1A and DOK1 were 16.2% and 12.0% in non-cirrhotic, 26.1% and 19.6% in cirrhotic, and 59.1% and 56.0% in HCC tissues, respectively, showing a gradual increase according to the progression of the disease, with significantly higher levels in tumor tissues. In addition, hypermethylation of RASSF1A and DOK1 was found in the vast majority (88%) of the HCC cases. Interestingly, DOK1 methylation levels in HCC samples were significantly higher in the group of younger (<40 years) patients, and higher in moderately differentiated than in poorly differentiated tumors (p < 0.05). Our results reinforce the hypothesis that hypermethylation of RASSF1A and DOK1 contributes to hepatocarcinogenesis and is associated to clinicopathological characteristics. RASSF1A and DOK1 promoter hypermethylation may be a valuable biomarker for early diagnosis of HCC and a potential molecular target for epigenetic-based therapy.