Complete genome sequences and phylogenetic analysis of Hepatitis B virus isolates from Serbia

Although the genotype and subgenotype distribution of HBV isolates from Serbia has previously been reported, data about whole genome sequences from this area are scarce. This study included plasma samples from 5 chronically infected patients. Full genome amplification of the HBV isolates was performed by nested-PCR using 7 primer pairs, and the whole genome nucleotide sequences were obtained by direct sequencing. Two complete genome sequences belonged to D2 subgenotype (ayw3 HBsAg subtype), one to D1 (ayw2) and two to A2 (adw2). All 5 Serbian isolates clustered with sequences from the expected geographic regions and had nucleotide and coded protein length in accordance to their assigned genotypes, except for one HBeAg-negative isolate displaying G1896A mutation leading to a premature stop codon in the Pre-C region. The first complete genome sequences of HBV D1, D2 and A2 subgenotypes from Serbian patients showed characteristics similar to the nucleotide sequences of HBV isolates from other European and Middle East countries.

Antigenic Diversity of Hepatitis B Virus Strains of Genotype F in Amerindians and Other Population Groups from Venezuela

The adw4 subtype of hepatitis B virus (HBV) belongs to a unique genomic group (genotype F) representing the original HBV strains from the New World. Data regarding the prevalence of this subtype among HBV carriers in South America are, however, scarce, and those concerning HBV genotype F are based on only a few samples from Latin America. In this study, serum samples were obtained from 141 hepatitis B surface antigen (HBsAg) carriers from Amerindians and urban populations from Venezuela. The HBsAg subtype was identified with monoclonal antibodies in 105 samples, and the HBV genotype was identified by reverse-phase hybridization with DNA fragments in 58 samples. The adw4 subtype was highly prevalent in the population studied (75%); among the Amerindians, the prevalence was 97%. The adw2 subtype was also present (10%), while other subtypes (ayw3 and ayw4) were only occasionally found. The HBV subtype was associated with the expected genotype in most cases (80%), and thus genotype F was highly prevalent. Sequencing of viral strains that gave genotypes unpredicted by the HBsAg subtyping confirmed seven of them as belonging to not previously described genotype-subtype associations: namely, adw2 and ayw4 within genotype F.

Inactivation Of Hepatitis B Virus By Heat In Antithrombin III Stabilized With Citrate

The recent report that antithrombin III (AT-III) prepared in solution with 0.5 M sodium citrate can withstand heating at 60°C for 10 hours suggested that this method of preparation could permit the heat-inactivation of hepatitis B virus (HBV) which might be present. Although heating at 60°C for 10 hours will inactivate HBV in albumin, this will not inactivate HBV in whole plasma or in some purified hepatitis B surface antigen (HBsAg) preparations. HBV, subtype ayw, hepatitis B e antigen positive, containing 103.5 chimpanzee infectious doses per ml, was added to each of two vials containing 690 plasma units each of AT- III. One was kept at 4°C for 10 hours (unheated AT-III), the second was heated at 60°C for 10 hours. The contents of both vials were separately dialyzed and inoculated intravenously into chimpanzees with no prior exposure to hepatitis B virus and with no serologic evidence of prior hepatitis B infection. No evidence of hepatitis B was detected in the chimpanzee inoculated with the heated AT-III during six months of evaluation. Aspartate and alanine aminotransferase levels (AST, ALT) remained within or near the normal range. Neither HBsAg, antibody to hepatitis B core antigen (anti-HBc), nor antibody to HBsAg (anti-HBs) was detected in any of the weekly serum samples. The chimpanzee inoculated with the unheated AT-III developed hepatitis B. AST and ALT became elevated 13 weeks after inoculation; HBsAg (subtype ayw) was detected six weeks after inoculation and remained detectable; anti-HBc became detectable 12 weeks after inoculation. Liver biopsies obtained 20 and 23 weeks after inoculation showed histologic changes of acute hepatitis. This study shows that the application of heat at 60°C for 10 hours to AT-III stabilized with 0.5 M sodium citrate can inactivate > 1,000 chimpanzee infectious doses of HBV.