Betty H. Robertson

Genetic Diversity of Hepatitis B Virus Strains Derived Worldwide: Genotypes, Subgenotypes, and HBsAg Subtypes

Sequences of 234 complete genomes and 631 hepatitis B surface antigen genes were used to assess the worldwide diversity of hepatitis B virus (HBV). Apart from the described two subgenotypes each for A and F, also B, C, and D divided into four subgenotypes each in the analysis of complete genomes supported by significant bootstrap values. The subgenotypes of B and C differed in their geographical distribution, with B1 dominating in Japan, B2 in China and Vietnam, B3 confined to Indonesia, and B4 confined to Vietnam, all strains specifying subtype ayw1. Subgenotype C1 was common in Japan, Korea, and China; C2 in China, South-East Asia, and Bangladesh, and C3 in the Oceania comprising strains specifying adrq–, and C4 specifying ayw3 is encountered in Aborigines from Australia. This pattern of defined geographical distribution was less evident for D1–D4, where the subgenotypes were widely spread in Europe, Africa, and Asia, possibly due to their divergence having occurred a longer time ago than for genotypes B and C, with D4 being the first split and still the dominating subgenotype of D in the Oceania. The genetic diversity of HBV and the geographical distribution of its subgenotypes provide a tool to reconstruct the evolutionary history of HBV and may help to complement genetic data in the understanding of the evolution and past migrations of man.

Genotype H: a new Amerindian genotype of hepatitis B virus revealed in Central America

The complete genomes were sequenced for ten hepatitis B virus (HBV) strains. Two of them, from Spain and Sweden, were most similar to genotype D, although encoding d specificity. Five of them were from Central America and belonged to genotype F. Two strains from Nicaragua and one from Los Angeles, USA, showed divergences of 3.1-4.1% within the small S gene from genotype F strains and were recognized previously as a divergent clade within genotype F. The complete genomes of the two genotype D strains were found to differ from published genotype D strains by 2.8-4.6%. Their S genes encoded Lys(122), Thr(127) and Lys(160), corresponding to the putative new subtype adw3 within this genotype, previously known to specify ayw2, ayw3 or, rarely, ayw4. The complete genomes of the three divergent strains diverged by 0.8-2.5% from each other, 7.2-10.2% from genotype F strains and 13.2-15.7% from other HBV strains. Since pairwise comparisons of 82 complete HBV genomes of intratypic and intertypic divergences ranged from 0.1 to 7.4% and 6.8 to 17.1%, respectively, the three sequenced strains should represent a new HBV genotype, for which the designation H is proposed. In the polymerase region, the three strains had 16 unique conserved amino acid residues not present in genotype F strains. So far, genotype H has been encountered in Nicaragua, Mexico and California. Phylogenetic analysis of the complete genomes and subgenomes of the three strains showed them clustering with genotype F but forming a separate branch supported by 100% bootstrap. Being most similar to genotype F, known to be an Amerindian genotype, genotype H has most likely split off from genotype F within the New World.

Classification, nomenclature, and database development for hepatitis C virus (HCV) and related viruses: proposals for standardization

SummaryThis paper presents a summary of the recommendations that were formulated for the purposes of unifying the nomenclature for hepatitis C virus (HCV), based upon guidelines of the International Committee on Virus Taxonomy (ICTV), and provides guidelines of the incorporation of sequence data into an HCV database that will be available to researchers through the internet. Based upon the available data, the genus Hepacivirus should be regarded as comprising as comprising a single species with HCV-1 as the prototype. All currently known isolates of HCV can be divided into six phylogenetically distinct groups, and we recommend that these groups are described as clades 1 to 6. Whether or not these should be regarded as different species within the Hepacivirus genus requires additional clinical, virological, and immunological information. Clades 1, 2, 4, and 5 would correspond to genotype 1, 2, 4, and 5 while clade 3 would comprise genotype 3 and genotype 10, and clade 6 comprise genotypes 6, 7, 8, 9, and 11. We propose that existing subtype designations are reassigned within these clades based upon publication priority, the existence of a complete genome sequence and prevalence. The assignment of isolates to new clades and subtypes should be confined to isolates characterized from epidemiologically unlinked individuals. Comparisons should be based on nucleotide sequence of at least two coding regions and preferably of complete genome sequences, and should be based on phylogenetic analysis rather than percent identity. A forum for discussion and contributions to these recommendations will be made available at the international HCV database at http://s2as02.genes.nig.ac.jp.

Genetic relatedness of hepatitis A virus strains recovered from different geographical regions

A pairwise comparison of the nucleic acid sequence of 168 bases from 152 wild-type or unique cell culture-adapted strains of hepatitis A virus (HAV) revealed that HAV strains can be differentiated genetically into seven unique genotypes (I to VII). In general, the nucleotide sequence of viruses in different genotypes differs at 15 to 25% of positions within this segment of the genome. Viruses from four of the genotypes (I, II, III and VII) were recovered from cases of hepatitis A in humans, whereas viruses from the other three genotypes (IV, V and VI) were isolated only from simian species developing a hepatitis A-like illness during captivity. Among non-epidemiologically related human HAV strains, 81 were characterized as genotype I, and 19 as genotype III. Within each of these major genotypes, there were two distinct groups (subgenotypes), which differed in sequence at approximately 7.5% of base positions. Each genotype and subgenotype has a characteristic amino acid sequence in this region of the polyprotein, with the most divergent genotypes differing at 10 of 56 residues. Strains recovered from some geographical regions belonged to a common (endemic) genotype, whereas strains from other regions belonged to several, probably imported, genotypes. Thus, HAV strains recovered in North America were for the most part closely related at the nucleotide sequence level, whereas in other regions, such as Japan and Western Europe, HAV strains were derived from multiple genotypes or sub-genotypes. These data indicate that patterns of endemic transmission can be differentiated from situations in which infections are imported due to travel.

Hepatitis B virus genotype assignment using restriction fragment length polymorphism patterns.

Hepatitis B virus (HBV) is classified into genotypes A–F, which is important for clinical and etiological investigations. To establish a simple genotyping method, 68 full‐genomic sequences and 106 S gene sequences were analyzed by the molecular evolutionary method. HBV genotyping with the S gene sequence is consistent with genetic analysis using the full‐genomic sequence. After alignment of the S sequences, genotype specific regions are identified and digested by the restriction enzymes, HphI, NciI, AlwI, EarI, and NlaIV. This HBV genotyping system using restriction fragment length polymorphism (RFLP) was confirmed to be correct when the PCR products of the S gene in 23 isolates collected from various countries were digested with this method. A restriction site for EarI in genotype B was absent in spite of its presence in all the other genotypes and genotype C has no restriction site for AlwI. Only genotype E is digested with NciI, while only genotype F has a restriction site for HphI. Genotype A can be distinguished by a single restriction enzyme site for NlaIV, while genotype D digestion with this enzyme results in two products that migrates at 265 and 186 bp. This simple and accurate HBV genotyping system using RFLP is considered to be useful for research on HBV.

Classification, nomenclature, and database development for hepatitis C virus (HCV) and related viruses

SummaryThis paper presents a summary of the recommendations that were formulated for the purposes of unifying the nomenclature for hepatitis C virus (HCV), based upon guidelines of the International Committee on Virus Taxonomy (ICTV), and provides guidelines of the incorporation of sequence data into an HCV database that will be available to researchers through the internet. Based upon the available data, the genus Hepacivirus should be regarded as comprising as comprising a single species with HCV-1 as the prototype. All currently known isolates of HCV can be divided into six phylogenetically distinct groups, and we recommend that these groups are described as clades 1 to 6. Whether or not these should be regarded as different species within the Hepacivirus genus requires additional clinical, virological, and immunological information. Clades 1, 2, 4, and 5 would correspond to genotype 1, 2, 4, and 5 while clade 3 would comprise genotype 3 and genotype 10, and clade 6 comprise genotypes 6, 7, 8, 9, and 11. We propose that existing subtype designations are reassigned within these clades based upon publication priority, the existence of a complete genome sequence and prevalence. The assignment of isolates to new clades and subtypes should be confined to isolates characterized from epidemiologically unlinked individuals. Comparisons should be based on nucleotide sequence of at least two coding regions and preferably of complete genome sequences, and should be based on phylogenetic analysis rather than percent identity. A forum for discussion and contributions to these recommendations will be made available at the international HCV database at http://s2as02.genes.nig.ac.jp.

Hepatitis E Virus Antibody Prevalence among Persons Who Work with Swine

Prevalence of antibody and risk factors to hepatitis E virus (HEV) infection were determined in a cross-sectional study of 2 group-matched populations: swine farmers (n=264) and persons without occupational exposure to swine (n=255) in Moldova, a country without reported cases of hepatitis E. The prevalence of HEV infection was higher among swine farmers than among the comparison group (51.1% vs. 24.7%; prevalence ratio, 2.07; 95% confidence interval [CI], 1.62-2.64). In multivariate analysis, HEV infection was associated with an occupational history of cleaning barns or assisting sows at birth (odds ratio [OR], 2.46; 95% CI, 1.52-4.01), years of occupational exposure (OR, 1.04 per year; 95% CI, 1.01-1.07), and a history of drinking raw milk (OR, 1.61; 95% CI, 1.08-2.40). HEV infection was not associated with civilian travel abroad or having piped water in the household. The increased prevalence of HEV infection among persons with occupational exposure to swine suggests animal-to-human transmission of this infection.

Aetiological Agent of Enterically Transmitted Non-A, Non-B Hepatitis

Virus-like particles (VLPs) with a mean diameter of 32 nm were recovered from the stools of three acute phase cases of enterically transmitted non-A, non-B hepatitis (ET-NANBH) occurring in the Soviet Union, North Africa and North America. VLPs from two of these cases were studied in detail and were shown to react specifically with antibody in acute phase sera obtained from other cases of ET-NANBH in Asia, the Soviet Union, North Africa and North America. Partially purified VLPs were found to sediment at 183S in sucrose gradients and to cross-react with antibody in acute phase sera from geographically isolated cases of ET-NANBH. The latter virus preparations were also used to document the seroconversion of experimentally ET-NANBH-infected cynomolgus macaques to 32 nm VLPs. Our findings indicate that one virus or class of viruses is responsible for the majority of ET-NANBH.

Characterization of hepatitis D virus genotype III among Yucpa Indians in Venezuela.

The complete genome sequences of hepatitis D virus (HDV) strains isolated from three Yucpa Amerindians in Venezuela were determined and found to be genotype III. Comparison of these three genotype III sequences demonstrated the presence of a hypervariable region containing numerous substitutions, insertions/deletions and a highly conserved region containing the self-cleavage domains, which have been reported previously for genotypes I and II. Amino acid changes within the first 90 amino acids of the hepatitis D antigen (HDAg) were found in the genotype III sequences, while the remainder of the HDAg-coding sequence was conserved. The secondary structure for the RNA-editing site differed between genotypes I and III. It was concluded that the serious delta hepatitis outbreaks characterized epidemiologically in the Yucpa Amerindians were caused by HDV genotype III isolates that were related to HDV genotype III isolates from other regions of South America.

Development and Evaluation of a Broadly Reactive TaqMan Assay for Rapid Detection of Hepatitis A Virus

ABSTRACT Primers and a TaqMan probe for the 5′-untranslated region (UTR) of the hepatitis A virus (HAV) genome were designed and evaluated. The assay detected 0.5 infectious units of HAV and 40 copies of a synthetic transcript and provides an important screening tool for rapid quantitative HAV detection in clinical or environmental samples.