Use of Molecular Epidemiology to Inform Response to a Hepatitis A Outbreak - Los Angeles County, California, October 2018-April 2019.

Los Angeles County comprises 4,058 square miles and is home to approximately 10 million residents (1), an estimated 59,000 (0.6%) of whom experience homelessness on a given night (2). In late 2018, Los Angeles County Department of Public Health (LAC DPH) was notified of a case of hepatitis A virus (HAV) infection in a person experiencing homelessness. LAC DPH conducted an investigation to determine the source of infection, identify additional cases, and identify contacts for postexposure prophylaxis (PEP). Over the next week, LAC DPH identified two additional hepatitis A cases in persons experiencing homelessness who knew one another socially and were known to congregate at a specific street intersection. To identify and respond rapidly to additional outbreak-associated cases, LAC DPH implemented enhanced surveillance procedures, including immediately obtaining specimens for molecular testing from all patients with suspected hepatitis A in the same geographic area. Enhanced surveillance identified four additional cases in persons linked to a senior living campus within two blocks of the intersection where the initial three patients reported congregating. These four cases were linked to the cluster in persons experiencing homelessness through HAV genotyping. Overall, DPH identified seven outbreak-associated hepatitis A cases during October 2018-January 2019. The DPH response to this community hepatitis A outbreak included conducting vaccination outreach to persons at risk, conducting environmental health outreach to restaurants in the outbreak area, and issuing health care provider alerts about the increased occurrence of hepatitis A. Implementation of near real-time molecular testing can improve hepatitis A outbreak responses by confirming HAV infections, linking additional cases to the outbreak, and informing the targeting of prevention efforts.

Redundant Late Domain Functions of Tandem VP2 YPX3L Motifs in Nonlytic Cellular Egress of Quasi-enveloped Hepatitis A Virus.

The quasi-envelopment of hepatitis A virus (HAV) capsids in exosome-like virions (eHAV) is an important but incompletely understood aspect of the hepatovirus life cycle. This process is driven by recruitment of newly assembled capsids to endosomal vesicles into which they bud to form multivesicular bodies with intraluminal vesicles that are later released at the plasma membrane as eHAV. The endosomal sorting complexes required for transport (ESCRT) are key to this process, as is the ESCRT-III-associated protein, ALIX, which also contributes to membrane budding of conventional enveloped viruses. YPX1or3L late domains in the structural proteins of these viruses mediate interactions with ALIX, and two such domains exist in the HAV VP2 capsid protein. Mutational studies of these domains are confounded by the fact that the Tyr residues (important for interactions of YPX1or3L peptides with ALIX) are required for efficient capsid assembly. However, single Leu-to-Ala substitutions within either VP2 YPX3L motif (L1-A and L2-A mutants) were well tolerated, albeit associated with significantly reduced eHAV release. In contrast, simultaneous substitutions in both motifs (L1,2-A) eliminated virus release but did not inhibit assembly of infectious intracellular particles. Immunoprecipitation experiments suggested that the loss of eHAV release was associated with a loss of ALIX recruitment. Collectively, these data indicate that HAV YPX3L motifs function as redundant late domains during quasi-envelopment and viral release. Since these motifs present little solvent-accessible area in the crystal structure of the naked extracellular capsid, the capsid structure may be substantially different during quasi-envelopment.IMPORTANCE Nonlytic release of hepatitis A virus (HAV) as exosome-like quasi-enveloped virions is a unique but incompletely understood aspect of the hepatovirus life cycle. Several lines of evidence indicate that the host protein ALIX is essential for this process. Tandem YPX3L "late domains" in the VP2 capsid protein could be sites of interaction with ALIX, but they are not accessible on the surface of an X-ray model of the extracellular capsid, raising doubts about this putative late domain function. Here, we describe YPX3L domain mutants that assemble capsids normally but fail to bind ALIX and be secreted as quasi-enveloped eHAV. Our data support late domain function for the VP2 YPX3L motifs and raise questions about the structure of the HAV capsid prior to and following quasi-envelopment.

Evaluation of candidate genes associated with hepatitis A and E virus infection in Chinese Han population.

BACKGROUND: Recent GWAS-associated studies reported that single nucleotide polymorphisms (SNPs) in ABCB1, TGFß1, XRCC1 genes were associated with hepatitis A virus (HAV) infection, and variants of APOA4 and APOE genes were associated with and hepatitis E virus (HEV) infection in US population. However, the associations of these loci with HAV or HEV infection in Chinese Han population remain unclear. METHODS: A total of 3082 Chinese Han persons were included in this study. Anti-HAV IgG and anti-HEV IgG were detected by enzyme-linked immunosorbent assay (ELISA). Genotypes in ABCB1, TGFß1, XRCC1, APOA4 and APOE SNPs were determined by TaqMan MGB technology. RESULTS: In Chinese Han population, rs1045642 C to T variation in ABCB1 was significantly associated with the decreased risk of HAV infection (P < 0.05). However, the effect direction was different with the previous US study. Rs1001581 A to G variation in XRCC1, which was not identified in US population, was significantly associated with the protection against HAV infection in our samples (P < 0.05). In addition, our results suggested that rs7412 C to T variation in APOE was significantly associated with lower risk of HEV infection in males (adjusted OR < 1.0, P < 0.05) but not in females. CONCLUSIONS: ABCB1 and XRCC1 genes variants are significantly associated with the protection against HAV infection. Additionally, Chinese Han males with rs7412 C to T variation in APOE gene are less prone to be infected by HEV.

Molecular characterization of hepatitis A virus from children hospitalized at a tertiary care centre in northwest India.

Background & objectives: Hepatitis A virus (HAV) infection is a major cause of childhood hepatitis, prevalent worldwide. HAV is classified into seven genotypes I-VII; genotypes III and I are the most common among humans. The present work was carried out to identify the genotypes prevalent in children suspected to have acute viral hepatitis (AVH), hospitalized at a tertiary care centre in northwest India. Methods: A total of 1269 blood samples from children (0-15 yr of age) clinically suspected of viral hepatitis were screened for anti-HAV IgM. Acute phase serum was processed for RNA extraction and amplified by nested polymerase chain reaction (PCR) followed by sequencing of representative samples. Results: Among the 1269 samples tested, 642 (50.59%) were positive for anti-HAV IgM; among the positive samples, 171 patients having a history of less than seven days were tested by PCR, of whom 141 (82.45%) were found to be PCR positive. Nucleotide sequencing of a representative 44 samples showed high homology; all the samples were found to be of genotype IIIA. Interpretation & conclusions: Hepatitis A was prevalent during July to September and in predominantly children less than five years age. Only genotype IIIA was detected in all the samples.

Relationship of Severity of Hepatitis A with Polymorphisms in Hepatitis A Virus Cellular Receptor 1 (HAVCR1) Gene.

INTRODUCTION AND AIM: HAVCR1 protein is the cellular receptor for hepatitis A virus (HAV). Genetic polymorphism in this gene may alter the outcome of HAV infection. In a previous study, a 6-amino acid insertion (157insMTTTVP) in HAVCR1 gene was associated with more severe disease. We decided to investigate this association further. MATERIAL AND METHODS: We sequenced exon 4 of the HAVCR1 gene in patients with clinical hepatitis A attending our institution, and a group of healthy controls in a disease-endemic setting in India. Frequencies of different haplotypes of a genomic region with two overlapping insertion-deletion polymorphisms (indels; rs141023871 and rs139041445) were compared between patients and controls, as well as between patients with and without a severe form of disease (liver failure). RESULTS: The gene had three haplotypes in the region of interest - a short form, an intermediate-form with a 5-amino acid 157insMTTVP insertion and a long-form with a 6-amino acid 157insMTTTVP insertion. The allele frequency (29/150 [19%] vs. 43/146 [29%]; p = ns) and haplotype frequency (29/75 [39%] vs. 39/73 [53%]; p = ns) of the 157insMTTTVP variant were similar in hepatitis A patients and healthy controls (30%). Further, the allele frequency (12/58 [21%] vs. 17/92 [18%]; p = ns) and haplotype frequency (12/29 [41%] vs.17/46 [37%]; p = ns) of the longest variant were also similar in patients with severe and mild disease. DISCUSSION: In the study population, the 157insMTTTVP variant of HAVCR1 gene was not associated with more severe outcome of HAV infection. Further studies in other populations around the world are needed to assess the relation of this genetic variation with disease outcome.

Micro RNAs mir-106a, mir-122 and mir-197 are increased in severe acute viral hepatitis with coagulopathy.

BACKGROUND & AIMS: The severity of acute viral hepatitis, which may be caused by several distinct viruses, varies among individual patients. In rare cases, severe hepatic injury with sudden loss of liver function may occur, which is clinically indicated by the occurrence of coagulopathy or encephalopathy. As the molecular mechanisms of this liver injury are largely unknown, we investigated extracellular micro RNA (miRNA) profiles in 54 patients acutely infected with one of four different hepatotropic viruses, in order to identify those miRNAs which indicate severe viral hepatitis associated with coagulopathy. METHODS: First, the profile of miRNAs was extensively analysed using a microarray-based approach in highly characterized 24 patients, matched in terms of sex, age and level of liver enzymes, as well as in three healthy controls. The cohort included samples from 18 patients with moderate and six individuals with severe hepatitis, indicated by abnormal prothrombin time and higher alanine aminotransferase and bilirubin levels. miRNAs found to be upregulated in severe hepatitis were then quantified by real-time PCR in the expanded cohort of 54 patients. RESULTS: Comprehensive microarray-based miRNA profiling identified upregulation of mir-106a, mir-122 and mir-197 in patients with severe acute viral hepatitis with coagulopathy, as compared to patients who did not develop coagulopathy. mir-106a, mir-122 and mir-197 were then proven to be significantly upregulated in patients with severe acute viral hepatitis by quantitative real-time PCR (P < 0.01, Mann-Whitney U-test). CONCLUSIONS: mir-106a, mir-122 and mir-197 could be potential markers for severe acute viral hepatitis associated with coagulopathy.

Molecular characterization of hepatitis A virus strains in a tertiary care health set up in north western India.

BACKGROUND & OBJECTIVES: Hepatitis A virus usually causes acute viral hepatitis (AVH) in the paediatric age group with a recent shift in age distribution and disease manifestations like acute liver failure (ALF). This has been attributed to mutations in 5'non-translated region (5'NTR) which affects the viral multiplication. The present study was aimed to carry out the molecular detection and phylogenetic analysis of hepatitis A virus strains circulating in north western India. METHODS: Serum samples from in patients and those attending out patient department of Pediatric Gastroenterology in a tertiary care hospital in north India during 2007-2011 with clinically suspected AVH were tested for anti-hepatitis A virus (HAV) IgM antibodies. Acute phase serum samples were subjected to nested PCR targeting the 5'NTR region followed by sequencing of the representative strains. RESULTS: A total of 1334 samples were tested, 290 (21.7%) were positive for anti-HAV IgM antibody. Of these, 78 serum samples (< 7 days old) were subjected to PCR and 47.4% (37/78) samples showed the presence of HAV RNA. Children < 15 yr of age accounted for majority (94%) of cases with highest seropositivity during rainy season. Sequencing of 15 representative strains was carried out and the circulating genotype was found to be III A. The nucleotide sequences showed high homology among the strains with a variation ranging from 0.1-1 per cent over the years. An important substitution of G to A at 324 position was shown by both AVH and ALF strains. The cumulative substitution in AVH strains Vs ALF strains as compared to GBM, Indian and prototype strain in the 200-500 region of 5' NTR was comparable. INTERPRETATION & CONCLUSION: Our results showed hepatitis A still a disease of children with III A as a circulating genotype in this region. The mutations at 5'NTR region warrant further analysis as these affect the structure of internal ribosomal entry site which is important for viral replication.

Whole genome sequencing to identify host genetic risk factors for severe outcomes of hepatitis a virus infection.

Acute liver failure is a severe, but rare, outcome of hepatitis A virus infection. Unusual presentations of prevalent infections have often been attributed to pathogen-specific immune deficits that exhibit Mendelian inheritance. Genome-wide resequencing of unrelated cases has proven to be a powerful approach for identifying highly penetrant risk alleles that underlie such syndromes. Rare mutations likely to affect protein expression or function can be identified from sequence data, and their association with a similarly rare phenotype rests on their existence in multiple affected individuals. A rare or novel sequence variant that is enriched to a significant degree in a genetically diverse cohort suggests a candidate susceptibility allele. Whole genome sequencing of ten individuals from ethnically diverse backgrounds with HAV-associated acute liver failure was performed. A set of rational filtering criteria was used to identify genetic variants that are rare in the population, but enriched in this cohort. Single nucleotide polymorphisms, insertions, and deletions were considered and autosomal dominant, autosomal recessive, and polygenic models were applied. Analysis of the protein-coding exome identified no single gene with putatively deleterious mutations shared by multiple individuals, arguing against a simple Mendelian model of inheritance. A number of rare variants were significantly enriched in this cohort, consistent with a complex and genetically heterogeneous trait. Several of the variants identified in this genome-wide study lie within genes important to hepatic pathophysiology and are candidate susceptibility alleles for hepatitis A virus infection.

Acute hepatitis A virus infection is associated with a limited type I interferon response and persistence of intrahepatic viral RNA.

Hepatitis A virus (HAV) is an hepatotropic human picornavirus that is associated only with acute infection. Its pathogenesis is not well understood because there are few studies in animal models using modern methodologies. We characterized HAV infections in three chimpanzees, quantifying viral RNA by quantitative RT-PCR and examining critical aspects of the innate immune response including intrahepatic IFN-stimulated gene expression. We compared these infection profiles with similar studies of chimpanzees infected with hepatitis C virus (HCV), an hepatotropic flavivirus that frequently causes persistent infection. Surprisingly, HAV-infected animals exhibited very limited induction of type I IFN-stimulated genes in the liver compared with chimpanzees with acute resolving HCV infection, despite similar levels of viremia and 100-fold greater quantities of viral RNA in the liver. Minimal IFN-stimulated gene 15 and IFIT1 responses peaked 1-2 wk after HAV challenge and then subsided despite continuing high hepatic viral RNA. An acute inflammatory response at 3-4 wk correlated with the appearance of virus-specific antibodies and apoptosis and proliferation of hepatocytes. Despite this, HAV RNA persisted in the liver for months, remaining present long after clearance from serum and feces and revealing dramatic differences in the kinetics of clearance in the three compartments. Viral RNA was detected in the liver for significantly longer (35 to >48 wk) than HCV RNA in animals with acute resolving HCV infection (10-20 wk). Collectively, these findings indicate that HAV is far stealthier than HCV early in the course of acute resolving infection. HAV infections represent a distinctly different paradigm in virus-host interactions within the liver.

Variants in ABCB1, TGFB1, and XRCC1 genes and susceptibility to viral hepatitis A infection in Mexican Americans.

UNLABELLED: Hepatitis A vaccination has dramatically reduced the incidence of hepatitis A virus (HAV) infection, but new infections continue to occur. To identify human genetic variants conferring a risk for HAV infection among the three major racial/ethnic populations in the United States, we assessed associations between 67 genetic variants (single nucleotide polymorphisms [SNPs]) among 31 candidate genes and serologic evidence of prior HAV infection using a population-based, cross-sectional study of 6,779 participants, including 2,619 non-Hispanic whites, 2,095 non-Hispanic blacks, and 2,065 Mexican Americans enrolled in phase 2 (1991-1994) of the Third National Health and Nutrition Examination Survey. Among the three racial/ethnic groups, the number (weighted frequency) of seropositivity for antibody to HAV was 958 (24.9%), 802 (39.2%), and 1540 (71.5%), respectively. No significant associations with any of the 67 SNPs were observed among non-Hispanic whites or non-Hispanic blacks. In contrast, among Mexican Americans, variants in two genes were found to be associated with an increased risk of HAV infection: TGFB1 rs1800469 (adjusted odds ratio [OR], 1.38; 95% confidence interval [CI], 1.14-1.68; P value adjusted for false discovery rate [FDR-P] = 0.017) and XRCC1 rs1799782 (OR, 1.57; 95% CI, 1.27-1.94; FDR-P = 0.0007). A decreased risk was found with ABCB1 rs1045642 (OR, 0.79; 95% CI, 0.71-0.89; FDR-P = 0.0007). CONCLUSION: Genetic variants in ABCB1, TGFB1, and XRCC1 appear to be associated with susceptibility to HAV infection among Mexican Americans. Replication studies involving larger population samples are warranted.

[The hepatitis A virus: structural and functional organization of the genome, its molecular diagnostic value and cultivation].

The analysis of recently published data on hepatitis A virus (HAV) genome clinical features, molecular diagnostic value and cell culture propagation are reviewed. The growing need in the study of the genetic diversity of HAV isolates and the search of its possible new antigenic variants are underlined. The results of the cultivation of different HAV strains are analyzed for possible application in vaccine and diagnostic kit production.

[Development of a multiplex technique for detecting simultaneously HAV RNA and HEV RNA]. / Mise au point d'une technique multiplex de dépistage génomique HEV-HAV.

The hepatitis E virus (HEV) is more and more frequently incriminated in hepatitis episodes. In non-endemic regions, it is clear now that this infection is autochthonous, and certainly a zoonosis and thus, that this virus must not only be assessed in hepatitis cases among travellers in endemic regions. In parallel, in endemic region, where HEV, like HAV, is mainly water-transmitted, important outbreaks still occur. This article describes the development and validation of a new molecular technique for detecting simultaneously HAV RNA and HEV RNA and its evaluation on clinical specimen.

Genotyping of acute hepatitis a virus isolates from China, 2003-2008.

Hepatitis A virus (HAV) is usually transmitted by an oral-fecal route and is prevalent not only in developing countries but also in developed countries. In the present study, the phylogenetic characterization of the VP1/2A junction region (321 nucleotides) of China HAV isolates was examined. Anti-HAV IgM-positive serum samples were collected from 8 provinces, including 20 cities or counties in China from 2003 to 2008; 337 isolates from 406 HAV patients' serum samples were amplified by RT-PCR, sequenced at the VP1/2A junction region and aligned with the published sequences from GenBank to establish phylogenetic analysis. All China HAV isolates in this study belonged to genotype I, with 98.8% (333/337) of samples clustering in sub-genotype IA and 1.2% (4/337) in sub-genotype IB. In addition, sub-genotype IA isolates clustered into four groups (92.7-100% nucleotide identity), and the samples collected from all China HAV isolates in this investigation showed 87.5-100% nucleotide identity, but the amino acids in this region were more conserved (95.2-100% identity). Few unique amino acid changes could be deduced (VP1-253: Glu → Gly; 2A-34: Pro → Ala; 2A-33: Leu → Phe). Genetically identical or similar HAV strains existed in some investigated areas in China during different years, suggesting that an indigenous strain has been circulating in those regions. This report provides new data on the genetic relatedness and molecular epidemiology of HAV isolates from China as well as the distribution of sub-genotype IA and IB in this part of the world.

[Role of hepatitis A and E viruses in the development of autoimmune diseases].

The mechanisms of development of autoimmune diseases may be associated with a complex of genetic, immune, hormonal, and infectious factors. Autoimmune diseases include a wide range of systemic and organ-specific diseases, including autoimmune hepatitis (AIH). It is currently assumed that the pathogenesis of AIH is due to compromised immune regulation in the presence of an exogenous triggering factor. Exogenous factors, such as viruses, may be triggers of AIH. There may be different ways of initiating an autoimmune response by viruses, which includes nonspecific T-lymphocyte activation and molecular mimicry. There is much evidence supporting the initiating role of hepatitis viruses in the development of AIH and other autoimmune diseases. The development of AIH symptoms during hepatitis A and E virus infections has been described elsewhere. The creation of animal models of viral hepatitis is required to confirm the hypothesis that the viruses trigger the development of AIH and other autoimmune manifestations.

IL-15 enhances CCR5-mediated migration of memory CD8+ T cells by upregulating CCR5 expression in the absence of TCR stimulation.

During microbial infection, bystander CD8+ T cells that are not specific to infecting pathogens can be activated by interleukin (IL)-15. However, the tissue-homing properties of bystander-activated CD8+ T cells have not been elucidated. Here, we examine the effects of IL-15 on the expression of chemokine receptors on CD8+ T cells and their migration. IL-15 upregulates CCR5 in memory CD8+ T cells in the absence of T cell receptor (TCR) stimulation and enhances CCR5-dependent migration. IL-15-induced CCR5 upregulation is abrogated by TCR stimulation, indicating that CCR5 is upregulated in bystander-activated CD8+ T cells. Moreover, CCR5 signals increase proliferation and cytotoxic protein expression in IL-15-treated memory CD8+ T cells, although the increase has a small extent. CCR5 upregulation in bystander-activated CD8+ T cells is associated with severe liver injury in patients with acute hepatitis A. Altogether, the results indicate that CCR5 upregulation by IL-15 mediates the migration of bystander-activated CD8+ T cells.

Inherited IL-18BP deficiency in human fulminant viral hepatitis.

Fulminant viral hepatitis (FVH) is a devastating and unexplained condition that strikes otherwise healthy individuals during primary infection with common liver-tropic viruses. We report a child who died of FVH upon infection with hepatitis A virus (HAV) at age 11 yr and who was homozygous for a private 40-nucleotide deletion in IL18BP, which encodes the IL-18 binding protein (IL-18BP). This mutation is loss-of-function, unlike the variants found in a homozygous state in public databases. We show that human IL-18 and IL-18BP are both secreted mostly by hepatocytes and macrophages in the liver. Moreover, in the absence of IL-18BP, excessive NK cell activation by IL-18 results in uncontrolled killing of human hepatocytes in vitro. Inherited human IL-18BP deficiency thus underlies fulminant HAV hepatitis by unleashing IL-18. These findings provide proof-of-principle that FVH can be caused by single-gene inborn errors that selectively disrupt liver-specific immunity. They also show that human IL-18 is toxic to the liver and that IL-18BP is its antidote.

Molecular characterization of hepatitis A virus in children with fulminant hepatic failure in Argentina.

BACKGROUND: Hepatitis A infection, a vaccine-preventable disease, is an important cause of fulminant hepatic failure (FHF) in children in Argentina. Universal vaccination in 1-year-old children was implemented in June 2005. The limited studies about the correlation between the characteristics of the hepatitis A virus (HAV) and FHF have been carried out in adults. METHODS: Samples from 41 children with FHF were studied from September 2003 to January 2006 and HAV RNA was detected, sequenced and analysed in the 5' non-coding region and VP1/2A region. RESULTS: Eighteen HAV strains were characterized and found to be different at the nucleotide level from the self-limited acute infection strains that have been circulating in Argentina with no temporal or geographical pattern. They did not form a genetic cluster, but some of them were identical in the largest fragment characterized and some of them seemed to be more closely related in time and/or geographically. CONCLUSION: Our results suggest that viral factors could be involved in the severity of the clinical presentation of HAV infection in children in Argentina.

Detection of hepatitis A virus RNA from children patients with acute and fulminant hepatitis of unknown etiology in Vietnam: Genomic characterization of Vietnamese HAV strain.

BACKGROUND: Although it is thought that Vietnam is a high endemic region of hepatitis A virus (HAV) infection, there is no report on genomic characterization of HAV spread in Vietnam. The purpose of the present paper was therefore to identify various virus infections from 33 children with acute or fulminant hepatitis of unknown etiology admitted to Children's Hospital No.1 in Ho Chi Minh City, Vietnam. METHODS: Anti-HAV IgM and IgG were assayed by ELISA. Viral RNA and DNA were determined by PCR method. HAV genes isolated by PCR were sequenced and characterized by phylogenetic analysis. RESULTS: Anti-HAV IgM was detected in 18 of 26 acute hepatitis (69.2%) and one of seven (14.3%) fulminant hepatitis patients. Furthermore, HAV-RNA in serum was identified in five of 26 acute (19.2%) and two of seven (28.6%) fulminant hepatitis patients, respectively, on nested reverse transcription-polymerase chain reaction. Among the seven HAV-RNA-positive patients tested, two (28.6%) were negative for anti-HAV IgM. We also obtained seven isolates containing the HAV genome with the viral protein 1 (VP1) region sequence. All Vietnamese HAV isolates formed a cluster and belonged to genotype IA according to phylogenetic analysis based on the short sequences of VP1-2A junction region. CONCLUSION: HAV is an important agent with regard to fulminant hepatitis among children in Vietnam. To the authors' knowledge this is the first report on Vietnamese HAV strain confirmed on sequencing.

Analysis of hepatitis A virus protein 2B in sera of hepatitis A of various severities.

BACKGROUND: In our recent study of the full-length hepatitis A virus (HAV) genome from some patients with fulminant hepatitis and acute hepatitis, possible associations were suggested between the severity of hepatitis A and the amino acid substitutions in the nonstructural protein 2B. We therefore analyzed HAV 2B from many patients with various clinical disease severities. METHODS: Serum samples from 30 Japanese patients with sporadic hepatitis A from five widely separated regions of Japan, comprising nine patients with fulminant hepatitis (FH), six with severe acute hepatitis (AHs), and 15 with acute hepatitis (AH), were examined for HAV RNA. The entire sequences of HAV 2B were analyzed. RESULTS: Compared with the sequence of the wild-type HAV strain GBM, nucleotide sequences of 2B had homology of 94.5 +/- 1.0% in FH, 95.2 +/- 1.2% in AHs, and 95.1 +/- 1.8% in AH. Deduced amino acid sequences had homology of 97.5 +/- 2.1% in FH, 97.9 +/- 2.4% in AHs, and 98.5 +/- 1.3% in AH. Differences were not statistically significant among the three groups. The average number of amino acid mutations between amino acids 100 and 200 was 5.0 +/- 5.2 per case in FH, 4.0 +/- 6.0 in AHs, and 1.9 +/- 2.9 in AH. The differences between FH and AH, AHs and AH, and between severe cases (FH and AHs) and nonsevere cases (AH) were not statistically significant (P = 0.13, P = 0.45, and P = 0.10, respectively). CONCLUSIONS: There were no obvious differences in the sequences among FH, AHs, and AH throughout the 2B region, but there seemed to be more mutations in the strains obtained from FH and AHs patients than in those obtained from AH patients in the central part of HAV 2B.