Nationwide epidemiologic and genetic surveillance of hepatitis E in Japan, 2014-2021.

Hepatitis E virus (HEV) is an emerging causative agent of acute hepatitis. To clarify the epidemiology of HEV and characterize the genetic diversity of the virus in Japan, nationwide enhanced surveillance and molecular characterization studies of HEV in Japan were undertaken from 2014 to 2021. In total, 2770 hepatitis E cases were reported, of which 88% were domestic cases, while only 4.1% represented cases following infection abroad. In addition, 57% of domestic infections occurred in males aged in their 40s-70s. For domestic cases, infection via pork meat consumption continued to be the most reported route. Analysis of the 324 sequences detected between 2016 and 2021 showed that the majority of domestic HEV strains belong to Genotype 3a (G3a) and G3b. In contrast, six of eight cases of G1 HEV reflected infection abroad. Our results suggest that HEV is circulating widely in Japan, with genotypes G3a and G3b being most prevalent. Continued surveillance is necessary to monitor future trends and changes in the epidemiology of HEV in Japan.

Delaying the third dose of Japanese aluminum-free hepatitis A vaccine Aimmugen elicits effective immune responses against hepatitis A in adults.

Inactivated aluminum-adsorbed hepatitis A vaccines such as Havrix, Vaqta, and Avaxim are commonly used worldwide. These vaccines are typically administered in a two-dose series (at 0 and 6-12 months). However, a lyophilized inactivated aluminum-free hepatitis A vaccine, Aimmugen, which is approved in Japan, is typically administered in a three-dose series (at 0, 2-4, and 24 weeks). Hence, individuals visiting endemic hepatitis A areas receive the primary two doses of Aimmugen before traveling and the third booster dose much later. It is currently uncertain whether boosting with a delayed third dose of Aimmugen is effective, or whether a new vaccination schedule should instead be initiated. Therefore, we investigated the anti-hepatitis-A viral immune response of adult travelers who received the third dose of Aimmugen more than 24 weeks after the first dose. Participants were vaccinated with the third dose of Aimmugen more than 2 years after the first two doses. Antibody titers were measured at Day 0 (prevaccination) and at 28-42 days after the third dose of Aimmugen. Twenty-nine adult participants were enrolled in the study (14 men and 15 women; mean age ± standard deviation age, 36.2 ± 8.1 years). The interval between the first two doses and the third dose was 3-14 years. The seroprotection rate (i.e., the percentage of participants with anti-hepatitis A virus antibody titers ≥ 10 mIU/mL) was 96.6 % (28/29) at Day 0 and increased to 100 % (29/29) at Days 28-42. Geometric mean concentration increased from 105 to 4,013 mIU/mL. We demonstrated that delaying the third dose of Aimmugen still elicited effective immune responses after priming with two doses of the vaccine. Trial registration: UMIN Clinical Trials Registry (UMIN-CTR): MIN000013624. Registered 03 April 2014. https://center6.umin.ac.jp/cgi-bin/ctr/ctr_view_reg.cgi?recptno=R000015906.

Seroepidemiology of hepatitis A virus infection in Japan: An area of very low endemicity.

The incidence of hepatitis A virus (HAV) infection has declined significantly worldwide, including in Japan. A nationwide seroepidemiological study on hepatitis A in Japan has taken place almost every 10 years since 1973, and the last study was performed in 2003. In the present study, we describe the latest seroepidemiological pattern of hepatitis A in Japan using 7867 serum specimens obtained from healthy individuals collected between 2013 and 2017, approximately 10 years after the last study. Among them, 223 were anti-HAV positive. About 68% of individuals aged 60 years and older had anti-HAV antibodies, whereas only 1.1% of those aged below 60 years old had immunity; thus, almost all individuals younger than 60 years of age were HAV susceptible. In comparison with previous investigations, the susceptible population has increased and aged. According to data from the National Epidemiological Surveillance of Infectious Diseases (NESID) program, between 1989 and 2016, the proportion of patients with hepatitis A aged 60 years and older continuously increased with each year. The NESID data also suggested that recently, typical large foodborne outbreaks of hepatitis A have become rare, and cases tend to be reported among at-risk groups; overseas travelers contributed to 25% of hepatitis A cases, and in 2018, the first nationwide hepatitis A outbreak that affected mostly men who have sex with men was reported. The purpose of this study was to determine the current status of HAV infection in Japan, based on both seroepidemiology and the national surveillance data from the NESID.

Neutralization of hepatitis B virus with vaccine-escape mutations by hepatitis B vaccine with large-HBs antigen.

Although the current hepatitis B (HB) vaccine comprising small-HBs antigen (Ag) is potent and safe, attenuated prophylaxis against hepatitis B virus (HBV) with vaccine-escape mutations (VEMs) has been reported. We investigate an HB vaccine consisting of large-HBsAg that overcomes the shortcomings of the current HB vaccine. Yeast-derived large-HBsAg is immunized into rhesus macaques, and the neutralizing activities of the induced antibodies are compared with those of the current HB vaccine. Although the antibodies induced by the current HB vaccine cannot prevent HBV infection with VEMs, the large-HBsAg vaccine-induced antibodies neutralize those infections. The HBV genotypes that exhibited attenuated neutralization via these vaccines are different. Here, we show that the HB vaccine consisting of large-HBsAg is useful to compensate for the shortcomings of the current HB vaccine. The combined use of these HB vaccines may induce antibodies that can neutralize HBV strains with VEMs or multiple HBV genotypes.

Comparison of the Clinical Features of Hepatitis A in People Living with HIV between Pandemics in 1999-2000 and 2017-2018 in the Metropolitan Area of Japan.

Since 2017, hepatitis A virus (HAV) infection has been an epidemic among men who have sex with men (MSM) in Japan. We have come across 11 MSM patients with hepatitis A who were also infected with HIV. In 1999-2000, we came across 5 HIV-infected patients with hepatitis A. Since the conditions of current HIV-infected patients have changed owing to the recent progress in anti-HIV therapies, we compared clinical features of hepatitis A between patients in 2017-2018 and those in 1999-2000. By comparing the background characteristics of the patients, we found that the CD4/CD8 ratio was significantly higher in the 2017-2018 group. After the onset of hepatitis, peak levels of hepatic transaminases were found to be higher in the 2017-2018 group, suggesting severe hepatocellular damage. In contrast, neither the peak level of total bilirubin nor the nadir of prothrombin time was significantly different among the 2 groups. We also analyzed the HAV genome derived from some of the recently infected patients, and found that the HAV strains were almost the same among these patients; slight differences were observed from the previously identified strain. Thus, we concluded that the recovery of immunity by recent anti-HIV therapies may result in more severe hepatocellular damages and differences in clinical features.

Serologic testing of randomly selected children after hepatitis B vaccination: a cross-sectional population-based study in Lao People's Democratic Republic.

BACKGROUND: Population immunity against hepatitis B virus (HBV) in Lao People's Demographic Republic (PDR) has not been examined since the national HBV vaccination program was started in 2002. Vaccine has been observed to be frozen at times during cold-chain transport in vaccination programs in Lao PDR and other developing countries, which will inactivate the vaccine. Therefore, this study used post-vaccination serologic testing to evaluate the effects of HBV immunization in Lao PDR. METHODS: A cross-sectional serologic study was conducted among children (age range, 5-9 years) and mothers (15-45 years) who were randomly selected using probability-proportional-to-size sampling from central Lao PDR. Blood samples were collected as dried blood spots (DBS) and analyzed using chemiluminescent microparticle immunoassay to detect anti-hepatitis B surface (HBs) titers. We also evaluated the correlation between anti-HBs levels measured in DBS and serum among healthy healthcare workers in Vientiane. RESULTS: Anti-HBs titers from DBS were strongly correlated with serum levels (correlation coefficient = 0.999) in all 12 healthcare workers evaluated. A linear regression model showed that 10 mIU/mL of serum anti-HBs was equivalent to 3.45 mIU/mL (95% CI: 3.06-3.85) of DBS. Among 911 mother-child pairs tested, 171 children had documentation of vaccination. Of the 147 children who had received ≥3 doses of the hepatitis B vaccine, 1 (0.7%) was positive for anti-HBs. The remaining 24 children received the hepatitis B vaccine only twice, once or no dose. CONCLUSIONS: The results showed extremely low positivity for anti-HBs among vaccinated children in central Lao PDR. Therefore, post-vaccination serologic testing is important to evaluate population immunity against HBV infection. DBS testing is a potential low-cost tool to evaluating the effectiveness of HBV vaccination programs.

Immunogenicity of aluminum-adsorbed hepatitis A vaccine (Havrix®) administered as a third dose after primary doses of Japanese aluminum-free hepatitis A vaccine (Aimmugen®) for Japanese travelers to endemic countries.

BACKGROUND: Hepatitis A vaccination is recommended for travelers to endemic countries. Several inactivated aluminum-adsorbed hepatitis A vaccines are available worldwide, but only one licensed hepatitis A vaccine is available in Japan. This vaccine is a lyophilized inactivated aluminum-free hepatitis A vaccine (Aimmugen®). The standard schedule of Aimmugen® is three doses (at 0, 2-4 weeks, and 6 months). Japanese people will go abroad after receiving 2 doses of Aimmugen®. Some long-term travelers will receive the third dose of hepatitis A vaccine at their destination, at 6-24 months after 2 doses of Aimmugen®. Aimmugen® is not available in countries other than Japan. They receive inactivated aluminum-adsorbed hepatitis A vaccine instead of a third dose of Aimmugen®. This study was undertaken to determine whether the booster vaccination with an aluminum-adsorbed hepatitis A vaccine is effective following two doses of Aimmugen®. METHODS: Subjects were healthy Japanese adults aged 20 years or older who had received two doses of Aimmugen®. Subjects received a booster dose of Havrix®1440 intramuscularly as the third dose. Serology samples for hepatitis A virus antibody titers were taken 4-6 weeks later. Anti-hepatitis A virus antibody titers were measured by an inhibition enzyme-linked immunosorbent assay. RESULTS: Subjects were 20 healthy Japanese adults, 6 men and 14 women. The mean age ± standard deviation was 37.2 ± 13.3. The seroprotection rate (SPR, anti-hepatitis A virus antibody titer ≥10 mIU/mL) was 85% at enrollment, and increased to 100% after vaccination with Havrix®. The geometric mean anti-hepatitis A virus antibody titer increased from 39.8 mIU/mL to 2938.2 mIU/mL. CONCLUSION: The three scheduled doses consisting of two doses of Aimmugen® plus a third dose with Havrix® is more immunogenic than using only two doses of Aimmugen®. The vaccination with Havrix® could be allowed to be used instead of a third dose of Aimmugen®. (UMIN000009351).

Clinical Features and Transmission Pattern of Hepatitis A: An Experience from a Hepatitis A Outbreak Caused by Two Cocirculating Genotypes in Sri Lanka.

Sri Lanka is one of the intermediate-endemic areas for hepatitis A virus (HAV), and concerns exist about the increasing HAV-susceptible population. In fact, Sri Lanka recorded a large hepatitis outbreak, possibly hepatitis A, around the end of the Sri Lankan war. It included more than 14,000 patients consisting of local residents, internally displaced personnel, and military personnel in the main combat zone. The outbreak had slowed down by October 2009; however, acute viral hepatitis continued to occur sequentially among military personnel. We obtained clinical information and serum samples from 222 patients with acute hepatitis who visited the Military Hospital Anuradhapura between January and September 2010. Samples were subjected to laboratory testing including HAV-immunoglobulin M and genotyping. Most patients (98.2%) were confirmed as having hepatitis A belonging to two subgenotypes: IA and IIIA. We did not observe any differences in clinical or biochemical features among patients with subgenotypes IA and IIIA except for pale stools and upper abdominal discomfort. During the investigation period, we observed a serial outbreak caused by identical HAV strains with an interval in line with that of typical HAV incubation periods. Most patients in the first outbreak were found in the training center, and patients in the second outbreak were found in multiple places where soldiers were assigned after the training center. These findings indicate that a strain of HAV diffused from one place to another along with movement of infected persons among the HAV-susceptible population. HAV vaccination for high-risk groups, such as young soldiers, is necessary.

Seroepidemiological study of hepatitis B virus markers in Japan.

BACKGROUND: In Japan, since 1986, selective vaccination has been implemented as a hepatitis B prevention strategy. The target of vaccination is the infant born to a hepatitis B surface antigen (HBsAg)-positive mother. The current Japanese hepatitis B prevention strategy focuses on reducing the number of HBV carriers but overlooks the risk to susceptible populations. We conducted a nationwide HBV seroepidemiological study to explore the next hepatitis B control strategy. METHODS: We used sera derived from healthy individuals collected nationwide from 2005 through 2011 to investigate the HBsAg seroprevalence among children aged 4-9 years and 10-15 years (3000 samples) and hepatitis B core antibody (HBcAb) seroprevalence among people 10-39 years of age (600 samples). FINDINGS: Among sera from 3000 children, 5 (0.17%) specimens were HBsAg-positive. There was no significant difference in HBsAg prevalence between age groups. Among 600 samples, 15 (2.5%) were HBcAb-positive. Out of 15 samples, 4 were from teenagers. Both HBsAg- and HBcAb-positive sera were found mainly in the Southern area of Japan. CONCLUSION: The prevalence of HBsAg among children was 0.17% in the present study. This is higher than the prevalence reported in previous studies performed in the local area or in blood donors. The prevalence of HBcAb is also higher than we estimated. One of the reasons for this discrepancy from previous studies may be due to the small sample size and the impact of HBV high-endemic areas included in the present nationwide study. Nevertheless, our findings revealed that the opportunities for acquiring HBV infection in the susceptible population were more frequent than we thought, especially in some localities. Hepatitis B vaccination should be introduced into the routine child immunization program for susceptible populations, and the selective vaccination program should be continued for high-risk children.

Epidemiological and genetic analysis of a 2014 outbreak of hepatitis A in Japan.

Hepatitis A virus (HAV) is one of the most common causes of feces-transmitted acute hepatitis worldwide. In Japan, most of HAV infections have been sporadic cases and a relatively low number of cases (approximately 100-150) of acute hepatitis A were reported in 2012 and 2013. However, in 2014, 342 cases were reported as of week 22. In order to characterize the viral agents causing this outbreak, we collected stool or sera (and both for three case) from patients with hepatitis A from many regions throughout Japan and performed genotyping of the VP1/P2A regions of HAV. We then used a multiple-alignment algorithm to compare the nucleotide sequences with those of reference strains. Phylogenetic tree analyses revealed that the 159 HAV isolates were divided into three subgenotypes: IA (137 cases), IB (4 cases), and IIIA (18 cases). The most unique feature of this outbreak was that for most subgenotype IA cases (103 out of 137 IA cases) the sequences analyzed shared 100% homology. Interestingly, the peak week for these IA infections was almost the same nationwide, suggesting that the epidemic of hepatitis A caused by this subgenotype IA strain may have expanded from a single source possibly because of one food-borne or waterborne source that was distributed nationwide at once.

Seroprevalence of chronic hepatitis B, as determined from dried blood spots, among children and their mothers in central Lao People's Democratic Republic: a multistage, stratified cluster sampling survey.

BACKGROUND: There is limited information regarding the prevalence of hepatitis B in Lao PDR, where the hepatitis disease burden is substantial. Thus, reliable seroprevalence data is needed for the disease, based on probability sampling. METHODS: A stratified, multistage, cluster sampling survey of hepatitis B surface antigen (HBsAg) positivity among children aged 5-9 years and their mothers aged 15-45 years was conducted. Participants were selected randomly from the central region of Lao PDR via probability-proportional-to-size sampling. Blood samples were collected onto filter paper and subsequently analyzed using a chemiluminescent microparticle immunoassay. RESULTS: A total of 911 mother-and-child pairs were collected; the seroprevalence of HBsAg was estimated to be 2.1% (95% confidence interval 0.8-3.4%) among children and 4.1% (95% confidence interval 2.6-5.5%) in their mothers after taking into account the sampling design and the weight of each sample. The children's HBsAg positivity was positively associated with maternal infection and being born in a non-health facility, while the maternal infection status was not associated with any background characteristic. CONCLUSIONS: Lao PDR has a relatively lower HBsAg prevalence in the general population compared to surrounding countries. To ensure comparability to other countries and to future data, rapid field tests are recommended for a nationwide prevalence survey.

Suppression of La antigen exerts potential antiviral effects against hepatitis A virus.

BACKGROUND: Despite the development and availability of hepatitis A virus (HAV) vaccine, HAV infection is still a major cause of acute hepatitis that occasionally leads to fatal liver disease. HAV internal ribosomal entry-site (IRES) is one of the attractive targets of antiviral agents against HAV. The aim of the present study is to evaluate the impact of La, one of the cellular proteins, on HAV IRES-mediated translation and HAV replication. METHODS AND FINDINGS: We investigated the therapeutic feasibility of siRNAs specific for cellular cofactors for HAV IRES-mediated translation in cell culture. It was revealed that siRNA against La could inhibit HAV IRES activities as well as HAV subgenomic replication. We also found that the Janus kinase (JAK) inhibitors SD-1029 and AG490, which reduce La expression, could inhibit HAV IRES activities as well as HAV replication. CONCLUSIONS: Inhibition of La by siRNAs and chemical agents could lead to the efficient inhibition of HAV IRES-mediated translation and HAV replication in cell culture models. La might play important roles in HAV replication and is being exploited as one of the therapeutic targets of host-targeting antivirals.

Interleukin-1 and tumor necrosis factor-α trigger restriction of hepatitis B virus infection via a cytidine deaminase activation-induced cytidine deaminase (AID).

Virus infection is restricted by intracellular immune responses in host cells, and this is typically modulated by stimulation of cytokines. The cytokines and host factors that determine the host cell restriction against hepatitis B virus (HBV) infection are not well understood. We screened 36 cytokines and chemokines to determine which were able to reduce the susceptibility of HepaRG cells to HBV infection. Here, we found that pretreatment with IL-1ß and TNFα remarkably reduced the host cell susceptibility to HBV infection. This effect was mediated by activation of the NF-κB signaling pathway. A cytidine deaminase, activation-induced cytidine deaminase (AID), was up-regulated by both IL-1ß and TNFα in a variety of hepatocyte cell lines and primary human hepatocytes. Another deaminase APOBEC3G was not induced by these proinflammatory cytokines. Knockdown of AID expression impaired the anti-HBV effect of IL-1ß, and overexpression of AID antagonized HBV infection, suggesting that AID was one of the responsible factors for the anti-HBV activity of IL-1/TNFα. Although AID induced hypermutation of HBV DNA, this activity was dispensable for the anti-HBV activity. The antiviral effect of IL-1/TNFα was also observed on different HBV genotypes but not on hepatitis C virus. These results demonstrate that proinflammatory cytokines IL-1/TNFα trigger a novel antiviral mechanism involving AID to regulate host cell permissiveness to HBV infection.

Interleukin-29 suppresses hepatitis A and C viral internal ribosomal entry site-mediated translation.

Our aim was to investigate the effects of interferons (IFNs)-λ (interleukin-29 [IL-29], IL-28A, and IL-28B) on hepatitis C virus (HCV) and hepatitis A virus (HAV) internal ribosomal entry site (IRES)-mediated translation. The effects of these IFNs on HCV/HAV translation from HAV/HCV IRES were investigated using bicistronic reporter constructs. We transfected HCV/HAV IRES constructs into these IFN-expressing cell lines. IL-29 showed stronger inhibition of their IRES-mediated translation. Combining IL-29 with IFN-α or amantadine resulted in stronger inhibition of HAV IRES activity. Our findings demonstrated a novel antiviral effect of IFNs-λ against HAV and HCV through the suppression of IRES-mediated translation.

Hepatitis A outbreak associated with a revolving sushi bar in Chiba, Japan: Application of molecular epidemiology.

AIM: The number of hepatitis A cases in Japan as well as in other developed countries has been progressively decreasing during the last several years. There is no universal hepatitis A vaccination program in Japan, and a hepatitis A virus (HAV) epidemic in Japan is not unlikely. In 2011, a hepatitis A outbreak associated with a revolving sushi bar occurred in Chiba, Japan. We aimed to analyze this outbreak. METHODS: Twenty-seven patients associated with this outbreak were admitted to the National Hospital Organization Chiba Medical Center. Molecular epidemiologic investigations were conducted. RESULTS: Twenty-six of the 27 patients had gone to the same revolving sushi bar, and then clinical symptoms appeared. HAV RNA was detected by reverse transcription polymerase chain reaction in 23 of the 27 (85.1%) patients whose sera had tested positive for anti-HAV immunoglobulin M. All isolates from this outbreak were clustered within subgenotype IA, displaying 100% sequence homology with each other in 232 bp from all 23 patients. All isolates belong to the IA-1 sublineage, which is endemic to Japan. CONCLUSION: A revolving sushi bar was associated with a hepatitis A outbreak, and molecular epidemiological investigations proved useful.

Possible widespread presence of hepatitis A virus subgenotype IIIA in Japan: Recent trend of hepatitis A causing acute liver failure.

AIM: Recently, the number of acute hepatitis A cases has decreased in Japan. However, six patients with acute liver failure caused by hepatitis A virus (HAV) have been admitted to Chiba University Hospital, Japan, in the last 18 months, between 2010 and June 2011. The aim of this study is to characterize the recent HAV genotypes from an urban hospital in Japan and to compare the clinical differences. METHODS: Hepatitis A virus RNA was detected by strand-specific reverse transcription. Then, HAV VP1/2A regions were amplified by nested polymerase chain reaction (PCR). Sequences were directly determined and phylogenetic trees were constructed for determining HAV subgenotypes. RESULTS: Analysis of these HAV genomes revealed that 4 and 2 belonged to subgenotypes IA and IIIA, respectively. CONCLUSIONS: Fujiwara et al. reported a frequency of HAV subgenotype IIIA of only 2.1% in Japan. We conclude that HAV subgenotype IIIA might be widespread in our country.

Antigen-loaded dissolving microneedle array as a novel tool for percutaneous vaccination.

Antigen-loaded dissolving microneedle array (dMNA) patches were investigated as novel systems for vaccine delivery into the skin, where immuno-competent dendritic cells are densely distributed. We fabricated micron-scale needles arrayed on patches, using chondroitin sulfate mixed with a model antigen, ovalbumin. Insertion of dMNA effectively delivered substantial amounts of ovalbumin into the skin within 3 min and induced robust antigen-specific antibody responses in the sera of mice. The antibody dose-response relationship showed that the efficiency of dMNA patch immunization was comparable to that of conventional intradermal injections. Thus, Antigen-loaded dMNA patches are a promising antigen-delivery system for percutaneous vaccination.

Epidemiological and genetic analyses of a diffuse outbreak of hepatitis A in Japan, 2010.

BACKGROUND: Hepatitis A virus (HAV) is still one of the most common causative agents of acute hepatitis in Japan. Although a relatively small number of annual acute hepatitis A cases (approximately 100-150, 0.78-1.17 per million) were recently reported, a larger number of cases (346, 2.71 per million) were reported in 2010. OBJECTIVES: To investigate the causes of the 2010 HAV resurgence in Japan by using molecular epidemiological and genetic analyses. STUDY DESIGN: HAV specimens were obtained from 61 cases from 22 different prefectures. These viral specimens were genotyped by PCR amplification and sequencing of the VP1/2A region of HAV genome. RESULTS: Phylogenetic analysis revealed that 61 HAV strains could be divided into three genotypes: IA (44 cases), IB (1 case) and IIIA (16 cases). The IA genotype consisted of two genomic sub-lineages. The sequences of one of the two IA sub-lineages (corresponding to 31 cases) were very similar, 26 of these 31 isolates had 100% identity. The other IA sub-lineage corresponded to strains endemic to Japan. The sequences of Japanese IIIA strains were similar to those of strains that caused a large epidemic in the Republic of Korea from 2007 to 2009. CONCLUSIONS: The resurgence of HAV in 2010 can be attributed to importation of two newly emerged HAV genotypes.

Inhibitory effects on HAV IRES-mediated translation and replication by a combination of amantadine and interferon-alpha.

Hepatitis A virus (HAV) causes acute hepatitis and sometimes leads to fulminant hepatitis. Amantadine is a tricyclic symmetric amine that inhibits the replication of many DNA and RNA viruses. Amantadine was reported to suppress HAV replication, and the efficacy of amantadine was exhibited in its inhibition of the internal ribosomal entry site (IRES) activities of HAV. Interferon (IFN) also has an antiviral effect through the induction of IFN stimulated genes (ISG) and the degradation of viral RNA. To explore the mechanism of the suppression of HAV replication, we examined the effects of the combination of amantadine and IFN-alpha on HAV IRES-mediated translation, HAV replicon replication in human hepatoma cell lines, and HAV KRM003 genotype IIIB strain replication in African green monkey kidney cell GL37. IFN-alpha seems to have no additive effect on HAV IRES-mediated translation inhibition by amantadine. However, suppressions of HAV replicon and HAV replication were stronger with the combination than with amantadine alone. In conclusion, amantadine, in combination of IFN-alpha, might have a beneficial effect in some patients with acute hepatitis A.