Biochemical analysis of the host factor activity of ZCCHC14 in hepatitis A virus replication.

Relatively little is known of the mechanisms underlying hepatitis A virus (HAV) genome replication. Unlike other well-studied picornaviruses, HAV RNA replication requires the zinc finger protein ZCCHC14 and non-canonical TENT4 poly(A) polymerases with which it forms a complex. The ZCCHC14-TENT4 complex binds to a stem-loop located within the internal ribosome entry site (IRES) in the 5' untranslated RNA (5'UTR) and is essential for viral RNA synthesis, but the underlying mechanism is unknown. Here, we describe how different ZCCHC14 domains contribute to its RNA-binding, TENT4-binding, and HAV host factor activities. We show that the RNA-binding activity of ZCCHC14 requires both a sterile alpha motif (SAM) and a downstream unstructured domain (D4) and that ZCCHC14 contains two TENT4-binding sites: one at the N-terminus and the other around D4. Both RNA-binding and TENT4-binding are required for HAV host factor activity of ZCCHC14. We also demonstrate that the location of the ZCCHC14-binding site within the 5'UTR is critical for its function. Our study provides a novel insight into the function of ZCCHC14 and helps elucidate the mechanism of the ZCCHC14-TENT4 complex in HAV replication.IMPORTANCEThe zinc finger protein ZCCHC14 is an essential host factor for both hepatitis A virus (HAV) and hepatitis B virus (HBV). It recruits the non-canonical TENT4 poly(A) polymerases to viral RNAs and most likely also a subset of cellular mRNAs. Little is known about the details of these interactions. We show here the functional domains of ZCCHC14 that are involved in binding to HAV RNA and interactions with TENT4 and describe previously unrecognized peptide sequences that are critical for the HAV host factor activity of ZCCHC14. Our study advances the understanding of the ZCCHC14-TENT4 complex and how it functions in regulating viral and cellular RNAs.

Circulation of a single hepatitis A virus genotype IIIA with two distinct clusters in different states of India.

With the changing hepatitis A epidemiology in India, focal viral outbreaks are being reported from different parts of the country. This study presents Hepatitis A Virus (HAV) strain characterization (period 2009-2020) from 18 states of India. For that, blood and stool samples (n â€‹= â€‹280) were screened for HAV RNA and sequences for 5'non-coding and VP3 regions were generated from positive samples (n â€‹= â€‹68). Presence of a single IIIA genotype in all samples indicated IIIA being the only HAV genotype currently circulating in India. Interestingly, it was evident that these strains form two distinct groups suggesting independent evolution of these two clusters.

The ZCCHC14/TENT4 complex is required for hepatitis A virus RNA synthesis.

Despite excellent vaccines, resurgent outbreaks of hepatitis A have caused thousands of hospitalizations and hundreds of deaths within the United States in recent years. There is no effective antiviral therapy for hepatitis A, and many aspects of the hepatitis A virus (HAV) replication cycle remain to be elucidated. Replication requires the zinc finger protein ZCCHC14 and noncanonical TENT4 poly(A) polymerases with which it associates, but the underlying mechanism is unknown. Here, we show that ZCCHC14 and TENT4A/B are required for viral RNA synthesis following translation of the viral genome in infected cells. Cross-linking immunoprecipitation sequencing (CLIP-seq) experiments revealed that ZCCHC14 binds a small stem-loop in the HAV 5' untranslated RNA possessing a Smaug recognition-like pentaloop to which it recruits TENT4. TENT4 polymerases lengthen and stabilize the 3' poly(A) tails of some cellular and viral mRNAs, but the chemical inhibition of TENT4A/B with the dihydroquinolizinone RG7834 had no impact on the length of the HAV 3' poly(A) tail, stability of HAV RNA, or cap-independent translation of the viral genome. By contrast, RG7834 inhibited the incorporation of 5-ethynyl uridine into nascent HAV RNA, indicating that TENT4A/B function in viral RNA synthesis. Consistent with potent in vitro antiviral activity against HAV (IC50 6.11 nM), orally administered RG7834 completely blocked HAV infection in Ifnar1-/- mice, and sharply reduced serum alanine aminotransferase activities, hepatocyte apoptosis, and intrahepatic inflammatory cell infiltrates in mice with acute hepatitis A. These results reveal requirements for ZCCHC14-TENT4A/B in hepatovirus RNA synthesis, and suggest that TENT4A/B inhibitors may be useful for preventing or treating hepatitis A in humans.

Hepatitis vaccination adherence and completion rates and factors associated with low compliance: A claims-based analysis of U.S. adults.

Poor compliance with multi-dose vaccine schedules by adults for whom hepatitis (Hep) A and B vaccines are recommended contributes to major Hep A and B disease burdens among high-risk U.S. adults. Evidence on hepatitis vaccine series adherence, completion, timeliness of completion, and factors associated with these outcomes, is limited and not readily generalizable for U.S. adults. This retrospective, observational study examined adherence, completion, its timeliness, and the impact of sociodemographic and clinical factors on these outcomes among a large, geographically representative sample of U.S. adults. We analyzed the Optum Clinformatics SES administrative claims database (1/1/2010-6/30/2020) for recipients of 2-dose (HepA, HepB2) or 3-dose (HepB3, HepAB) hepatitis vaccines. Adherence was defined as receipt of booster doses within specified assessment periods, per label-recommended schedules. Completion (receipt of all doses) was assessed at 6, 12, 18, and 24 months.The study included 356,828 adults ≥19 years old who were continuously enrolled in a medical benefit plan for one (HepB2), six (HepB3; HepAB), or 18 months (HepA) prior to and following the index date (first observed vaccine dose). Adherence and 24-month completion rates were: HepA (27.0%, 28.4%), HepB2 (32.2%, 44.8%), HepB3 (14.3%, 37.3%), HepAB, (15.3%, 33.8%). Kaplan-Meier completion curves plateaued after about 6 months for HepB2 and about 12 months for HepA, HepB3, and HepAB vaccines. Logistic regression analyses showed risk for low adherence/completion was generally associated with male gender, younger age, Black or Hispanic race/ethnicity, lower educational or household income attainment, and more comorbidities. Adherence and completion rates for all hepatitis vaccine series are low, especially for males, younger adults, those with lower socio-economic status and more comorbidities. To our knowledge, this is the largest claims-based analysis of adherence and completion rates for U.S. adults initiating all currently available HepA and HepB vaccines. Findings may inform hepatitis vaccination programming.

Increase in Hepatitis A Cases Linked to Imported Strains to Rio de Janeiro, Brazil: A Cross-Sectional Study.

This study aims to evaluate the epidemiological and molecular features associated with HAV transmission in adults in Rio de Janeiro during a period of increased registered cases of HAV (2017-2018). Socio-epidemiological data and serum samples from anti-HAV IgM+ individuals were obtained. HAV RNA was RT-PCR amplified and sequenced for further phylogenetic and phylogeographic analyses. From fifty-two HAV IgM+ individuals, most were men (78.85%; p = 0.024), aged 20-30 years old (84.61%; p < 0.001), resided in the Rio de Janeiro north zone (31/52; 59.62%; p = 0.001), and are men who have sex with men (MSM) (57.69%; p = 0.002). Sexual practices were more frequent (96%) than others risk factors (food-borne (44%), water-borne (42.31%), and parenteral (34.62%)). Individuals who traveled to endemic regions had a 7.19-fold (1.93-36.04; p < 0.01) increased risk of HAV. Phylogenetic analysis revealed four distinct clades of subgenotype IA, three of them comprised sequences from European/Asian MSM outbreaks and one from Brazilian endemic strains. Bayesian Inference showed that the imported strains were introduced to Brazil during large mass sportive events. Sexual orientation and sexual practices may play a role in acquiring HAV infection. Public policies targeting key populations must be implemented to prevent further dissemination of HAV and other STIs.

Development and Evaluation of a Molecular Hepatitis A Virus Assay for Serum and Stool Specimens.

Hepatitis A virus (HAV) is an emerging public health concern and there is an urgent need for ways to rapidly identify cases so that outbreaks can be managed effectively. Conventional testing for HAV relies on anti-HAV IgM seropositivity. However, studies estimate that 10-30% of patients may not be diagnosed by serology. Molecular assays that can directly detect viral nucleic acids have the potential to improve diagnosis, which is key to prevent the spread of infections. In this study, we developed a real-time PCR (RT-PCR) assay to detect HAV RNA for the identification of acute HAV infection. Primers were designed to target the conserved 5'-untranslated region (5'-UTR) of HAV, and the assay was optimized on both the Qiagen Rotor-Gene and the BD MAX. We successfully detected HAV from patient serum and stool samples with moderate differences in sensitivity and specificity depending on the platform used. Our results highlight the clinical utility of using a molecular assay to detect HAV from various specimen types that can be implemented in hospitals to assist with diagnostics, treatment and prevention.

Prolonged Gut Dysbiosis and Fecal Excretion of Hepatitis A Virus in Patients Infected with Human Immunodeficiency Virus.

Hepatitis A virus (HAV) causes transient acute infection, and little is known of viral shedding via the duodenum and into the intestinal environment, including the gut microbiome, from the period of infection until after the recovery of symptoms. Therefore, in this study, we aimed to comprehensively observe the amount of virus excreted into the intestinal tract, the changes in the intestinal microbiome, and the level of inflammation during the healing process. We used blood and stool specimens from patients with human immunodeficiency virus who were infected with HAV during the HAV outbreak in Japan in 2018. Moreover, we observed changes in fecal HAV RNA and quantified the plasma cytokine level and gut microbiome by 16S rRNA analysis from clinical onset to at least 6 months after healing. HAV was detected from clinical onset up to a period of more than 150 days. Immediately after infection, many pro-inflammatory cytokines were elicited, and some cytokines showed different behaviors. The intestinal microbiome changed significantly after infection (dysbiosis), and the dysbiosis continued for a long time after healing. These observations suggest that the immunocompromised state is associated with prolonged viral shedding into the intestinal tract and delayed recovery of the intestinal environment.

Synthesis, delivery, and molecular docking of fused quinolines as inhibitor of Hepatitis A virus 3C proteinase.

It is widely accepted that Hepatitis A virus (HAV) is responsible for liver failure and even death in older people and in people with other serious health issues; so, proposing new compounds with inhibitory activity can help to treated of these disease's. In current study, a new class of quinolines is proposed with inhibitor activity of the HAV proteinase. So, in the first step, fused quinoline derivatives has been synthesized in short reaction time (12.0 min) and high efficiency yields (94%) in presence of 1-carboxymethyl-2,3-dimethylimidazolium iodide ([cmdmim]I) ionic liquid catalyst using a new method. In the following, chemical reactivity and inhibitory activity of synthesized quinolines were evaluated in density functional theory (DFT) framework and molecular docking methodologies. High global softness (0.67 eV), low HOMOSWBNNT-LUMO4a gap (4.78 eV), and more negative adsorption energy (- 87.9 kJ mol-1) in these quinolines reveal that the 4a and 4b compounds have better delivery than other quinolines using SWBNNT as suitable carrier to target cells. Molecular docking shows that the best cavity of the HAV has - 134.2 kJ mol-1 interaction energy involving bonding and non-bonding interactions. In fact, these interactions are between fused quinolines with especial geometries and sidechain flexibility amino acids residues inside the best binding site of the HAV, as hydrogen bonding, steric, and electrostatic interactions. So, these interactions imply that proposed fused quinolines have good inhibitor activity for the HAV.

Evidence of Circulation of Several HAV Genetic Variants and Emergence of Potential Antigenic Variants in an Endemo-Epidemic Country before Vaccine Introduction.

Similar to several other countries in the world, the epidemiology of hepatitis A virus changed from high to intermediate endemicity level in Tunisia, which led to the occurrence of outbreaks. This study aimed to determine the genetic and antigenic variability of HAV strains circulating in Tunisia during the last few years. Genotyping using complete VP1 gene and VP1-2A junction confirmed the predominance of genotype IA, with co-circulation of several genetic and antigenic variants. Phylogenetic analysis including Tunisian and strains from other regions of the world showed the presence of at least two IA-variants within IA subgenotype. Amino-acid analysis showed several mutations in or close to epitope regions in the VP1-region. This study provides a baseline on the genetic and antigenic variability of HAV circulating strains before the introduction of vaccination into the national immunization schedule.

Knockdown of Mitogen-Activated Protein Kinase Kinase 3 Negatively Regulates Hepatitis A Virus Replication.

Zinc chloride is known to be effective in combatting hepatitis A virus (HAV) infection, and zinc ions seem to be especially involved in Toll-like receptor (TLR) signaling pathways. In the present study, we examined this involvement in human hepatoma cell lines using a human TLR signaling target RT-PCR array. We also observed that zinc chloride inhibited mitogen-activated protein kinase kinase 3 (MAP2K3) expression, which could downregulate HAV replication in human hepatocytes. It is possible that zinc chloride may inhibit HAV replication in association with its inhibition of MAP2K3. In that regard, this study set out to determine whether MAP2K3 could be considered a modulating factor in the development of the HAV pathogen-associated molecular pattern (PAMP) and its triggering of interferon-ß production. Because MAP2K3 seems to play a role in antiviral immunity against HAV infection, it is a promising target for drug development. The inhibition of MAP2K3 may also prevent HAV patients from developing a severe hepatitis A infection.

An outbreak of acute jaundice syndrome (AJS) among the Rohingya refugees in Cox's Bazar, Bangladesh: Findings from enhanced epidemiological surveillance.

In the summer of 2017, an estimated 745,000 Rohingya fled to Bangladesh in what has been described as one of the largest and fastest growing refugee crises in the world. Among numerous health concerns, an outbreak of acute jaundice syndrome (AJS) was detected by the disease surveillance system in early 2018 among the refugee population. This paper describes the investigation into the increase in AJS cases, the process and results of the investigation, which were strongly suggestive of a large outbreak due to hepatitis A virus (HAV). An enhanced serological investigation was conducted between 28 February to 26 March 2018 to determine the etiologies and risk factors associated with the outbreak. A total of 275 samples were collected from 18 health facilities reporting AJS cases. Blood samples were collected from all patients fulfilling the study specific case definition and inclusion criteria, and tested for antibody responses using enzyme-linked immunosorbent assay (ELISA). Out of the 275 samples, 206 were positive for one of the agents tested. The laboratory results confirmed multiple etiologies including 154 (56%) samples tested positive for hepatitis A, 1 (0.4%) positive for hepatitis E, 36 (13%) positive for hepatitis B, 25 (9%) positive for hepatitis C, and 14 (5%) positive for leptospirosis. Among all specimens tested 24 (9%) showed evidence of co-infections with multiple etiologies. Hepatitis A and E are commonly found in refugee camps and have similar clinical presentations. In the absence of robust testing capacity when the epidemic was identified through syndromic reporting, a particular concern was that of a hepatitis E outbreak, for which immunity tends to be limited, and which may be particularly severe among pregnant women. This report highlights the challenges of identifying causative agents in such settings and the resources required to do so. Results from the month-long enhanced investigation did not point out widespread hepatitis E virus (HEV) transmission, but instead strongly suggested a large-scale hepatitis A outbreak of milder consequences, and highlighted a number of other concomitant causes of AJS (acute hepatitis B, hepatitis C, Leptospirosis), albeit most likely at sporadic level. Results strengthen the need for further water and sanitation interventions and are a stark reminder of the risk of other epidemics transmitted through similar routes in such settings, particularly dysentery and cholera. It also highlights the need to ensure clinical management capacity for potentially chronic conditions in this vulnerable population.

Hepatitis A virus strains identified in jogaejeot associated with outbreaks in Seoul, South Korea.

Jogaejeot, seasoned Venerupis philippinarum, is a traditional Korean fermented food, and hepatitis A virus (HAV) can be transmitted through contaminated food, especially bivalve shellfish, causing acute gastroenteritis worldwide. Here, we carried out a phylogenetic analysis to identify and characterize HAV strains in jogaejeot samples associated with hepatitis A (HA) outbreaks in Seoul, South Korea, in 2019. The HAV strains were identified using blast and molecular analysis of the amplified HAV VP1-P2B genome region. The HAV strains identified in the five jogaejeot samples shared at least 99% sequence identity, were all classified as genotype IA and were most closely related to strains that are widespread in East Asia. These results support a link between the consumption of jogaejeot and the HA outbreaks observed in 2019 in Seoul. In addition, they indicate a need for more stringent enforcement of food safety regulations for the shellfish industry, especially against HAV, and the value of widespread vaccination.

Virus-specific memory T cell responses unmasked by immune checkpoint blockade cause hepatitis.

Treatment of advanced melanoma with combined PD-1/CTLA-4 blockade commonly causes serious immune-mediated complications. Here, we identify a subset of patients predisposed to immune checkpoint blockade-related hepatitis who are distinguished by chronic expansion of effector memory CD4+ T cells (TEM cells). Pre-therapy CD4+ TEM cell expansion occurs primarily during autumn or winter in patients with metastatic disease and high cytomegalovirus (CMV)-specific serum antibody titres. These clinical features implicate metastasis-dependent, compartmentalised CMV reactivation as the cause of CD4+ TEM expansion. Pre-therapy CD4+ TEM expansion predicts hepatitis in CMV-seropositive patients, opening possibilities for avoidance or prevention. 3 of 4 patients with pre-treatment CD4+ TEM expansion who received αPD-1 monotherapy instead of αPD-1/αCTLA-4 therapy remained hepatitis-free. 4 of 4 patients with baseline CD4+ TEM expansion given prophylactic valganciclovir and αPD-1/αCTLA-4 therapy remained hepatitis-free. Our findings exemplify how pathogen exposure can shape clinical reactions after cancer therapy and how this insight leads to therapeutic innovations.

Male-Dominant Hepatitis A Outbreak Observed among Non-HIV-Infected Persons in the Northern Part of Tokyo, Japan.

Recently, we experienced an outbreak of acute hepatitis A virus (HAV) infection between 2018 and 2020. Herein, we describe this male-dominant HAV infection outbreak observed among non-human immunodeficiency virus (HIV)-infected persons in the northern part of Tokyo, Japan. Clinical information was collected from patient interviews and from medical record descriptions. In the present study, 21 patients were retrospectively analyzed. A total of 90.4 and 33.3% of patients were males, and men who have sex with men (MSM), respectively. The total bilirubin levels and platelet counts tended to be lower in the MSM group than in the non-MSM group. C-reactive protein (CRP) levels tended to be higher in acute liver failure (ALF) patients than in non-ALF patients. Prolonged cholestasis was observed in one patient (4.8%). We also found that 18 HAV isolates belonged to HAV subgenotype IA/subgroup 13 (S13), which clustered with the HAV isolate (KX151459) that was derived from an outbreak of HAV infection among MSM in Taiwan in 2015. Our results suggest that the application of antivirals against HAV, as well as HAV vaccines, would be useful for the treatment and prevention of severe HAV infection.

The Molecular Characterization of Hepatitis A Virus Strains Circulating during Hepatitis A Outbreaks in São Paulo, Brazil, from September 2017 to May 2019.

Outbreaks of hepatitis A may occur in countries of medium and high socioeconomic levels in which the population generally exhibits an increased susceptibility in young adults to this infection if they are not vaccinated against the hepatitis A virus (HAV). In Europe, an outbreak involved approximately 22 European countries with 4475 cases reported from 2016 to 2018; most of them were men who have sex with men (MSM). This outbreak expanded to North and South America, including Brazil, particularly in São Paulo city with 1547 reported cases from 2016 to 2019. In the present study, we characterized the HAV strains involved in the acute hepatitis A cases identified in the reference centers of São Paulo city during this outbreak. A total of 51 cases with positive anti-HAV IgM were included, 80.4% male, 68.6% of them between 20 and 40 years old and 41.7% MSM. HAV RNA was detected in 92% (47/51) of the cases. Subgenotype IA of HAV was identified and most of the strains were closely related to that isolated in outbreaks that occurred in different European countries in 2016. These results showed the epidemiological relation between these outbreaks and reinforce the need to implement vaccination against hepatitis A for the adult population, particularly for a population with a high-risk behavior.

Prognostic, clinical, and therapeutic importance of RANTES-CCR5 axis in hepatitis A infection: A multiapproach study.

Fulminant hepatic failure (FHF) is a lethal manifestation of hepatitis A virus (HAV) infection, whose underlying mechanisms are poorly understood. We aimed to evaluate the importance of the modulation of the RANTES-chemokine receptor type 5 (CCR5) signaling axis and its immunomodulatory effects in directing hepatitis A disease pathogenesis using an in silico, in vitro and patient cohort-based approach. In silico interaction studies were performed using computation approaches with suitable software. Differential expression of relevant cytokines and immune cell markers were studied using real-time quantitative reverse transcription PCR (qRT-PCR), enzyme-linked immunosorbent assay, and flow-cytometry-based methods. In the HepG2 cell line, we studied inflammatory responses and susceptibility to HAV infection following RANTES stimulation and antibody blockade of CCR5. The HAV-VP3 region exhibited high interaction in CCR5: HAV complexes. RANTES levels were significantly increased in FHF cases. Reduced monocyte and T-cell activation were observed in FHF cases. RANTES expression inversely correlated with viremia but positively correlated with proinflammatory responses. Hyper Th1-biased immune responses, marked by high interleukin (IL)-12/IL-10 ratio were observed in FHF cases, which were also characterized by upregulated tumor necrosis factor-alpha (TNF-α) expression and reduced interferon-gamma expression. In vitro, RANTES was protective against HAV infection but resulted in upregulated TNF-α expression. Although viral load increased upon the regulation of inflammatory responses by CCR5 blocking, it was still significantly lower compared to control HAV-infected cells. Our study suggests the importance of RANTES-CCR5 signaling and linked-immunomodulation in HAV disease pathogenesis, as well as highlights the utility of CCR5 antagonists as a risk-reduction strategy in FHF patients. Our findings, therefore, have important implications for the management of high-risk HAV infections.

Factors Associated With Hepatitis A Mortality During Person-to-Person Outbreaks: A Matched Case-Control Study-United States, 2016-2019.

BACKGROUND AND AIMS: During 2016-2020, the United States experienced person-to-person hepatitis A outbreaks that are unprecedented in the vaccine era, during which case-fatality ratios reported by some jurisdictions exceeded those historically associated with hepatitis A. APPROACH AND RESULTS: To identify factors associated with hepatitis A-related mortality, we performed a matched case-control study (matched on age [±5 years] and county of residence in a 1:4 ratio) using data collected from health department and hospital medical records of outbreak-associated patients in Kentucky, Michigan, and West Virginia. Controls were hepatitis A outbreak-associated patients who did not die. There were 110 cases (mean age 53.6 years) and 414 matched controls (mean age 51.9 years); most cases (68.2%) and controls (63.8%) were male. Significantly (P < 0.05) higher odds of mortality were associated with preexisting nonviral liver disease (adjusted odds ratio [aOR], 5.2), history of hepatitis B (aOR, 2.4), diabetes (aOR, 2.2), and cardiovascular disease (aOR, 2.2), as well as initial Model for End-Stage Liver Disease (MELD) score ≥ 30 (aOR, 10.0), aspartate aminotransferase (AST)/alanine aminotransferase (ALT) ratio > 2 (aOR, 10.3), and platelet count < 150,000/µL (aOR, 3.7) among hepatitis A outbreak-associated patients in the independent multivariable conditional logistic regression analyses (each model adjusted for sex). CONCLUSIONS: Preexisting liver disease, diabetes, cardiovascular disease, and initial MELD score ≥ 30, AST/ALT ratio ≥ 1, and platelet count < 150,000/µL among hepatitis A patients were independently associated with higher odds of mortality. Providers should be vigilant for such features and have a low threshold to escalate care and consider consultation for liver transplantation. Our findings support the recommendation of the Advisory Committee on Immunization Practices to vaccinate persons with chronic liver disease, though future recommendations to include adults with diabetes and cardiovascular disease should be considered.

Genetic diversity, haplotype analysis, and risk factor assessment of hepatitis A virus isolates from the West Bank, Palestine during the period between 2014 and 2016.

BACKGROUND: Hepatitis A virus (HAV) infection is one of the major causes of acute viral hepatitis. HAV genotypes and its genetic diversity is rarely investigated in our region as well as worldwide. AIMS: The aims of the present study were to determine the HAV genotypes and its risk factors and to investigate the genetic diversity of the HAV isolates in the West Bank, Palestine. STUDY DESIGN: A cohort of 161 clinically and laboratory-confirmed HAV (IgM-positive) cases and 170 apparently healthy controls from all the districts of the West Bank, Palestine during the period of 2014 to 2016 were tested for HAV infection using IgM antibodies, RT-PCR and sequence analysis of the VP3/VP1 junction region of the HAV genome. Phylogenetic analysis, genetic diversity and haplotypes analysis were used to characterize the VP3/VP1 sequences. RESULTS: All the 34 sequences of the HAV were found to be of HAV-IB sub-genotype. The phylogenetic analysis showed four main clusters with cluster III exclusively consisting of 18 Palestinian isolates (18/23-78%), but with weak bootstrap values. A high haplotype diversity (Hd) and low nucleotide diversity (π) were observed. Cluster III showed high number of haplotypes (h = 8), but low haplotype (gene) diversity (Hd = 0.69). A total of 28 active haplotypes with some consisting of more than one sequence were observed using haplotype network analysis. The Palestinian haplotypes are characterized by closely related viral haplotypes with one SNV away from each other which ran parallel to cluster III in the phylogenetic tree. A smaller Palestinian haplotype (4 isolates) was three SNVs away from the major haplotype cluster (n = 10) and closer to others haplotypes from Iran, Spain, and South Africa. Young age, low level of parent's education, infrequent hand washing before meals, and drinking of un-treated water were considered the major HAV risk factors in the present study. CONCLUSION: Haplotype network analysis revealed haplotype variation among the HAV Palestinian sequences despite low genetic variation and nucleotide diversity. In addition, this study reconfirmed that age and parent's level of education as HAV risk factors, while hand washing and treating drinking water as protective factors.

Hepatitis A Virus Strains Circulating in the Campania Region (2015-2018) Assessed through Bivalve Biomonitoring and Environmental Surveillance.

The genetic diversity of Hepatitis A Virus (HAV) circulating in the Campania Region in years 2015-2018 was investigated through the monitoring of sentinel bivalve shellfish and water matrices. Overall, 463 water samples (71 sewage samples, 353 coastal discharge waters, and 39 seawaters samples), and 746 bivalve shellfish samples were analyzed. Positivity for HAV was detected in 20/71 sewage samples, 14/353 coastal discharge waters, 5/39 seawaters, and 102/746 bivalve shellfish. Sixty-one of the positive samples were successfully sequenced and were characterized as genotype IA (n = 50) and IB (n = 11). The prevalent strain circulating in 2015 in both bivalves and waters was the IA strain responsible for the outbreak occurring around the same time in the Naples area. This variant was no longer identified in subsequent years (2017-2018) when, instead, appeared two of the IA variants of the multistate outbreak affecting men who have sex with men (MSM), VRD_521_2016, and RIVM-HAV16-090, with the former prevailing in both shellfish and water environments. HAV IB isolates were detected over the years in shellfish and in water matrices, but not in clinical samples, suggesting that this genotype had been circulating silently. An integrated surveillance system (environment/food/clinical cases) can be a useful tool to monitor changes in viral variants in the population, as well as an early warning system.

Co-Occurrence of Hepatitis A Infection and Chronic Liver Disease.

Hepatitis A virus (HAV) infection occasionally leads to a critical condition in patients with or without chronic liver diseases. Acute-on-chronic liver disease includes acute-on-chronic liver failure (ACLF) and non-ACLF. In this review, we searched the literature concerning the association between HAV infection and chronic liver diseases in PubMed. Chronic liver diseases, such as metabolic associated fatty liver disease and alcoholic liver disease, coinfection with other viruses, and host genetic factors may be associated with severe hepatitis A. It is important to understand these conditions and mechanisms. There may be no etiological correlation between liver failure and HAV infection, but there is an association between the level of chronic liver damage and the severity of acute-on-chronic liver disease. While the application of an HAV vaccination is important for preventing HAV infection, the development of antivirals against HAV may be important for preventing the development of ACLF with HAV infection as an acute insult. The latter is all the more urgent given that the lives of patients with HAV infection and a chronic liver disease of another etiology may be at immediate risk.