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Brown HM, Marlet J, León-Janampa N, Brand D, Fletcher NF. Enhanced hepatitis E virus infection of polarised hepatocytes in vitro. Sci Rep 2025; 15:7598. [PMID: 40038434 PMCID: PMC11880378 DOI: 10.1038/s41598-025-92164-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2024] [Accepted: 02/25/2025] [Indexed: 03/06/2025] Open
Abstract
Hepatitis E virus (HEV) is a leading cause of acute viral hepatitis worldwide, and the only zoonotic hepatitis virus. HEV genotype 3 (HEV3) is associated with a range of clinical presentations including chronic infection in immunocompromised individuals in developed nations as well as sporadic cases of autochthonous HEV3 in Europe. Current in vitro models support low levels of HEV infection, hampering our understanding of viral pathogenesis and development of therapeutics. We developed modified culture methods for two widely used hepatoma cell lines, PLC-PRF-5 and Huh-7.5, and evaluated HEV infection. Simple epithelial-like polarity and differentiation formed in PLC-PRF-5 cells, evidenced by localisation of tight junction proteins occludin and zona-occludin 1 to intercellular junctions, and increased albumin production. Complex hepatocyte-like polarity was observed in Huh-7.5 cells, with tight junction proteins localised to shared internal bile canaliculi-like structures and retention of the fluorescent molecule, 5(6)-Carboxyfluorescein diacetate. Cells were infected with genotype 3 HEV, and enhanced infection and replication of HEV was observed using RT-qPCR and immunofluorescent labelling of HEV ORF2 and dsRNA. We describe robust, accessible models for HEV infection in vitro. These models will allow studies to further our understanding of this emerging zoonotic pathogen and develop therapeutic interventions.
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Affiliation(s)
- Hannah M Brown
- Veterinary Sciences Centre and Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland
| | - Julien Marlet
- INSERM U1259 MAVIVHe, CHRU de Tours, Université de Tours et CHRU de Tours and Service de Bactériologie-Virologie-Hygiène, Tours, France
| | - Nancy León-Janampa
- INSERM U1259 MAVIVHe, Université de Tours et CHRU de Tours, Tours, France
| | - Denys Brand
- INSERM U1259 MAVIVHe, CHRU de Tours, Université de Tours et CHRU de Tours and Service de Bactériologie-Virologie-Hygiène, Tours, France
| | - Nicola F Fletcher
- Veterinary Sciences Centre and Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
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2
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Liu X, Liu T, Shao Z, Xiong X, Qi S, Guan J, Wang M, Tang YD, Feng Z, Wang L, Yin X. Palmitoylation-dependent association with Annexin II directs hepatitis E virus ORF3 sorting into vesicles and quasi-enveloped virions. Proc Natl Acad Sci U S A 2025; 122:e2418751122. [PMID: 39793027 PMCID: PMC11725905 DOI: 10.1073/pnas.2418751122] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2024] [Accepted: 11/25/2024] [Indexed: 01/12/2025] Open
Abstract
Historically considered to be nonenveloped, hepatitis E virus (HEV), an important zoonotic pathogen, has recently been discovered to egress from infected cells as quasi-enveloped virions. These quasi-enveloped virions circulating in the blood are resistant to neutralizing antibodies, thereby facilitating the stealthy spread of infection. Despite abundant evidence of the essential role of the HEV-encoded ORF3 protein in quasi-enveloped virus formation, the underlying mechanism remains unclear. Here, we demonstrate that the HEV ORF3 protein possesses an inherent capacity for self-secretion and that palmitoylation at two cysteine residues within the ORF3 N-terminal region is essential for its secretion and quasi-enveloped virus formation. We further found that only palmitoylated ORF3 proteins hijacked Annexin II for transport to the cytoskeleton and are then directed into multivesicular bodies through the nSMase-endosomal sorting complexes required for transport-III pathway for secretion. Finally, we show that infection of gerbils with HEV mutants harboring mutations at palmitoylation sites within ORF3 showed no fecal viral shedding but competent replication in the liver. Our study fills a gap in the understanding of the assembly and release of quasi-enveloped virions mediated by ORF3 and offers the potential for designing therapeutic strategies to control HEV infection.
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Affiliation(s)
- Xing Liu
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Tianxu Liu
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing100191, China
| | - Zhen Shao
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Xiaoyan Xiong
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
- Department of Animal Sciences, Quantitative Veterinary Epidemiology Group, Wageningen University, Wageningen6700 AH, The Netherlands
| | - Shuhui Qi
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Junyong Guan
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Menghang Wang
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Yan-Dong Tang
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
| | - Zongdi Feng
- Center for Vaccines and Immunity, The Research Institute at Nationwide Children’s Hospital, Columbus, OH43205
- Department of Pediatrics, The Ohio State University College of Medicine, Columbus, OH43205
| | - Lin Wang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing100191, China
| | - Xin Yin
- Division of Livestock Infectious Diseases, State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin150069, China
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Nagashima S, Primadharsini PP, Takahashi M, Nishiyama T, Murata K, Okamoto H. Role of Rab13, Protein Kinase A, and Zonula Occludens-1 in Hepatitis E Virus Entry and Cell-to-Cell Spread: Comparative Analysis of Quasi-Enveloped and Non-Enveloped Forms. Pathogens 2024; 13:1130. [PMID: 39770389 PMCID: PMC11678111 DOI: 10.3390/pathogens13121130] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2024] [Revised: 12/16/2024] [Accepted: 12/17/2024] [Indexed: 01/11/2025] Open
Abstract
Hepatitis E virus (HEV) exists in two distinct forms: a non-enveloped form (neHEV), which is present in feces and bile, and a quasi-enveloped form (eHEV), found in circulating blood and culture supernatants. This study aimed to elucidate the roles of Ras-associated binding 13 (Rab13) and protein kinase A (PKA) in the entry mechanisms of both eHEV and neHEV, utilizing small interfering RNA (siRNA) and chemical inhibitors. The results demonstrated that the entry of both viral forms is dependent on Rab13 and PKA. Further investigation into the involvement of tight junction (TJ) proteins revealed that the targeted knockdown of zonula occludens-1 (ZO-1) significantly impaired the entry of both eHEV and neHEV. In addition, in ZO-1 knockout (KO) cells inoculated with either viral form, HEV RNA levels in culture supernatants did not increase, even up to 16 days post-inoculation. Notably, the absence of ZO-1 did not affect the adsorption efficiency of eHEV or neHEV, nor did it influence HEV RNA replication. In cell-to-cell spread assays, ZO-1 KO cells inoculated with eHEV showed a lack of expression of HEV ORF2 and ORF3 proteins. In contrast, neHEV-infected ZO-1 KO cells showed markedly reduced ORF2 and ORF3 protein expression within virus-infected foci, compared to non-targeting knockout (NC KO) cells. These findings underscore the crucial role of ZO-1 in facilitating eHEV entry and mediating the cell-to-cell spread of neHEV in infected cells.
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Affiliation(s)
- Shigeo Nagashima
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke-shi 329-0498, Tochigi, Japan; (P.P.P.); (M.T.); (T.N.); (K.M.)
| | | | | | | | | | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke-shi 329-0498, Tochigi, Japan; (P.P.P.); (M.T.); (T.N.); (K.M.)
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4
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Santos-Silva S, Romalde JL, Bento JT, Cruz AVS, López-López P, Gonçalves HMR, Van der Poel WHM, Nascimento MSJ, Rivero-Juarez A, Mesquita JR. Serological and Molecular Survey of Hepatitis E Virus in Small Ruminants from Central Portugal. FOOD AND ENVIRONMENTAL VIROLOGY 2024; 16:516-524. [PMID: 39235492 PMCID: PMC11525313 DOI: 10.1007/s12560-024-09612-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/20/2024] [Accepted: 08/26/2024] [Indexed: 09/06/2024]
Abstract
Hepatitis E virus (HEV) is currently recognized as an emerging problem and a growing concern for public health in developed countries, with HEV infections mainly attributable to foodborne transmission of HEV-3. The zoonotic HEV genotype 3 infects a wide range of mammalian hosts, with swine considered as the primary host. This study investigates the occurrence of HEV among small ruminants in Portugal. The primary aim of the present research was to evaluate the circulation and the potential for HEV infection among sheep and goats. A total of 400 bile samples and 493 blood samples were collected from sheep and goats at a slaughterhouse in the center region of Portugal, between January 2022 and March 2023. The HEV RNA detection in bile samples was performed using a nested broad-spectrum RT-PCR targeting the ORF1 region. Serological analysis to detect anti-HEV antibodies was conducted using a commercial double-antigen sandwich multi-species ELISA. The HEV RNA was not detected in any bile samples using the nested broad-spectrum RT-PCR. Serological analysis revealed an overall HEV antibody seroprevalence of 2% (10/493, 95% CI: 0.98-3.70) among the small ruminants, namely 2.2% in goats and 2.0% in sheep. Curiously, no statistically significant association among the factors, age, sex and species and HEV seroprevalence was observed. Although HEV RNA was not detected in the bile of sheep and goats, this study the evidence of seroprevalence in these small ruminant species. Further research could provide additional insights into the factors influencing HEV transmission dynamics in small ruminants in Portugal and its potential implications for public health.
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Affiliation(s)
- Sérgio Santos-Silva
- School of Medicine and Biomedical Sciences (ICBAS), University of Porto, Porto, Portugal
| | - Jesús L Romalde
- Department of Microbiology and Parasitology, CIBUS-Faculty of Biology, Cross-Disciplinary Research Center in Environmental Technologies (CRETUS), Universidad de Santiago de Compostela, Santiago de Compostela, Spain
| | | | - Andreia V S Cruz
- School of Medicine and Biomedical Sciences (ICBAS), University of Porto, Porto, Portugal
| | - Pedro López-López
- Unit of Infectious Diseases, Clinical Virology and Zoonoses, Hospital Universitario Reina Sofia, Instituto Maimonides de Investigación Biomédica de Córdoba (IMIBIC), Universidad de Córdoba (UCO), Cordoba, Spain
- Center for Biomedical Research Network (CIBER) in Infectious Diseases, Health Institute Carlos III, Madrid, Spain
| | - Helena M R Gonçalves
- LAQV, REQUIMTE, Department of Chemistry and Biochemistry, Faculty of Sciences, University of Porto, Porto, Portugal
| | - Wim H M Van der Poel
- Quantitative Veterinary Epidemiology, Wageningen University, Wageningen, The Netherlands
- Department Virology & Molecular Biology, Wageningen Bioveterinary Research, Lelystad, The Netherlands
| | | | - António Rivero-Juarez
- Unit of Infectious Diseases, Clinical Virology and Zoonoses, Hospital Universitario Reina Sofia, Instituto Maimonides de Investigación Biomédica de Córdoba (IMIBIC), Universidad de Córdoba (UCO), Cordoba, Spain
- Center for Biomedical Research Network (CIBER) in Infectious Diseases, Health Institute Carlos III, Madrid, Spain
| | - João R Mesquita
- School of Medicine and Biomedical Sciences (ICBAS), University of Porto, Porto, Portugal.
- Epidemiology Research Unit (EPIUnit), Instituto de Saúde Pública da Universidade Do Porto, Porto, Portugal.
- Laboratory for Integrative and Translational Research in Population Health (ITR), Porto, Portugal.
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5
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Chu YD, Chen MC, Yeh CT, Lai MW. Hijacking host extracellular vesicle machinery by hepatotropic viruses: current understandings and future prospects. J Biomed Sci 2024; 31:97. [PMID: 39369194 PMCID: PMC11453063 DOI: 10.1186/s12929-024-01063-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2024] [Accepted: 06/25/2024] [Indexed: 10/07/2024] Open
Abstract
Recent advances in studies exploring the roles of extracellular vesicles (EVs) in viral transmission and replication have illuminated hepatotropic viruses, such as hepatitis A (HAV), hepatitis B (HBV), hepatitis C (HCV), hepatitis D (HDV), and hepatitis E (HEV). While previous investigations have uncovered these viruses' ability to exploit cellular EV pathways for replication and transmission, most have focused on the impacts of exosomal pathways. With an improved understanding of EVs, four main subtypes, including exosomes, microvesicles, large oncosomes, and apoptotic bodies, have been categorized based on size and biogenic pathways. However, there remains a noticeable gap in comprehensive reviews summarizing recent findings and outlining future perspectives for EV studies related to hepatotropic viruses. This review aims to consolidate insights into EV pathways utilized by hepatotropic viruses, offering guidance for the future research direction in this field. By comprehending the diverse range of hepatotropic virus-associated EVs and their role in cellular communication during productive viral infections, this review may offer valuable insights for targeting therapeutics and devising strategies to combat virulent hepatotropic virus infections and the associated incidence of liver cancer.
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Affiliation(s)
- Yu-De Chu
- Liver Research Center, Chang Gung Memorial Hospital, 5F., No. 15, Wenhua 1st Rd., Guishan Dist., Taoyuan City, 333, Taiwan
| | - Mi-Chi Chen
- Liver Research Center, Chang Gung Memorial Hospital, 5F., No. 15, Wenhua 1st Rd., Guishan Dist., Taoyuan City, 333, Taiwan
- Department of Pediatric, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Chau-Ting Yeh
- Liver Research Center, Chang Gung Memorial Hospital, 5F., No. 15, Wenhua 1st Rd., Guishan Dist., Taoyuan City, 333, Taiwan.
- Institute of Stem Cell and Translational Cancer Research, Chang Gung Memorial Hospital, Taoyuan, Taiwan.
| | - Ming-Wei Lai
- Liver Research Center, Chang Gung Memorial Hospital, 5F., No. 15, Wenhua 1st Rd., Guishan Dist., Taoyuan City, 333, Taiwan.
- Department of Pediatric, Chang Gung Memorial Hospital, Taoyuan, Taiwan.
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6
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Brüggemann Y, Klöhn M, Wedemeyer H, Steinmann E. Hepatitis E virus: from innate sensing to adaptive immune responses. Nat Rev Gastroenterol Hepatol 2024; 21:710-725. [PMID: 39039260 DOI: 10.1038/s41575-024-00950-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 05/29/2024] [Indexed: 07/24/2024]
Abstract
Hepatitis E virus (HEV) infections are a major cause of acute viral hepatitis in humans worldwide. In immunocompetent individuals, the majority of HEV infections remain asymptomatic and lead to spontaneous clearance of the virus, and only a minority of individuals with infection (5-16%) experience symptoms of acute viral hepatitis. However, HEV infections can cause up to 30% mortality in pregnant women, become chronic in immunocompromised patients and cause extrahepatic manifestations. A growing body of evidence suggests that the host immune response to infection with different HEV genotypes is a critical determinant of distinct HEV infection outcomes. In this Review, we summarize key components of the innate and adaptive immune responses to HEV, including the underlying immunological mechanisms of HEV associated with acute and chronic liver failure and interactions between T cell and B cell responses. In addition, we discuss the current status of vaccines against HEV and raise outstanding questions regarding the immune responses induced by HEV and treatment of the disease, highlighting areas for future investigation.
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Affiliation(s)
- Yannick Brüggemann
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Mara Klöhn
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Heiner Wedemeyer
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany
- German Center for Infection Research (DZIF), Partner Sites Hannover-Braunschweig, Hannover, Germany
- Cluster of Excellence RESIST (EXC 2155), Hannover Medical School, Hannover, Germany
| | - Eike Steinmann
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany.
- German Center for Infection Research (DZIF), External Partner Site, Bochum, Germany.
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7
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Falkenhagen A, Panajotov J, Johne R. Colon-derived Caco-2 cells support replication of hepatitis E virus genotype 1 strain Sar55 generated by reverse genetics. Virus Res 2024; 347:199427. [PMID: 38917940 PMCID: PMC11261143 DOI: 10.1016/j.virusres.2024.199427] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2024] [Revised: 06/20/2024] [Accepted: 06/22/2024] [Indexed: 06/27/2024]
Abstract
The hepatitis E virus (HEV) is infecting over 20 million people annually with a high morbidity especially in pregnant women and immune-suppressed individuals. While HEV genotype 1 (HEV-1) infects only humans, genotype 3 (HEV-3) is zoonotic and commonly transmitted from infected animals to humans. Whereas a few reverse genetics systems enabling targeted genome manipulations exist for HEV-3, those for HEV-1 are still very limited, mainly because of inefficient cell culture replication. Here, the generation of HEV-1 strain Sar55 and HEV-3 strain 47832mc by transfecting in vitro-transcribed and capped virus genomes into different cell lines was attempted. Culture supernatants of colon-derived colorectal adenocarcinoma cell line Caco-2 contained HEV-1 and HEV-3 capable of infecting Caco-2 cells. Density gradient centrifugation analyses of culture supernatants confirmed that HEV-1 particles were quasi-enveloped in analogy to HEV-3 and that non-virion-associated capsid protein was secreted from cells. Following transfection or infection of Caco-2 cells, HEV-1 consistently reached higher titers than HEV-3 in culture supernatants, but HEV-1 generated by transfection of Caco-2 cells was unable to efficiently infect hepatoma cell lines PLC/PRF/5 or HuH7-Lunet BLR. Taken together, our results indicate that HEV-1 is able to exert a complete replication cycle in Caco-2 cells. An efficient cell culture system for this genotype will be useful for studying species tropism, but further research is required to determine the significance of HEV-1 replication in colon-derived cells.
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Affiliation(s)
- Alexander Falkenhagen
- Department of Biological Safety, German Federal Institute for Risk Assessment, Max-Dohrn-Str. 8-10, 10589 Berlin, Germany.
| | - Jessica Panajotov
- Department of Biological Safety, German Federal Institute for Risk Assessment, Max-Dohrn-Str. 8-10, 10589 Berlin, Germany
| | - Reimar Johne
- Department of Biological Safety, German Federal Institute for Risk Assessment, Max-Dohrn-Str. 8-10, 10589 Berlin, Germany
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Verbrugghe G, Soudan-Foulques C, Fraisse A, Waldman Vigne P, Perelle S, Ndoye FT, Martin-Latil S. A Useful Method to Provide Infectious and Cultivable In Vitro Naked Viral Particles of Hepatitis A Virus. Viruses 2024; 16:1360. [PMID: 39339837 PMCID: PMC11435643 DOI: 10.3390/v16091360] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/23/2024] [Revised: 08/15/2024] [Accepted: 08/20/2024] [Indexed: 09/30/2024] Open
Abstract
Hepatitis A virus (HAV) is an enteric virus mainly transmitted by the faecal-oral route. Belonging to the Picornaviridae family, HAV was first described as small naked particles, like all viruses of this family. However, for about a decade, it was demonstrated that HAV particles can exist surrounded by a lipid bilayer. This type of particle, called enveloped HAV (eHAV), acquires its lipid bilayer by hijacking a part of cell membranes during the virion egress in the last steps of the viral cycle. In vitro culture systems produce mainly eHAV, and so, to date, most of the studies on HAV have been carried out using this type of viral particle. In this study, a method based on lipid bilayer removal by chemical delipidation is proposed for the production of naked HAV particles. The resulting naked HAV particles conserve their infectivity and are therefore fully cultivable in vitro. By using this method, naked HAV particles can easily be produced in vitro and can be useful to perform further studies such as inactivation processes for the food industry, as HAV is a main concern for food safety.
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Affiliation(s)
- Gwenaëlle Verbrugghe
- Université Paris-Saclay, INRAE, UR FRISE, 92160 Antony, France;
- ANSES, Laboratory for Food Safety, UVE, 94700 Maisons-Alfort, France; (A.F.); (S.P.)
| | - Chloé Soudan-Foulques
- ANSES, Animal Health Laboratory, UMR1161 Virology, INRAe, Anses, ENVA, 94700 Maisons-Alfort, France;
| | - Audrey Fraisse
- ANSES, Laboratory for Food Safety, UVE, 94700 Maisons-Alfort, France; (A.F.); (S.P.)
| | | | - Sylvie Perelle
- ANSES, Laboratory for Food Safety, UVE, 94700 Maisons-Alfort, France; (A.F.); (S.P.)
| | | | - Sandra Martin-Latil
- ANSES, Laboratory for Food Safety, UVE, 94700 Maisons-Alfort, France; (A.F.); (S.P.)
- ANSES, Animal Health Laboratory, UMR1161 Virology, INRAe, Anses, ENVA, 94700 Maisons-Alfort, France;
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Letafati A, Taghiabadi Z, Roushanzamir M, Memarpour B, Seyedi S, Farahani AV, Norouzi M, Karamian S, Zebardast A, Mehrabinia M, Ardekani OS, Fallah T, Khazry F, Daneshvar SF, Norouzi M. From discovery to treatment: tracing the path of hepatitis E virus. Virol J 2024; 21:194. [PMID: 39180020 PMCID: PMC11342613 DOI: 10.1186/s12985-024-02470-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2024] [Accepted: 08/14/2024] [Indexed: 08/26/2024] Open
Abstract
The hepatitis E virus (HEV) is a major cause of acute viral hepatitis worldwide. HEV is classified into eight genotypes, labeled HEV-1 through HEV-8. Genotypes 1 and 2 exclusively infect humans, while genotypes 3, 4, and 7 can infect both humans and animals. In contrast, genotypes 5, 6, and 8 are restricted to infecting animals. While most individuals with a strong immune system experience a self-limiting infection, those who are immunosuppressed may develop chronic hepatitis. Pregnant women are particularly vulnerable to severe illness and mortality due to HEV infection. In addition to liver-related complications, HEV can also cause extrahepatic manifestations, including neurological disorders. The immune response is vital in determining the outcome of HEV infection. Deficiencies in T cells, NK cells, and antibody responses are linked to poor prognosis. Interestingly, HEV itself contains microRNAs that regulate its replication and modify the host's antiviral response. Diagnosis of HEV infection involves the detection of HEV RNA and anti-HEV IgM/IgG antibodies. Supportive care is the mainstay of treatment for acute infection, while chronic HEV infection may be cleared with the use of ribavirin and pegylated interferon. Prevention remains the best approach against HEV, focusing on sanitation infrastructure improvements and vaccination, with one vaccine already licensed in China. This comprehensive review provides insights into the spread, genotypes, prevalence, and clinical effects of HEV. Furthermore, it emphasizes the need for further research and attention to HEV, particularly in cases of acute hepatitis, especially among solid-organ transplant recipients.
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Affiliation(s)
- Arash Letafati
- Department of Virology, Faculty of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran.
| | - Zahra Taghiabadi
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Mahshid Roushanzamir
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
- Department of Pharmacological and Biomolecular Science, University of Milan, Milan, Italy
| | - Bahar Memarpour
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
- Shahid Chamran University of Ahvaz, Ahvaz, Iran
| | - Saba Seyedi
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | | | - Masoomeh Norouzi
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Saeideh Karamian
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Arghavan Zebardast
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Marzieh Mehrabinia
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Omid Salahi Ardekani
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Tina Fallah
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Fatemeh Khazry
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Samin Fathi Daneshvar
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
| | - Mehdi Norouzi
- Department of Virology, Faculty of Public Health, Tehran University of Medical Sciences, Tehran, Iran
- Research Center for Clinical Virology, Tehran University of Medical Science, Tehran, Iran
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10
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Jiao F, Zhao Y, Zhou G, Meng C, Wang L, Wu S, Li J, Cao L, Zhou B, Luo Y, Jiao H. Multiple Functions of Hepatitis E Virus ORF3. Microorganisms 2024; 12:1405. [PMID: 39065173 PMCID: PMC11278674 DOI: 10.3390/microorganisms12071405] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2024] [Revised: 07/05/2024] [Accepted: 07/08/2024] [Indexed: 07/28/2024] Open
Abstract
Hepatitis E (Hepatitis E, HE) is an acute and chronic infectious hepatitis caused by hepatitis E virus (Hepatitis E Virus, HEV) infection, which is responsible for most acute hepatitis in the world and is a significant public health problem. The pathogen, HEV, has three Open Reading Frames (ORFs) ORF1, ORF2, and ORF3, each of which has a different function. Most of the current research is focused on ORF1 and ORF2, while the research on ORF3 is still relatively small. To provide more ideas for the study of HEV pathogenesis and the prevention and treatment of HE, this paper reviews the effects of ORF3 on the ERK pathway, growth factors, immune response, and virus release.
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Affiliation(s)
- Fengyuan Jiao
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Yu Zhao
- Ministry of Agriculture and Rural Affairs Key Laboratory of Crop Genitic Resources and Germplasm Innovation in Karst Region, Institute of Animal Husbandry and Veterinary Medicine of Guizhou Academy of Agricultural Science, Guiyang 550005, China;
| | - Gengxu Zhou
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Chi Meng
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Lingjie Wang
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Shengping Wu
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Jixiang Li
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Liting Cao
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Bo Zhou
- Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Yujinxiang Street 573, Changchun 130102, China;
| | - Yichen Luo
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
| | - Hanwei Jiao
- The College of Veterinary Medicine, Southwest University, Chongqing 402460, China; (F.J.); (G.Z.); (C.M.); (L.W.); (S.W.); (J.L.); (L.C.)
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Orosz L, Sárvári KP, Dernovics Á, Rosztóczy A, Megyeri K. Pathogenesis and clinical features of severe hepatitis E virus infection. World J Virol 2024; 13:91580. [PMID: 38984076 PMCID: PMC11229844 DOI: 10.5501/wjv.v13.i2.91580] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/31/2023] [Revised: 02/08/2024] [Accepted: 04/15/2024] [Indexed: 06/24/2024] Open
Abstract
The hepatitis E virus (HEV), a member of the Hepeviridae family, is a small, non-enveloped icosahedral virus divided into eight distinct genotypes (HEV-1 to HEV-8). Only genotypes 1 to 4 are known to cause diseases in humans. Genotypes 1 and 2 commonly spread via fecal-oral transmission, often through the consumption of contaminated water. Genotypes 3 and 4 are known to infect pigs, deer, and wild boars, often transferring to humans through inadequately cooked meat. Acute hepatitis caused by HEV in healthy individuals is mostly asymptomatic or associated with minor symptoms, such as jaundice. However, in immunosuppressed individuals, the disease can progress to chronic hepatitis and even escalate to cirrhosis. For pregnant women, an HEV infection can cause fulminant liver failure, with a potential mortality rate of 25%. Mortality rates also rise amongst cirrhotic patients when they contract an acute HEV infection, which can even trigger acute-on-chronic liver failure if layered onto pre-existing chronic liver disease. As the prevalence of HEV infection continues to rise worldwide, highlighting the particular risks associated with severe HEV infection is of major medical interest. This text offers a brief summary of the characteristics of hepatitis developed by patient groups at an elevated risk of severe HEV infection.
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Affiliation(s)
- László Orosz
- Department of Medical Microbiology, University of Szeged, Szeged 6720, Csongrád-Csanád, Hungary
| | - Károly Péter Sárvári
- Department of Medical Microbiology, University of Szeged, Szeged 6720, Csongrád-Csanád, Hungary
| | - Áron Dernovics
- Department of Medical Microbiology, University of Szeged, Szeged 6720, Csongrád-Csanád, Hungary
| | - András Rosztóczy
- Department of Internal Medicine, Division of Gastroenterology, University of Szeged, Szeged 6725, Csongrád-Csanád, Hungary
| | - Klára Megyeri
- Department of Medical Microbiology, University of Szeged, Szeged 6720, Csongrád-Csanád, Hungary
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12
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Shahini E, Argentiero A, Andriano A, Losito F, Maida M, Facciorusso A, Cozzolongo R, Villa E. Hepatitis E Virus: What More Do We Need to Know? MEDICINA (KAUNAS, LITHUANIA) 2024; 60:998. [PMID: 38929615 PMCID: PMC11205503 DOI: 10.3390/medicina60060998] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/10/2024] [Revised: 06/14/2024] [Accepted: 06/17/2024] [Indexed: 06/28/2024]
Abstract
Hepatitis E virus (HEV) infection is typically a self-limiting, acute illness that spreads through the gastrointestinal tract but replicates in the liver. However, chronic infections are possible in immunocompromised individuals. The HEV virion has two shapes: exosome-like membrane-associated quasi-enveloped virions (eHEV) found in circulating blood or in the supernatant of infected cell cultures and non-enveloped virions ("naked") found in infected hosts' feces and bile to mediate inter-host transmission. Although HEV is mainly spread via enteric routes, it is unclear how it penetrates the gut wall to reach the portal bloodstream. Both virion types are infectious, but they infect cells in different ways. To develop personalized treatment/prevention strategies and reduce HEV impact on public health, it is necessary to decipher the entry mechanism for both virion types using robust cell culture and animal models. The contemporary knowledge of the cell entry mechanism for these two HEV virions as possible therapeutic target candidates is summarized in this narrative review.
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Affiliation(s)
- Endrit Shahini
- Gastroenterology Unit, National Institute of Gastroenterology-IRCCS “Saverio de Bellis”, Castellana Grotte, 70013 Bari, Italy; (F.L.); (R.C.)
| | | | - Alessandro Andriano
- Department of Precision and Regenerative Medicine and Ionian Area, University of Bari Aldo Moro Medical School, 70124 Bari, Italy;
| | - Francesco Losito
- Gastroenterology Unit, National Institute of Gastroenterology-IRCCS “Saverio de Bellis”, Castellana Grotte, 70013 Bari, Italy; (F.L.); (R.C.)
| | - Marcello Maida
- Gastroenterology and Endoscopy Unit, S. Elia-Raimondi Hospital, 93100 Caltanissetta, Italy;
| | - Antonio Facciorusso
- Department of Medical and Surgical Sciences, University of Foggia, 71122 Foggia, Italy;
| | - Raffaele Cozzolongo
- Gastroenterology Unit, National Institute of Gastroenterology-IRCCS “Saverio de Bellis”, Castellana Grotte, 70013 Bari, Italy; (F.L.); (R.C.)
| | - Erica Villa
- Gastroenterology Unit, CHIMOMO Department, University of Modena & Reggio Emilia, Via del Pozzo 71, 41121 Modena, Italy
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13
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Li X, Sun X, Pinpin J, Zhao Q, Sun Y. Multifunctional ORF3 protein of hepatitis E virus. J Med Virol 2024; 96:e29691. [PMID: 38783788 DOI: 10.1002/jmv.29691] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2024] [Revised: 04/23/2024] [Accepted: 05/14/2024] [Indexed: 05/25/2024]
Abstract
Hepatitis E virus (HEV) is an emerging zoonotic pathogen that is transmitted primarily through the fecal-oral route and can cause acute hepatitis in humans. Since HEV was identified as a zoonotic pathogen, different species of HEV strains have been globally identified from various hosts, leading to an expanding range of hosts. The HEV genome consists of a 5' noncoding region, three open reading frames (ORFs), and a 3' noncoding region. The ORF3 protein is the smallest but has many functions in HEV release and pathogenesis. In this review, we systematically summarize recent progress in understanding the functions of the HEV ORF3 protein in virion release, biogenesis of quasi-enveloped viruses, antigenicity, and host environmental regulation. This review will help us to understand HEV replication and pathogenesis mechanisms better.
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Affiliation(s)
- Xiaoxuan Li
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
| | - Xuwen Sun
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
| | - Ji Pinpin
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
| | - Qin Zhao
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
| | - Yani Sun
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
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Primadharsini PP, Takahashi M, Nishizawa T, Sato Y, Nagashima S, Murata K, Okamoto H. The Full-Genome Analysis and Generation of an Infectious cDNA Clone of a Genotype 6 Hepatitis E Virus Variant Obtained from a Japanese Wild Boar: In Vitro Cultivation in Human Cell Lines. Viruses 2024; 16:842. [PMID: 38932135 PMCID: PMC11209168 DOI: 10.3390/v16060842] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2024] [Revised: 05/22/2024] [Accepted: 05/22/2024] [Indexed: 06/28/2024] Open
Abstract
Hepatitis E virus (HEV) can cause self-limiting acute and chronic hepatitis infections, particularly in immunocompromised individuals. In developing countries, HEV is mainly transmitted via drinking contaminated water, whereas zoonotic transmission dominates the route of infection in developed countries, including Japan. Pigs are an important reservoir for HEV infection. Wild boars, which share the same genus and species as domestic pigs, are also an HEV reservoir. During our nationwide study of HEV infection in wild boar populations in Japan, a genotype 6 (HEV-6) strain, wbJHG_23, was isolated in Hyogo Prefecture in 2023. The genomic length was 7244 nucleotides, excluding the poly(A) tract. The wbJHG_23 strain exhibited the highest nucleotide identity throughout its genome with two previously reported HEV-6 strains (80.3-80.9%). Conversely, it displayed lower similarity (73.3-78.1%) with the HEV-1-5, HEV-7, and HEV-8 strains, indicating that, although closely related, the wbJHG_23 strain differs significantly from the reported HEV-6 strains and might represent a novel subtype. The wbJHG_23 strain successfully infected the human-derived cancer cell lines, PLC/PRF/5 and A549 1-1H8 cells, suggesting that HEV-6 has the potential for zoonotic infection. An infectious cDNA clone was constructed using a reverse genetics system, and a cell culture system supporting the efficient propagation of the HEV-6 strain was established, providing important tools for further studies on this genotype. Using this cell culture system, we evaluated the sensitivity of the wbJHG_23 strain to ribavirin treatment. Its good response to this treatment suggested that it could be used to treat human infections caused by HEV-6.
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Affiliation(s)
- Putu Prathiwi Primadharsini
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
| | - Masaharu Takahashi
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
| | - Tsutomu Nishizawa
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
| | - Yukihiro Sato
- Department of Internal Medicine, Kamiichi General Hospital, Nakaniikawa-Gun, Toyama 930-0391, Japan;
| | - Shigeo Nagashima
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
| | - Kazumoto Murata
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
| | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0498, Japan; (P.P.P.); (M.T.); (T.N.); (S.N.); (K.M.)
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Sheng Y, Deng Y, Li X, Ji P, Sun X, Liu B, Zhu J, Zhao J, Nan Y, Zhou EM, Hiscox JA, Stewart JP, Sun Y, Zhao Q. Hepatitis E virus ORF3 protein hijacking thioredoxin domain-containing protein 5 (TXNDC5) for its stability to promote viral particle release. J Virol 2024; 98:e0164923. [PMID: 38548704 PMCID: PMC11019958 DOI: 10.1128/jvi.01649-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2024] [Accepted: 03/06/2024] [Indexed: 04/17/2024] Open
Abstract
Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis worldwide, responsible for approximately 20 million infections annually. Among the three open reading frames (ORFs) of the HEV genome, the ORF3 protein is involved in virus release. However, the host proteins involved in HEV release need to be clarified. In this study, a host protein, thioredoxin domain-containing protein 5 (TXNDC5), interacted with the non-palmitoylated ORF3 protein by co-immunoprecipitation analysis. We determined that the overexpression or knockdown of TXNDC5 positively regulated HEV release from the host cells. The 17FCL19 mutation of the ORF3 protein lost the ability to interact with TXNDC5. The releasing amounts of HEV with the ORF3 mutation (FCL17-19SSP) were decreased compared with wild-type HEV. The overexpression of TXNDC5 can stabilize and increase ORF3 protein amounts, but not the TXNDC5 mutant with amino acids 1-88 deletion. Meanwhile, we determined that the function of TXNDC5 on the stabilization of ORF3 protein is independent of the Trx-like domains. Knockdown of TXNDC5 could lead to the degradation of ORF3 protein by the endoplasmic reticulum (ER)-associated protein degradation-proteasome system. However, the ORF3 protein cannot be degraded in the knockout-TXNDC5 stable cells, suggesting that it may hijack other proteins for its stabilization. Subsequently, we found that the other members of protein disulfide isomerase (PDI), including PDIA1, PDIA3, PDIA4, and PDIA6, can increase ORF3 protein amounts, and PDIA3 and PDIA6 interact with ORF3 protein. Collectively, our study suggested that HEV ORF3 protein can utilize TXNDC5 for its stability in ER to facilitate viral release. IMPORTANCE Hepatitis E virus (HEV) infection is the leading cause of acute viral hepatitis worldwide. After the synthesis and modification in the cells, the mature ORF3 protein is essential for HEV release. However, the host protein involved in this process has yet to be determined. Here, we reported a novel host protein, thioredoxin domain-containing protein 5 (TXNDC5), as a chaperone, contributing to HEV release by facilitating ORF3 protein stability in the endoplasmic reticulum through interacting with non-palmitoylated ORF3 protein. However, we also found that in the knockout-TXNDC5 stable cell lines, the HEV ORF3 protein may hijack other proteins for its stabilization. For the first time, our study demonstrated the involvement of TXNDC5 in viral particle release. These findings provide some new insights into the process of the HEV life cycle, the interaction between HEV and host factors, and a new direction for antiviral design.
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Affiliation(s)
- Yamin Sheng
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Yingying Deng
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Xiaoxuan Li
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Pinpin Ji
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Xuwen Sun
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Baoyuan Liu
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Jiahong Zhu
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Jiakai Zhao
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Yuchen Nan
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - En-Min Zhou
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Julian A. Hiscox
- Department of Infection Biology and Microbiomes, Institute of Infection, Veterinary and Ecological Sciences, University of Liverpool, Liverpool, United Kingdom
| | - James P. Stewart
- Department of Infection Biology and Microbiomes, Institute of Infection, Veterinary and Ecological Sciences, University of Liverpool, Liverpool, United Kingdom
| | - Yani Sun
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Qin Zhao
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
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16
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Csernalabics B, Marinescu MS, Maurer L, Kelsch L, Werner J, Baumann K, Zoldan K, Panning M, Reuken P, Bruns T, Bengsch B, Neumann-Haefelin C, Hofmann M, Thimme R, Dao Thi VL, Boettler T. Efficient formation and maintenance of humoral and CD4 T-cell immunity targeting the viral capsid in acute-resolving hepatitis E infection. J Hepatol 2024; 80:564-575. [PMID: 38154741 DOI: 10.1016/j.jhep.2023.12.016] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/04/2023] [Revised: 12/06/2023] [Accepted: 12/11/2023] [Indexed: 12/30/2023]
Abstract
BACKGROUND & AIMS CD4 T cells shape the neutralizing antibody (nAb) response and facilitate viral clearance in various infections. Knowledge of their phenotype, specificity and dynamics in hepatitis E virus (HEV) infection is limited. HEV is enterically transmitted as a naked virus (nHEV) but acquires a host-derived quasi-envelope (eHEV) when budding from cells. While nHEV is composed of the open reading frame (ORF)-2-derived capsid, eHEV particles also contain ORF3-derived proteins. We aimed to longitudinally characterize the HEV-specific CD4 T cells targeting ORF1, 2 and 3 and antibodies against nHEV or eHEV in immunocompetent individuals with acute and resolved HEV infection. METHODS HEV-specific CD4 T cells were analyzed by intracellular cytokine staining after stimulation with in silico-predicted ORF1- and ORF2-derived epitopes and overlapping peptides spanning the ORF3 region. Ex vivo multiparametric characterization of capsid-specific CD4 T cells was performed using customized MHC class II tetramers. Total and neutralizing antibodies targeting nHEV or eHEV particles were determined. RESULTS HEV-specific CD4 T-cell frequencies and antibody titers are highest in individuals with acute infection and decline in a time-dependent process with an antigen hierarchy. HEV-specific CD4 T cells strongly target the ORF2-derived capsid and ORF3-specific CD4 T cells are hardly detectable. NAbs targeting nHEV are found in high titers while eHEV particles are less efficiently neutralized. Capsid-specific CD4 T cells undergo memory formation and stepwise contraction, accompanied by dynamic phenotypical and transcriptional changes over time. CONCLUSION The viral capsid is the main target of HEV-specific CD4 T cells and antibodies in acute-resolving infection, correlating with efficient neutralization of nHEV. Capsid-specific immunity rapidly emerges followed by a stepwise contraction several years after infection. IMPACT AND IMPLICATIONS The interplay of CD4 T cells and neutralizing antibody responses is critical in the host defense against viral infections, yet little is known about their characteristics in hepatitis E virus (HEV) infection. We conducted a longitudinal study of immunocompetent individuals with acute and resolved HEV infection to understand the characteristics of HEV-specific CD4 T cells and neutralizing antibodies targeting different viral proteins and particles. We found that HEV-specific CD4 T cells mainly target capsid-derived epitopes. This correlates with efficient neutralization of naked virions while quasi-enveloped particles are less susceptible to neutralization. As individuals with pre-existing liver disease and immunocompromised individuals are at risk for fulminant or chronic courses of HEV infection, these individuals might benefit from the development of vaccination strategies which require a detailed knowledge of the composition and longevity of HEV-specific CD4 T-cell and antibody immunity.
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Affiliation(s)
- Benedikt Csernalabics
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Mircea Stefan Marinescu
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Lars Maurer
- Schaller Research Group, Department of Infectious Diseases and Virology, Heidelberg University Hospital, Germany
| | - Lara Kelsch
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Jill Werner
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Katharina Baumann
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Katharina Zoldan
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Marcus Panning
- Institute of Virology, University Hospital Freiburg, Germany
| | - Philipp Reuken
- Department of Internal Medicine IV, University Hospital Jena, Germany
| | - Tony Bruns
- Department of Internal Medicine IV, University Hospital Jena, Germany; Department of Internal Medicine III, University Hospital RWTH Aachen, Germany
| | - Bertram Bengsch
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany; Signalling Research Centres BIOSS and CIBSS, University of Freiburg, Freiburg, Germany
| | - Christoph Neumann-Haefelin
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Maike Hofmann
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Robert Thimme
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany
| | - Viet Loan Dao Thi
- Schaller Research Group, Department of Infectious Diseases and Virology, Heidelberg University Hospital, Germany; German Centre for Infection Research (DZIF), Partner Site Heidelberg, Heidelberg, Germany
| | - Tobias Boettler
- Department of Medicine II, Medical Center - University of Freiburg, Germany; Faculty of Medicine, University of Freiburg, Germany.
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17
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Lu Q, Wu H, Meng J, Wang J, Wu J, Liu S, Tong J, Nie J, Huang W. Multi-epitope vaccine design for hepatitis E virus based on protein ORF2 and ORF3. Front Microbiol 2024; 15:1372069. [PMID: 38577684 PMCID: PMC10991829 DOI: 10.3389/fmicb.2024.1372069] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2024] [Accepted: 03/07/2024] [Indexed: 04/06/2024] Open
Abstract
Introduction Hepatitis E virus (HEV), with heightened virulence in immunocompromised individuals and pregnant women, is a pervasive threat in developing countries. A globaly available vaccine against HEV is currently lacking. Methods We designed a multi-epitope vaccine based on protein ORF2 and ORF3 of HEV using immunoinformatics. Results The vaccine comprised 23 nontoxic, nonallergenic, soluble peptides. The stability of the docked peptide vaccine-TLR3 complex was validated by molecular dynamic simulations. The induction of effective cellular and humoral immune responses by the multi-peptide vaccine was verified by simulated immunization. Discussion These findings provide a foundation for future HEV vaccine studies.
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Affiliation(s)
- Qiong Lu
- Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Product Control, National Institutes for Food and Drug Control and WHO Collaborating Center for Standardization and Evaluation of Biologicals, Beijing, China
| | - Hao Wu
- Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Product Control, National Institutes for Food and Drug Control and WHO Collaborating Center for Standardization and Evaluation of Biologicals, Beijing, China
- Wuhan Institute of Biological Products Co., Ltd., Wuhan, China
| | - Jing Meng
- State Key Laboratory of Common Mechanism Research for Major Diseases, Suzhou Institute of Systems Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Suzhou, Jiangsu, China
| | | | - Jiajing Wu
- Research and Development Department, Beijing Yunling Biotechnology Co., Ltd., Beijing, China
| | - Shuo Liu
- Changping Laboratory, Beijing, China
- Graduate School of Peking Union Medical College, Beijing, China
| | - Jincheng Tong
- Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Product Control, National Institutes for Food and Drug Control and WHO Collaborating Center for Standardization and Evaluation of Biologicals, Beijing, China
| | - Jianhui Nie
- Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Product Control, National Institutes for Food and Drug Control and WHO Collaborating Center for Standardization and Evaluation of Biologicals, Beijing, China
| | - Weijin Huang
- Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Product Control, National Institutes for Food and Drug Control and WHO Collaborating Center for Standardization and Evaluation of Biologicals, Beijing, China
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18
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Tanaka A, Matsubayashi K, Odajima T, Sakata H, Iida J, Kai K, Goto N, Satake M. Universal nucleic acid donor screening revealed epidemiological features of hepatitis E and prevented transfusion-transmitted infection in Japan. Transfusion 2024; 64:335-347. [PMID: 38152964 DOI: 10.1111/trf.17696] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2023] [Revised: 11/29/2023] [Accepted: 11/30/2023] [Indexed: 12/29/2023]
Abstract
BACKGROUND More than 45 cases of transfusion-transmitted hepatitis E virus infection (TT-HEV) have been reported in Japan. Therefore, in 2020, universal individual donation nucleic acid amplification testing (ID-NAT) was implemented for HEV. STUDY DESIGN AND METHODS We characterized HEV NAT-positive blood donors. The number of new HEV infections and the asymptomatic infection rate were estimated using the HEV NAT-positive rate. HEV RNA quantitation, phylogenetic analysis, and antibody tests were performed, and the residual risk of TT-HEV was assessed based on the lookback study results. RESULTS A total of 5,075,100 blood donations were screened with ID-NAT during the first year of implementation, among which 2804 (0.055%; males: 0.060%, females: 0.043%) were NAT-positive with regional differences. Approximately 270,000 new HEV infection cases were estimated to occur annually in Japan, with an asymptomatic infection rate of 99.9%. The median HEV RNA concentration, excluding cases below the limit of quantification, was 205 IU/mL. Among the 1113 cases where the genotype could be determined, HEV-3 and HEV-4 accounted for 98.8% (1100) and 1.2% (13), respectively. The maximum duration of HEV viremia, including the pre- and post-ID-NAT window periods, was estimated to be 88.2 days. Within the 3 years since ID-NAT implementation, no confirmed cases of breakthrough TT-HEV were observed. DISCUSSION Multiple indigenous HEV strains are prevalent in Japan, infecting a significant number of individuals. However, since the implementation of ID-NAT, TT-HEV has been prevented due to the test's high sensitivity.
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Affiliation(s)
- Ami Tanaka
- Central Blood Institute, Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
| | - Keiji Matsubayashi
- Central Blood Institute, Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
| | - Takeshi Odajima
- Central Blood Institute, Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
| | | | - Juri Iida
- Japanese Red Cross Hokkaido Block Blood Center, Sapporo, Japan
| | - Kazuhiro Kai
- Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
| | - Naoko Goto
- Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
| | - Masahiro Satake
- Central Blood Institute, Blood Service Headquarters, Japanese Red Cross Society, Tokyo, Japan
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19
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Gremmel N, Keuling O, Eiden M, Groschup MH, Johne R, Becher P, Baechlein C. Hepatitis E virus neutralization by porcine serum antibodies. J Clin Microbiol 2023; 61:e0037323. [PMID: 37823649 PMCID: PMC10662371 DOI: 10.1128/jcm.00373-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2023] [Accepted: 08/26/2023] [Indexed: 10/13/2023] Open
Abstract
The consumption of raw or undercooked meat products poses a serious risk for human hepatitis E virus (HEV) infections. In many high-income countries, domestic pigs and wild boars represent the main animal reservoirs for HEV and are usually identified by reverse transcription-PCR and antibody enzyme-linked immunosorbent assay (ELISA). In order to characterize the humoral immune response in more detail, a cell culture-based serum neutralization assay using a culture-adapted HEV strain was established here. Measurement of neutralizing antibodies was only possible after removing the viral quasi-envelope by detergent treatment. Serum samples of 343 wild boars from Northern Germany were first analyzed for anti-HEV IgG using an in-house ELISA, resulting in 19% positive samples. Subsequently, a subset of 41 representative samples was tested with the neutralization assay, and the results correlated well with those obtained by ELISA. Not only the human HEV strain 47832c but also two porcine HEV strains were shown to be neutralized by porcine serum antibodies. Neutralizing activity was also found in samples containing both HEV-specific antibodies and HEV RNA. Testing of serum samples derived from two experimentally infected domestic pigs showed a steep increase in neutralizing activity at 24 or 51 days post infection, dependent on the used infectious dose. The developed assay can be useful for characterization of the humoral immune response after HEV infection and for assessing the efficiency of HEV vaccine candidates.
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Affiliation(s)
- Nele Gremmel
- Department of Infectious Diseases, Institute of Virology, University of Veterinary Medicine, Hannover, Germany
| | - Oliver Keuling
- Institute for Terrestrial and Aquatic Wildlife Research, University of Veterinary Medicine, Hannover, Germany
| | - Martin Eiden
- Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Greifswald, Insel Riems, Germany
| | - Martin H. Groschup
- Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Greifswald, Insel Riems, Germany
| | - Reimar Johne
- Department of Biological Safety, German Federal Institute for Risk Assessment, Berlin, Germany
| | - Paul Becher
- Department of Infectious Diseases, Institute of Virology, University of Veterinary Medicine, Hannover, Germany
| | - Christine Baechlein
- Department of Infectious Diseases, Institute of Virology, University of Veterinary Medicine, Hannover, Germany
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20
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Nagashima S, Primadharsini PP, Nishiyama T, Takahashi M, Murata K, Okamoto H. Development of a HiBiT-tagged reporter hepatitis E virus and its utility as an antiviral drug screening platform. J Virol 2023; 97:e0050823. [PMID: 37681960 PMCID: PMC10537679 DOI: 10.1128/jvi.00508-23] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2023] [Accepted: 06/15/2023] [Indexed: 09/09/2023] Open
Abstract
Previously, we developed an infectious hepatitis E virus (HEV) harboring the nanoKAZ gene in the hypervariable region of the open reading frame 1 (ORF1) of the HEV3b (JE03-1760F/P10) genome and demonstrated the usefulness for screening anti-HEV drugs that inhibit the early infection process. In the present study, we constructed another reporter HEV (HEV3b-HiBiT) by placing a minimized HiBiT tag derived from NanoLuc luciferase at the 3'-end of the viral capsid (ORF2) coding sequence. It replicated efficiently in PLC/PRF/5 cells, produced membrane-associated particles identical to those of the parental virus, and was genetically stable and infectious. The HiBiT tag was fused to both secreted ORF2s (ORF2s-HiBiT) and ORF2c capsid protein (ORF2c-HiBiT). The ORF2c-HiBiT formed membrane-associated HEV particles (eHEV3b-HiBiT). By treating these particles with digitonin, we demonstrated that the HiBiT tag was expressed on the surface of capsid and was present inside the lipid membrane. To simplify the measurement of luciferase activity and provide a more convenient screening platform, we constructed an ORF2s-defective mutant (HEV3b-HiBiT/ΔORF2s) in which the secreted ORF2s are suppressed. We used this system to evaluate the effects of introducing small interfering RNAs and treatment with an inhibitor or accelerator of exosomal release on HEV egress and demonstrated that the effects on virus release can readily be analyzed. Therefore, HEV3b-HiBiT and HEV3b-HiBiT/ΔORF2s reporters may be useful for investigating the virus life cycle and can serve as a more convenient screening platform to search for candidate drugs targeting the late stage of HEV infection such as particle formation and release. IMPORTANCE The construction of recombinant infectious viruses harboring a stable luminescence reporter gene is essential for investigations of the viral life cycle, such as viral replication and pathogenesis, and the development of novel antiviral drugs. However, it is difficult to maintain the stability of a large foreign gene inserted into the viral genome. In the present study, we successfully generated a recombinant HEV harboring the 11-amino acid HiBiT tag in the ORF2 coding region and demonstrated the infectivity, efficient virus growth, particle morphology, and genetic stability, suggesting that this recombinant HEV is useful for in vitro assays. Furthermore, this system can serve as a more convenient screening platform for anti-HEV drugs. Thus, an infectious recombinant HEV is a powerful approach not only for elucidating the molecular mechanisms of the viral life cycle but also for the screening and development of novel antiviral agents.
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Affiliation(s)
- Shigeo Nagashima
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Putu Prathiwi Primadharsini
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Takashi Nishiyama
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Masaharu Takahashi
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Kazumoto Murata
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
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21
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Primadharsini PP, Nagashima S, Nishiyama T, Okamoto H. Three Distinct Reporter Systems of Hepatitis E Virus and Their Utility as Drug Screening Platforms. Viruses 2023; 15:1989. [PMID: 37896767 PMCID: PMC10611241 DOI: 10.3390/v15101989] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2023] [Revised: 09/20/2023] [Accepted: 09/22/2023] [Indexed: 10/29/2023] Open
Abstract
The hepatitis E virus (HEV) is increasingly acknowledged as the primary cause of acute hepatitis. While most HEV infections are self-limiting, cases of chronic infection and fulminant hepatitis necessitate the administration of anti-HEV medications. However, there is a lack of specific antiviral drugs designed for HEV, and the currently available drug (ribavirin) has been associated with significant adverse effects. The development of innovative antiviral drugs involves targeting distinct steps within the viral life cycle: the early step (attachment and internalization), middle step (translation and RNA replication), and late step (virus particle formation and virion release). We recently established three HEV reporter systems, each covering one or two of these steps. Using these reporter systems, we identified various potential drug candidates that target different steps of the HEV life cycle. Through rigorous in vitro testing using our robust cell culture system with the genotype 3 HEV strain (JE03-1760F/P10), we confirmed the efficacy of these drugs, when used alone or in combination with existing anti-HEV drugs. This underscores their significance in the quest for an effective anti-HEV treatment. In the present review, we discuss the development of the three reporter systems, their applications in drug screening, and their potential to advance our understanding of the incompletely elucidated HEV life cycle.
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Affiliation(s)
- Putu Prathiwi Primadharsini
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke 329-0498, Tochigi, Japan; (P.P.P.); (S.N.)
| | - Shigeo Nagashima
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke 329-0498, Tochigi, Japan; (P.P.P.); (S.N.)
| | - Takashi Nishiyama
- Laboratory of Membrane Proteins, Research Division for Quantitative Life Sciences, Institute for Quantitative Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan;
| | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke 329-0498, Tochigi, Japan; (P.P.P.); (S.N.)
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22
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Das A, Rivera-Serrano EE, Yin X, Walker CM, Feng Z, Lemon SM. Cell entry and release of quasi-enveloped human hepatitis viruses. Nat Rev Microbiol 2023; 21:573-589. [PMID: 37185947 PMCID: PMC10127183 DOI: 10.1038/s41579-023-00889-z] [Citation(s) in RCA: 25] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 03/22/2023] [Indexed: 05/17/2023]
Abstract
Infectious hepatitis type A and type E are caused by phylogenetically distinct single-stranded, positive-sense RNA viruses that were once considered to be non-enveloped. However, studies show that both are released nonlytically from hepatocytes as 'quasi-enveloped' virions cloaked in host membranes. These virion types predominate in the blood of infected individuals and mediate virus spread within the liver. They lack virally encoded proteins on their surface and are resistant to neutralizing anti-capsid antibodies induced by infection, yet they efficiently enter cells and initiate new rounds of virus replication. In this Review, we discuss the mechanisms by which specific peptide sequences in the capsids of these quasi-enveloped virions mediate their endosomal sorting complexes required for transport (ESCRT)-dependent release from hepatocytes through multivesicular endosomes, what is known about how they enter cells, and the impact of capsid quasi-envelopment on host immunity and pathogenesis.
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Affiliation(s)
- Anshuman Das
- Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
- Lentigen Technology, Inc., Gaithersburg, MD, USA
| | - Efraín E Rivera-Serrano
- Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
- Department of Biology, Elon University, Elon, NC, USA
| | - Xin Yin
- Center for Vaccines and Immunity, The Research Institute at Nationwide Children's Hospital, Columbus, OH, USA
- State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin, China
| | - Christopher M Walker
- Center for Vaccines and Immunity, The Research Institute at Nationwide Children's Hospital, Columbus, OH, USA
- Department of Paediatrics, The Ohio State University College of Medicine, Columbus, OH, USA
| | - Zongdi Feng
- Center for Vaccines and Immunity, The Research Institute at Nationwide Children's Hospital, Columbus, OH, USA.
- Department of Paediatrics, The Ohio State University College of Medicine, Columbus, OH, USA.
| | - Stanley M Lemon
- Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
- Department of Medicine, The University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
- Department of Microbiology & Immunology, The University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
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23
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Nemes K, Persson S, Simonsson M. Hepatitis A Virus and Hepatitis E Virus as Food- and Waterborne Pathogens-Transmission Routes and Methods for Detection in Food. Viruses 2023; 15:1725. [PMID: 37632066 PMCID: PMC10457876 DOI: 10.3390/v15081725] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2023] [Revised: 08/07/2023] [Accepted: 08/09/2023] [Indexed: 08/27/2023] Open
Abstract
Foodborne viruses are an important threat to food safety and public health. Globally, there are approximately 5 million cases of acute viral hepatitis due to hepatitis A virus (HAV) and hepatitis E virus (HEV) every year. HAV is responsible for numerous food-related viral outbreaks worldwide, while HEV is an emerging pathogen with a global health burden. The reported HEV cases in Europe have increased tenfold in the last 20 years due to its zoonotic transmission through the consumption of infected meat or meat products. HEV is considered the most common cause of acute viral hepatitis worldwide currently. This review focuses on the latest findings on the foodborne transmission routes of HAV and HEV and the methods for their detection in different food matrices.
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Affiliation(s)
- Katalin Nemes
- European Union Reference Laboratory for Foodborne Viruses, Swedish Food Agency, Dag Hammarskjölds väg 56 A, 75237 Uppsala, Sweden; (S.P.); (M.S.)
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24
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Koutsoumanis K, Allende A, Alvarez Ordoñez A, Bolton D, Bover‐Cid S, Chemaly M, Herman L, Hilbert F, Lindqvist R, Nauta M, Nonno R, Peixe L, Skandamis P, Suffredini E, Fernandez Escamez P, Gonzales‐Barron U, Roberts H, Ru G, Simmons M, Cruz RB, Lourenço Martins J, Messens W, Ortiz‐Pelaez A, Simon AC, De Cesare A. Assessment on the efficacy of methods 2 to 5 and method 7 set out in Commission Regulation (EU) No 142/2011 to inactivate relevant pathogens when producing processed animal protein of porcine origin intended to feed poultry and aquaculture animals. EFSA J 2023; 21:e08093. [PMID: 37416785 PMCID: PMC10320699 DOI: 10.2903/j.efsa.2023.8093] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 07/08/2023] Open
Abstract
An assessment was conducted on the level of inactivation of relevant pathogens that could be present in processed animal protein of porcine origin intended to feed poultry and aquaculture animals when methods 2 to 5 and method 7, as detailed in Regulation (EU) No 142/2011, are applied. Five approved scenarios were selected for method 7. Salmonella Senftenberg, Enterococcus faecalis, spores of Clostridium perfringens and parvoviruses were shortlisted as target indicators. Inactivation parameters for these indicators were extracted from extensive literature search and a recent EFSA scientific opinion. An adapted Bigelow model was fitted to retrieved data to estimate the probability that methods 2 to 5, in coincidental and consecutive modes, and the five scenarios of method 7 are able to achieve a 5 log10 and a 3 log10 reduction of bacterial indicators and parvoviruses, respectively. Spores of C. perfringens were the indicator with the lowest probability of achieving the target reduction by methods 2 to 5, in coincidental and consecutive mode, and by the five considered scenarios of method 7. An expert knowledge elicitation was conducted to estimate the certainty of achieving a 5 log10 reduction of spores of C. perfringens considering the results of the model and additional evidence. A 5 log10 reduction of C. perfringens spores was judged: 99-100% certain for methods 2 and 3 in coincidental mode; 98-100% certain for method 7 scenario 3; 80-99% certain for method 5 in coincidental mode; 66-100% certain for method 4 in coincidental mode and for method 7 scenarios 4 and 5; 25-75% certain for method 7 scenario 2; and 0-5% certain for method 7 scenario 1. Higher certainty is expected for methods 2 to 5 in consecutive mode compared to coincidental mode.
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25
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Gabrielli F, Alberti F, Russo C, Cursaro C, Seferi H, Margotti M, Andreone P. Treatment Options for Hepatitis A and E: A Non-Systematic Review. Viruses 2023; 15:1080. [PMID: 37243166 PMCID: PMC10221699 DOI: 10.3390/v15051080] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2023] [Revised: 04/27/2023] [Accepted: 04/27/2023] [Indexed: 05/28/2023] Open
Abstract
Hepatitis A and hepatitis E are relatively common causes of liver disease. Both viruses are mainly transmitted through the faecal-oral route and, consequently, most outbreaks occur in countries with poor sanitation. An important role of the immune response as the driver of liver injury is also shared by the two pathogens. For both the hepatitis A (HAV) and hepatitis E (HEV) viruses, the clinical manifestations of infection mainly consist of an acute disease with mild liver injury, which results in clinical and laboratory alterations that are self-limiting in most cases. However, severe acute disease or chronic, long-lasting manifestations may occur in vulnerable patients, such as pregnant women, immunocompromised individuals or those with pre-existing liver disease. Specifically, HAV infection rarely results in fulminant hepatitis, prolonged cholestasis, relapsing hepatitis and possibly autoimmune hepatitis triggered by the viral infection. Less common manifestations of HEV include extrahepatic disease, acute liver failure and chronic HEV infection with persistent viraemia. In this paper, we conduct a non-systematic review of the available literature to provide a comprehensive understanding of the state of the art. Treatment mainly consists of supportive measures, while the available evidence for aetiological treatment and additional agents in severe disease is limited in quantity and quality. However, several therapeutic approaches have been attempted: for HAV infection, corticosteroid therapy has shown outcome improvement, and molecules, such as AZD 1480, zinc chloride and heme oxygenase-1, have demonstrated a reduction in viral replication in vitro. As for HEV infection, therapeutic options mainly rely on the use of ribavirin, and some studies utilising pegylated interferon-alpha have shown conflicting results. While a vaccine for HAV is already available and has led to a significant reduction in the prevalence of the disease, several vaccines for HEV are currently being developed, with some already available in China, showing promising results.
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Affiliation(s)
- Filippo Gabrielli
- Postgraduate School of Internal Medicine, University of Modena and Reggio Emilia, 41126 Modena, Italy
- Department of Surgical Sciences, University of Bologna, 40126 Bologna, Italy
| | - Francesco Alberti
- Postgraduate School of Internal Medicine, University of Modena and Reggio Emilia, 41126 Modena, Italy
| | - Cristina Russo
- Postgraduate School of Internal Medicine, University of Modena and Reggio Emilia, 41126 Modena, Italy
| | - Carmela Cursaro
- Internal and Metabolic Medicine, Department of Medical and Surgical Sciences, Maternal-Infantile and Adult, AOU di Modena, University of Modena and Reggio Emilia, 41126 Modena, Italy
| | - Hajrie Seferi
- Internal and Metabolic Medicine, Department of Medical and Surgical Sciences, Maternal-Infantile and Adult, AOU di Modena, University of Modena and Reggio Emilia, 41126 Modena, Italy
| | - Marzia Margotti
- Internal and Metabolic Medicine, Department of Medical and Surgical Sciences, Maternal-Infantile and Adult, AOU di Modena, University of Modena and Reggio Emilia, 41126 Modena, Italy
| | - Pietro Andreone
- Internal and Metabolic Medicine, Department of Medical and Surgical Sciences, Maternal-Infantile and Adult, AOU di Modena, University of Modena and Reggio Emilia, 41126 Modena, Italy
- Division of Internal Medicine, Department of Medical and Surgical Sciences, Maternal-Infantile and Adult, University of Modena and Reggio Emilia, 41126 Modena, Italy
- Postgraduate School of Allergology and Clinical Immunology, University of Modena and Reggio Emilia, 41126 Modena, Italy
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26
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Primadharsini PP, Nagashima S, Tanaka T, Jirintai S, Takahashi M, Murata K, Okamoto H. Development and Characterization of Efficient Cell Culture Systems for Genotype 1 Hepatitis E Virus and Its Infectious cDNA Clone. Viruses 2023; 15:v15040845. [PMID: 37112827 PMCID: PMC10146093 DOI: 10.3390/v15040845] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2023] [Revised: 03/21/2023] [Accepted: 03/25/2023] [Indexed: 03/29/2023] Open
Abstract
Hepatitis E virus (HEV) is a major cause of acute viral hepatitis globally. Genotype 1 HEV (HEV-1) is responsible for multiple outbreaks in developing countries, causing high mortality rates in pregnant women. However, studies on HEV-1 have been hindered by its poor replication in cultured cells. The JE04-1601S strain recovered from a Japanese patient with fulminant hepatitis E who contracted HEV-1 while traveling to India was serially passaged 12 times in human cell lines. The cell-culture-generated viruses (passage 12; p12) grew efficiently in human cell lines, but the replication was not fully supported in porcine cells. A full-length cDNA clone was constructed using JE04-1601S_p12 as a template. It was able to produce an infectious virus, and viral protein expression was detectable in the transfected PLC/PRF/5 cells and culture supernatants. Consistently, HEV-1 growth was also not fully supported in the cell culture of cDNA-derived JE04-1601S_p12 progenies, potentially recapitulating the narrow tropism of HEV-1 observed in vivo. The availability of an efficient cell culture system for HEV-1 and its infectious cDNA clone will be useful for studying HEV species tropism and mechanisms underlying severe hepatitis in HEV-1-infected pregnant women as well as for discovering and developing safer treatment options for this condition.
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Affiliation(s)
- Putu Prathiwi Primadharsini
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
| | - Shigeo Nagashima
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
| | - Toshinori Tanaka
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
| | - Suljid Jirintai
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
- Division of Pathology, Department of Basic Veterinary Medicine, Inner Mongolia Agricultural University College of Veterinary Medicine, Hohhot 010018, China
| | - Masaharu Takahashi
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
| | - Kazumoto Murata
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
| | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Shimotsuke, Tochigi 329-0414, Japan
- Correspondence: ; Tel.: +81-285-58-7404
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27
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Ju X, Dong L, Ding Q. Hepatitis E Virus Life Cycle. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1417:141-157. [PMID: 37223864 DOI: 10.1007/978-981-99-1304-6_10] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/25/2023]
Abstract
Hepatitis E virus (HEV) infects over 20 million people worldwide per year, leading to 30,000-40,000 deaths. In most cases HEV infection in a self-limited, acute illness. However, chronic infections could occur in immunocompromised individuals. Due to scarcity of robust cell culture models in vitro and genetic tractable animal models in vivo, the details of HEV life cycle, as well as its interaction with host cells still remain elusive, which dampens antivirals discovery. In this chapter, we present an update in the HEV infectious cycle steps: entry, genome replication/subgenomic RNA transcription, assembly, and release. Moreover, we discussed the future prospective on HEV research and illustrates important questions urgently to be addressed.
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Affiliation(s)
- Xiaohui Ju
- School of Medicine, Tsinghua University, Beijing, China
| | - Lin Dong
- School of Medicine, Tsinghua University, Beijing, China
| | - Qiang Ding
- School of Medicine, Tsinghua University, Beijing, China.
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28
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Wang L, Wang Y, Zhuang H. Puzzles for Hepatitis E Virus. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1417:247-256. [PMID: 37223871 DOI: 10.1007/978-981-99-1304-6_17] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 05/25/2023]
Abstract
Hepatitis E virus (HEV) is an important but understudied virus that has been the major cause of acute viral hepatitis worldwide. In recent decades, our understanding of this neglected virus has changed greatly: novel forms of viral proteins and their functions have been discovered; HEV can transmit via blood transfusion and organ transplantation; HEV can infect many animal species and the number is still increasing; HEV can induce chronic hepatitis and extra-hepatic manifestations. However, we are short of effective treatment measures to counter the virus. In this chapter we tend to briefly introduce the puzzles and major knowledge gaps existed in the field of HEV research.
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Affiliation(s)
- Lin Wang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Youchun Wang
- Institute of Medical Biology, Chinese Academy of Medical Science & Peking Union Medical College, Kunming, China
| | - Hui Zhuang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
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29
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Liu X, Qi S, Yin X. Morphogenesis of Hepatitis E Virus. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1417:159-169. [PMID: 37223865 DOI: 10.1007/978-981-99-1304-6_11] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 05/25/2023]
Abstract
Hepatitis E virus, a leading cause of acute hepatitis worldwide, has been recognized as non-enveloped virus since its discovery in the 1980s. However, the recent identification of lipid membrane-associated form termed as "quasi-enveloped" HEV has changed this long-held notion. Both naked HEV and quasi-enveloped HEV play important roles in the pathogenesis of hepatitis E. However, the biogenesis and the mechanisms underlying the composition, biogenesis regulation, and functions of the novel quasi-enveloped virions remain enigmatic. In this chapter, we highlight the most recent discoveries on the dual life cycle of these two different types of virions, and further discuss the implication of the quasi-envelopment in our understanding of the molecular biology of HEV.
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Affiliation(s)
- Xing Liu
- State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, China
| | - Shuhui Qi
- State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, China
| | - Xin Yin
- State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, China.
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30
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Zhang F, Wang Y. HEV Cell Culture. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1417:119-131. [PMID: 37223862 DOI: 10.1007/978-981-99-1304-6_8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 05/25/2023]
Abstract
Cell culture is an important research method in virology. Although many attempts have been conducted to culture HEV in cells, only a few cell culture systems were considered to be efficient enough for usage. Concentration of virus stocks, host cells, and medium components affects the culture efficiency and the genetic mutations during HEV passage were found to be associated with the increased virulence in cell culture. As an alternative method for traditional cell culture, the infectious cDNA clones were constructed. The viral thermal stability, factors that impact the host range, post-translation of viral proteins, and function of different viral proteins were studied using the infectious cDNA clones. HEV cell culture studies on progeny virus showed that the viruses secreted from host cells have an envelope and its formation was associated with pORF3. This result explained the phenomenon that virus could infect host cells in the presence of anti-HEV antibodies.
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Affiliation(s)
- Feng Zhang
- Division of Therapeutical Monoclonal Antibodies, National Institutes for Food and Drug Control, Beijing, China
| | - Youchun Wang
- Institute of Medical Biology, Chinese Academy of Medical Science & Peking Union Medical College, Kunming, Yunnan Province, China.
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31
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He Q, Zhang Y, Gong W, Zeng H, Wang L. Genetic Evolution of Hepatitis E Virus. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1417:59-72. [PMID: 37223859 DOI: 10.1007/978-981-99-1304-6_5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 05/25/2023]
Abstract
Comparative analysis of the genomic sequences of multiple hepatitis E virus (HEV) isolates has revealed extensive genomic diversity among them. Recently, a variety of genetically distinct HEV variants have also been isolated and identified from large numbers of animal species, including birds, rabbits, rats, ferrets, bats, cutthroat trout, and camels, among others. Furthermore, it has been reported that recombination in HEV genomes takes place in animals and in human patients. Also, chronic HEV infection in immunocompromised individuals has revealed the presence of viral strains carrying insertions from human genes. This paper reviews current knowledge on the genomic variability and evolution of HEV.
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Affiliation(s)
- Qiyu He
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Yulin Zhang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Wanyun Gong
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Hang Zeng
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Ling Wang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China.
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Costafreda MI, Sauleda S, Rico A, Piron M, Bes M. Detection of Nonenveloped Hepatitis E Virus in Plasma of Infected Blood Donors. J Infect Dis 2022; 226:1753-1760. [PMID: 34865052 DOI: 10.1093/infdis/jiab589] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2021] [Accepted: 12/01/2021] [Indexed: 12/16/2022] Open
Abstract
BACKGROUND Transfusion-transmitted hepatitis E virus (HEV) infections have raised many concerns regarding the safety of blood products. To date, enveloped HEV particles have been described in circulating blood, whereas nonenveloped HEV virions have only been found in feces; however, no exhaustive studies have been performed to fully characterize HEV particles in blood. METHODS Using isopycnic ultracentrifugation, we determined the types of HEV particles in plasma of HEV-infected blood donors. RESULTS Nonenveloped HEV was detected in 8 of 23 plasma samples, whereas enveloped HEV was found in all of them. No association was observed between the presence of nonenveloped HEV and viral load, gender, or age at infection. However, samples with HEV-positive serology and/or increased levels of liver injury markers contained a higher proportion of nonenveloped HEV than samples with HEV-negative serology and normal levels of liver enzymes. These results were further confirmed by analyzing paired donation and follow-up samples of 10 HEV-infected donors who were HEV seronegative at donation but had anti-HEV antibodies and/or increased levels of liver enzymes at follow up. CONCLUSIONS The HEV-contaminated blood products may contain nonenveloped HEV, which may pose an additional risk to blood safety by behaving differently to pathogen inactivation treatments or increasing infectivity.
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Affiliation(s)
- Maria Isabel Costafreda
- Catalan Blood Bank (Banc de Sang i Teixits de Catalunya), Transfusion Safety Laboratory, Barcelona, Spain.,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, Instituto de Salud Carlos III, Madrid, Spain.,Vall d'Hebron Institute of Research, Vall d'Hebron Universitary Hospital, Barcelona, Spain
| | - Silvia Sauleda
- Catalan Blood Bank (Banc de Sang i Teixits de Catalunya), Transfusion Safety Laboratory, Barcelona, Spain.,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, Instituto de Salud Carlos III, Madrid, Spain.,Vall d'Hebron Institute of Research, Vall d'Hebron Universitary Hospital, Barcelona, Spain
| | - Angie Rico
- Catalan Blood Bank (Banc de Sang i Teixits de Catalunya), Transfusion Safety Laboratory, Barcelona, Spain
| | - Maria Piron
- Catalan Blood Bank (Banc de Sang i Teixits de Catalunya), Transfusion Safety Laboratory, Barcelona, Spain.,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, Instituto de Salud Carlos III, Madrid, Spain.,Vall d'Hebron Institute of Research, Vall d'Hebron Universitary Hospital, Barcelona, Spain
| | - Marta Bes
- Catalan Blood Bank (Banc de Sang i Teixits de Catalunya), Transfusion Safety Laboratory, Barcelona, Spain.,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, Instituto de Salud Carlos III, Madrid, Spain.,Vall d'Hebron Institute of Research, Vall d'Hebron Universitary Hospital, Barcelona, Spain
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Primadharsini PP, Nagashima S, Takahashi M, Murata K, Okamoto H. Ritonavir Blocks Hepatitis E Virus Internalization and Clears Hepatitis E Virus In Vitro with Ribavirin. Viruses 2022; 14:v14112440. [PMID: 36366538 PMCID: PMC9697947 DOI: 10.3390/v14112440] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2022] [Revised: 10/29/2022] [Accepted: 10/31/2022] [Indexed: 11/06/2022] Open
Abstract
Hepatitis E virus (HEV) is increasingly recognized as the leading cause of acute hepatitis. Although HEV infections are mostly self-limiting, a chronic course can develop especially in those with immunocompromised state. Ribavirin is currently used to treat such patients. According to various reports on chronic HEV infections, a sustained virological response (SVR) was achieved in approximately 80% of patients receiving ribavirin monotherapy. To increase the SVR rate, drug combination might be a viable strategy, which we attempted in the current study. Ritonavir was identified in our previous drug screening while searching for candidate novel anti-HEV drugs. It demonstrated potent inhibition of HEV growth in cultured cells. In the present study, ritonavir blocked HEV internalization as shown through time-of-addition and immunofluorescence assays. Its combination with ribavirin significantly increased the efficiency of inhibiting HEV growth compared to that shown by ribavirin monotherapy, even in PLC/PRF/5 cells with robust HEV production, and resulted in viral clearance. Similar efficiency was seen for HEV genotypes 3 and 4, the main causes of chronic infection. The present findings provide insight concerning the advantage of combination therapy using drugs blocking different steps in the HEV life cycle (internalization and RNA replication) as a potential novel treatment strategy for chronic hepatitis E.
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Presence of Intact Hepatitis B Virions in Exosomes. Cell Mol Gastroenterol Hepatol 2022; 15:237-259. [PMID: 36184032 PMCID: PMC9676402 DOI: 10.1016/j.jcmgh.2022.09.012] [Citation(s) in RCA: 30] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/20/2022] [Revised: 09/23/2022] [Accepted: 09/23/2022] [Indexed: 02/21/2023]
Abstract
BACKGROUND & AIMS Hepatitis B virus (HBV) was identified as an enveloped DNA virus with a diameter of 42 nm. Multivesicular bodies play a central role in HBV egress and exosome biogenesis. In light of this, it was studied whether intact virions wrapped in exosomes are released by HBV-producing cells. METHODS Robust methods for efficient separation of exosomes from virions were established. Exosomes were subjected to limited detergent treatment for release of viral particles. Electron microscopy of immunogold labeled ultrathin sections of purified exosomes was performed for characterization of exosomal HBV. Exosome formation/release was affected by inhibitors or Crispr/Cas-mediated gene silencing. Infectivity/uptake of exosomal HBV was investigated in susceptible and non-susceptible cells. RESULTS Exosomes could be isolated from supernatants of HBV-producing cells, which are characterized by the presence of exosomal and HBV markers. These exosomal fractions could be separated from the fractions containing free virions. Limited detergent treatment of exosomes causes stepwise release of intact HBV virions and naked capsids. Inhibition of exosome morphogenesis impairs the release of exosome-wrapped HBV. Electron microscopy confirmed the presence of intact virions in exosomes. Moreover, the presence of large hepatitis B virus surface antigen on the surface of exosomes derived from HBV expressing cells was observed, which conferred exosome-encapsulated HBV initiating infection in susceptible cells in a , large hepatitis B virus surface antigen/Na+-taurocholate co-transporting polypeptide-dependent manner. The uptake of exosomal HBV with low efficiency was also observed in non-permissive cells. CONCLUSION These data indicate that a fraction of intact HBV virions can be released as exosomes. This reveals a so far not described release pathway for HBV.
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van der Grein SG, Defourny KAY, Rabouw HH, Goerdayal SS, van Herwijnen MJC, Wubbolts RW, Altelaar M, van Kuppeveld FJM, Nolte-'t Hoen ENM. The encephalomyocarditis virus Leader promotes the release of virions inside extracellular vesicles via the induction of secretory autophagy. Nat Commun 2022; 13:3625. [PMID: 35750662 PMCID: PMC9232559 DOI: 10.1038/s41467-022-31181-y] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2021] [Accepted: 06/07/2022] [Indexed: 11/08/2022] Open
Abstract
Naked viruses can escape host cells before the induction of lysis via release in extracellular vesicles (EVs). These nanosized EVs cloak the secreted virus particles in a host-derived membrane, which alters virus-host interactions that affect infection efficiency and antiviral immunity. Currently, little is known about the viral and host factors regulating this form of virus release. Here, we assessed the role of the encephalomyocarditis virus (EMCV) Leader protein, a 'viral security protein' that subverts the host antiviral response. EV release upon infection with wildtype virus or a Leader-deficient mutant was characterized at the single particle level using high-resolution flow cytometry. Inactivation of the Leader abolished EV induction during infection and strongly reduced EV-enclosed virus release. We demonstrate that the Leader promotes the release of virions within EVs by stimulating a secretory arm of autophagy. This newly discovered role of the EMCV Leader adds to the variety of mechanisms via which this protein affects virus-host interactions. Moreover, these data provide first evidence for a crucial role of a non-structural viral protein in the non-lytic release of picornaviruses via packaging in EVs.
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Affiliation(s)
- Susanne G van der Grein
- Division of Cell Biology, Metabolism & Cancer, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 2, 3584 CM, Utrecht, The Netherlands
| | - Kyra A Y Defourny
- Division of Cell Biology, Metabolism & Cancer, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 2, 3584 CM, Utrecht, The Netherlands
| | - Huib H Rabouw
- Virology Section, Division Infectious Diseases & Immunology, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht, The Netherlands
| | - Soenita S Goerdayal
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands
| | - Martijn J C van Herwijnen
- Division of Cell Biology, Metabolism & Cancer, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 2, 3584 CM, Utrecht, The Netherlands
| | - Richard W Wubbolts
- Division of Cell Biology, Metabolism & Cancer, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 2, 3584 CM, Utrecht, The Netherlands
| | - Maarten Altelaar
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands
| | - Frank J M van Kuppeveld
- Virology Section, Division Infectious Diseases & Immunology, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht, The Netherlands
| | - Esther N M Nolte-'t Hoen
- Division of Cell Biology, Metabolism & Cancer, Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 2, 3584 CM, Utrecht, The Netherlands.
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Chew N, Situ J, Wu S, Yao W, Sridhar S. Independent Evaluation of Cell Culture Systems for Hepatitis E Virus. Viruses 2022; 14:v14061254. [PMID: 35746725 PMCID: PMC9227121 DOI: 10.3390/v14061254] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2022] [Revised: 06/01/2022] [Accepted: 06/07/2022] [Indexed: 02/04/2023] Open
Abstract
Hepatitis E virus (HEV) infection in humans is primarily caused by genotypes within Paslahepevirus species balayani (HEV-A). Rocahepevirus species ratti (HEV-C1, otherwise known as rat HEV) can also infect humans. HEV grows poorly in cell culture. Recent studies have reported that hyper-confluent cell layers, amphotericin B, MgCl2, progesterone, and dimethyl sulfoxide (DMSO) increase HEV yield in vitro. Here, we describe an independent evaluation of the effectiveness of these modifications in improving the yield of HEV-A genotype 4 (HEV-A4) and HEV-C1 from clinical samples in PLC/PRF/5 cells. We found that amphotericin B, MgCl2, and DMSO increased HEV yield from high-viral-load patient stool samples, while progesterone was not effective. Yield of HEV-C1 was lower than HEV-A4 across all medium conditions, but was boosted by DMSO. HEV-A4 could be maintained for over 18 months in amphotericin B- and MgCl2-containing medium, with the demonstration of viral antigen in supernatants and infected cells. We also evaluated various protocols to remove pseudo-envelopes from cell culture-derived HEV. Treating cell culture supernatant with NP-40 was the most effective. Our findings identify key modifications that boost HEV growth in vitro and illustrate the importance of independent verification of such studies using diverse HEV variants and cell lines.
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Affiliation(s)
- Nicholas Chew
- Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; (N.C.); (J.S.); (S.W.); (W.Y.)
| | - Jianwen Situ
- Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; (N.C.); (J.S.); (S.W.); (W.Y.)
| | - Shusheng Wu
- Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; (N.C.); (J.S.); (S.W.); (W.Y.)
| | - Weiming Yao
- Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; (N.C.); (J.S.); (S.W.); (W.Y.)
| | - Siddharth Sridhar
- Department of Microbiology, School of Clinical Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; (N.C.); (J.S.); (S.W.); (W.Y.)
- State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong, China
- Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong, China
- Correspondence: ; Tel.: +852-22552408
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Behrendt P, Friesland M, Wißmann JE, Kinast V, Stahl Y, Praditya D, Hueffner L, Nörenberg PM, Bremer B, Maasoumy B, Steinmann J, Becker B, Paulmann D, Brill FHH, Steinmann J, Ulrich RG, Brüggemann Y, Wedemeyer H, Todt D, Steinmann E. Hepatitis E virus is highly resistant to alcohol-based disinfectants. J Hepatol 2022; 76:1062-1069. [PMID: 35085595 DOI: 10.1016/j.jhep.2022.01.006] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/14/2021] [Revised: 12/23/2021] [Accepted: 01/11/2022] [Indexed: 12/26/2022]
Abstract
BACKGROUND & AIMS Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis worldwide and is mainly transmitted via the fecal-oral route or through consumption of contaminated food products. Due to the lack of efficient cell culture systems for the propagation of HEV, limited data regarding its sensitivity to chemical disinfectants are available. Consequently, preventive and evidence-based hygienic guidelines on HEV disinfection are lacking. METHODS We used a robust HEV genotype 3 cell culture model which enables quantification of viral infection of quasi-enveloped and naked HEV particles. For HEV genotype 1 infections, we used the primary isolate Sar55 in a fecal suspension. Standardized quantitative suspension tests using end point dilution and large-volume plating were performed for the determination of virucidal activity of alcohols (1-propanol, 2-propanol, ethanol), WHO disinfectant formulations and 5 different commercial hand disinfectants against HEV. Iodixanol gradients were conducted to elucidate the influence of ethanol on quasi-enveloped viral particles. RESULTS Naked and quasi-enveloped HEV was resistant to alcohols as well as alcohol-based formulations recommended by the WHO. Of the tested commercial hand disinfectants only 1 product displayed virucidal activity against HEV. This activity could be linked to phosphoric acid as an essential ingredient. Finally, we observed that ethanol and possibly non-active alcohol-based disinfectants disrupt the quasi-envelope structure of HEV particles, while leaving the highly transmissible and infectious naked virions intact. CONCLUSIONS Different alcohols and alcohol-based hand disinfectants were insufficient to eliminate HEV infectivity with the exception of 1 commercial ethanol-based product that included phosphoric acid. These findings have major implications for the development of measures to reduce viral transmission in clinical practice. LAY SUMMARY Hepatitis E virus (HEV) showed a high level of resistance to alcohols and alcohol-based hand disinfectants. The addition of phosphoric acid to alcohol was essential for virucidal activity against HEV. This information should be used to guide improved hygiene measures for the prevention of HEV transmission.
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Affiliation(s)
- Patrick Behrendt
- Institute for Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, A Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany; Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Germany; German Centre for Infection Research (DZIF), Hannover-Braunschweig, Germany.
| | - Martina Friesland
- Institute for Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, A Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany
| | - Jan-Erik Wißmann
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Volker Kinast
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Yannick Stahl
- Institute for Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, A Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany
| | - Dimas Praditya
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Lucas Hueffner
- Institute for Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, A Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany
| | - Pia Maria Nörenberg
- Institute for Experimental Virology, TWINCORE Centre for Experimental and Clinical Infection Research, A Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), Hannover, Germany
| | - Birgit Bremer
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Germany
| | - Benjamin Maasoumy
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Germany; German Centre for Infection Research (DZIF), Hannover-Braunschweig, Germany; Centre for Individualised Infection Medicine (CIIM), Hannover, Germany
| | - Jochen Steinmann
- Dr. Brill + Partner GmbH Institute for Hygiene and Microbiology, Bremen, Germany
| | - Britta Becker
- Dr. Brill + Partner GmbH Institute for Hygiene and Microbiology, Bremen, Germany
| | - Dajana Paulmann
- Dr. Brill + Partner GmbH Institute for Hygiene and Microbiology, Bremen, Germany
| | - Florian H H Brill
- Dr. Brill + Partner GmbH Institute for Hygiene and Microbiology, Bremen, Germany
| | - Joerg Steinmann
- Institute of Medical Microbiology, University Hospital Essen, University of Duisburg-Essen, Germany; Institute for Clinical Hygiene, Medical Microbiology and Infectiology, Clinic Nuernberg, Paracelsus Medical University, Nuremberg, Germany
| | - Rainer G Ulrich
- Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany and German Centre for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Insel Riems, Germany
| | - Yannick Brüggemann
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany
| | - Heiner Wedemeyer
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Germany; German Centre for Infection Research (DZIF), Hannover-Braunschweig, Germany
| | - Daniel Todt
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany; European Virus Bioinformatics Center (EVBC), Jena, Germany
| | - Eike Steinmann
- Department of Molecular and Medical Virology, Ruhr University Bochum, Bochum, Germany; German Centre for Infection Research (DZIF), External Partner Site, Bochum, Germany.
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Tréguier Y, Bull-Maurer A, Roingeard P. Apolipoprotein E, a Crucial Cellular Protein in the Lifecycle of Hepatitis Viruses. Int J Mol Sci 2022; 23:ijms23073676. [PMID: 35409035 PMCID: PMC8998859 DOI: 10.3390/ijms23073676] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2022] [Revised: 03/19/2022] [Accepted: 03/25/2022] [Indexed: 02/01/2023] Open
Abstract
Apolipoprotein E (ApoE) is a multifunctional protein expressed in several tissues, including those of the liver. This lipoprotein component is responsible for maintaining lipid content homeostasis at the plasma and tissue levels by transporting lipids between the liver and peripheral tissues. The ability of ApoE to interact with host-cell surface receptors and its involvement in several cellular pathways raised questions about the hijacking of ApoE by hepatotropic viruses. Hepatitis C virus (HCV) was the first hepatitis virus reported to be dependent on ApoE for the completion of its lifecycle, with ApoE being part of the viral particle, mediating its entry into host cells and contributing to viral morphogenesis. Recent studies of the hepatitis B virus (HBV) lifecycle have revealed that this virus and its subviral envelope particles also incorporate ApoE. ApoE favors HBV entry and is crucial for the morphogenesis of infectious particles, through its interaction with HBV envelope glycoproteins. This review summarizes the data highlighting the crucial role of ApoE in the lifecycles of HBV and HCV and discusses its potential role in the lifecycle of other hepatotropic viruses.
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Affiliation(s)
- Yannick Tréguier
- INSERM U1259 MAVIVH, Université de Tours et CHU de Tours, 37032 Tours, France; (Y.T.); (A.B.-M.)
| | - Anne Bull-Maurer
- INSERM U1259 MAVIVH, Université de Tours et CHU de Tours, 37032 Tours, France; (Y.T.); (A.B.-M.)
| | - Philippe Roingeard
- INSERM U1259 MAVIVH, Université de Tours et CHU de Tours, 37032 Tours, France; (Y.T.); (A.B.-M.)
- Plateforme IBiSA des Microscopies, Université de Tours et CHU de Tours, 37032 Tours, France
- Correspondence: ; Tel.: +33-0247-366-232
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Fang L, Zhang J, Chen H, Lv F, Yu Y, Du X. Epidemiological Characteristics and Clinical Manifestations of Hepatitis E in a Tertiary Hospital in China: A Retrospective Study. Front Microbiol 2022; 12:831968. [PMID: 35310389 PMCID: PMC8928388 DOI: 10.3389/fmicb.2021.831968] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2021] [Accepted: 12/31/2021] [Indexed: 01/12/2023] Open
Abstract
Background Hepatitis E virus (HEV) infection is the most common cause of acute viral hepatitis worldwide and one of the main causes of death in the last decade, causing chronic hepatitis and liver failure in some populations. The aging population and obesity are two major factors threatening human health. Therefore, we want to understand the relationship between these two groups and HEV infection. Objectives The study aimed to analyze the epidemiological, clinical, and laboratory features of HEV infection and evaluate probable high-risk factors for disease progression and the current diagnostic strategies of hepatitis E infection. Study Design Patients diagnosed with acute hepatitis E with symptoms and liver dysfunction were enrolled. For statistical analysis, clinical features and laboratory findings were collected between the elderly and non-elderly and HEV+ fatty liver disease (FLD) groups. Statistical analysis was performed using Excel and the platform VassarStats, and statistical significance was taken as P < 0.05. Results Jaundice and the bilirubin peak were significantly deeper, the duration of hospitalization was significantly longer, and the proportion of ascites and liver failure was significantly higher in the elderly group. The aging population is one of the risk factors of severe hepatitis E. Hepatitis E becomes more serious in the HEV + FLD group, although the results did not reach statistical significance. Conclusion The aging and FLD were suggested to aggravate HEV infection. However, the diagnosis of HEV infection remains a challenge. A prospective study with sufficient sample size is needed to confirm this conclusion.
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Affiliation(s)
- Li Fang
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
| | - Junli Zhang
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
| | - Huiying Chen
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
| | - Fangfang Lv
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
| | - Yunsong Yu
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
| | - Xiaoxing Du
- Department of Infectious Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China
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Shafat Z, Ahmed A, Parvez MK, Islam A, Parveen S. Intrinsically disordered regions in the rodent hepevirus proteome. Bioinformation 2022; 18:111-118. [PMID: 36420436 PMCID: PMC9649497 DOI: 10.6026/97320630018111] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2021] [Revised: 01/03/2022] [Accepted: 01/03/2022] [Indexed: 09/19/2023] Open
Abstract
Hepatitis E virus (HEV) is the causative agent of Hepatitis E infections across the world. Intrinsically disordered protein regions (IDPRs) or intrinsically disordered proteins (IDPs) are regions or proteins that are characterized by lack of definite structure. These IDPRs or IDPs play significant roles in a wide range of biological processes, such as cell cycle regulation, control of signaling pathways, etc. IDPR/IDP in proteins is associated with the virus's pathogenicity and infectivity. The prevalence of IDPR/IDP in rat HEV proteome remains undetermined. Hence, we examined the unstructured/disordered regions of the open reading frame (ORF) encoded proteins of rat HEV by analyzing the prevalence of intrinsic disorder. The intrinsic disorder propensity analysis showed that the different ORF proteins consisted of varying fraction of intrinsic disorder. The protein ORF3 was identified with maximum propensity for intrinsic disorder while the ORF6 protein had the least fraction of intrinsic disorder. The analysis revealed ORF6 as a structured protein (ORDP); ORF1 and ORF4 as moderately disordered proteins (IDPRs); and ORF3 and ORF5 as highly disordered proteins (IDPs). The protein ORF2 was found to be moderately as well as highly disordered using different predictors, thus, was categorized into both IDPR and IDP. Such disordered regions have important roles in pathogenesis and replication of viruses.
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Affiliation(s)
- Zoya Shafat
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Anwar Ahmed
- Centre of Excellence in Biotechnology Research, College of Science, King Saud University, Riyadh, Saudi Arabia
| | - Mohammad K Parvez
- Department of Pharmacognosy, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia
| | - Asimul Islam
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Shama Parveen
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
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Development of recombinant infectious hepatitis E virus harboring the nanoKAZ gene and its application in drug screening. J Virol 2022; 96:e0190621. [PMID: 35107380 DOI: 10.1128/jvi.01906-21] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Hepatitis E virus (HEV) is a quasi-enveloped virus with a single-stranded positive-sense RNA genome belonging to family Hepeviridae. Studies on molecular aspects of HEV and drug screening have benefited from the discovery of bioluminescent reporter genes. However, the stability of large foreign genes is difficult to maintain after insertion into the viral genome. Currently, ribavirin is used to treat HEV-infected patients who require antiviral therapy. This has several major drawbacks. Thus, the development of novel anti-HEV drugs is of great importance. We developed a system consisting of recombinant infectious HEV harboring small luciferase gene (nanoKAZ) in the hypervariable region (HVR) of the open reading frame 1 (ORF1) (HEV-nanoKAZ). It replicated efficiently in cultured cells, was genetically stable, and had morphological characteristics similar to the parental virus. Both membrane-associated (eHEV-nanoKAZ) and membrane-unassociated (neHEV-nanoKAZ) particles were infectious. HEV particles circulating in the blood stream and attaching to hepatocytes in HEV-infected patients are membrane-associated, thus, eHEV-nanoKAZ was applied in drug screening. The eHEV-nanoKAZ system is able to cover at least the inhibitor of HEV entry and inhibitor of HEV RNA replication. Four drugs with anti-HEV activity were identified. Their effectiveness in cultured cells was confirmed in naïve and HEV-producing PLC/PRF/5 cells. Two hit drugs (azithromycin and ritonavir) strongly inhibited HEV production in culture supernatants, as well as intracellular expression of ORF2 protein, and may therefore be candidate novel anti-HEV drugs. The HEV-nanoKAZ system was developed and applied in drug screening, and is expected to be useful for investigating the HEV life cycle. IMPORTANCE Bioluminescent reporter viruses are essential tools in molecular virological research. It has been widely used to investigate viral life cycles and in the development of antiviral drugs. For drug screening, the use of a bioluminescent reporter virus helps shorten the time required to perform the assay. A system, consisting of recombinant infectious HEV harboring the nanoKAZ gene in the HVR of ORF1 (HEV-nanoKAZ), was developed in this study, and was successfully applied to drug screening in which four hit drugs with anti-HEV activity were identified. The results of this study provide evidence supporting the use of this system in more variable HEV studies. In addition, both forms of viral particles (eHEV-nanoKAZ and neHEV-nanoKAZ) are infectious, which will enable their application in HEV studies requiring both forms of viral particles, such as in the investigation of unknown HEV receptors and the elucidation of host factors important for HEV entry.
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Behrendt P, Wedemeyer H. [Vaccines against hepatitis E virus: state of development]. Bundesgesundheitsblatt Gesundheitsforschung Gesundheitsschutz 2022; 65:192-201. [PMID: 35099576 PMCID: PMC8802100 DOI: 10.1007/s00103-022-03487-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2021] [Accepted: 12/22/2021] [Indexed: 11/30/2022]
Abstract
In Europa ist aktuell kein Impfstoff gegen das Hepatitis-E-Virus (HEV) zugelassen. Demgegenüber steht in China bereits seit 10 Jahren mit HEV-239 (Hecolin®, Xiamen Innovax Biotech Co., Xiamen, China) ein Vakzin gegen den HEV-Genotyp 4 zur Verfügung. Herausforderungen für die Entwicklung von Impfstoffen ergeben sich v. a. aus den Unterschieden zwischen den Genotypen bezüglich Verbreitung, Übertragungswege und Risikogruppen. Weitere Hindernisse sind die Umhüllung von HEV im Blut durch Wirtsmembranen, die Replikation in verschiedenen Organen außerhalb der Leber sowie schwächere Immunantworten in vulnerablen Gruppen. In diesem Artikel wird der aktuelle Stand der verfügbaren und in fortgeschrittener präklinischer Evaluation befindlichen Vakzine gegen HEV mit Fokus auf Strategien der Impfstoffentwicklung dargestellt. Herausforderungen und Limitationen werden beschrieben. Aktuelle Impfkandidaten fokussieren auf proteinbasierte Immunisierungen mit dem Ziel der Induktion von schützenden, neutralisierenden Antikörperantworten. Das Ziel der HEV-239-Zulassungsstudie mit mehr als 100.000 Studienteilnehmern war die Verhinderung von akuten symptomatischen Infektionen. Es ist jedoch unklar, inwieweit asymptomatische Infektionen durch das Vakzin verhindert wurden und ob es in Risikopatienten für einen komplizierten Verlauf, wie Patienten mit Leberzirrhose, Immunsupprimierten und Schwangeren, effektiv genug wirkt. Effiziente In-vitro-Modelle ermöglichen zunehmend die Entwicklung von monoklonalen neutralisierenden Antikörpern zur passiven Immunisierung oder Therapie. Zukünftige Vakzine sollten neben einem sehr guten Sicherheitsprofil eine eindeutige Protektion gegenüber allen Genotypen demonstrieren. Die Entwicklung einer effizienten passiven Immunisierungsstrategie, insbesondere für immunsupprimierte Personen, ist wünschenswert.
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Affiliation(s)
- Patrick Behrendt
- Klinik für Gastroenterologie, Hepatologie und Endokrinologie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625, Hannover, Deutschland
| | - Heiner Wedemeyer
- Klinik für Gastroenterologie, Hepatologie und Endokrinologie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625, Hannover, Deutschland.
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Shafat Z, Ahmed A, Parvez MK, Parveen S. Sequence to structural analysis of ORF5 protein in Norway rat Hepatitis E Virus. Bioinformation 2022; 18:19-25. [PMID: 35815200 PMCID: PMC9200610 DOI: 10.6026/97320630018019] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2021] [Revised: 11/14/2021] [Accepted: 11/14/2021] [Indexed: 11/23/2022] Open
Abstract
Hepatitis E virus (HEV) is a major causative agent of acute hepatitis in developing countries. The Norway rat HEV genome consists of six open reading frames (ORFs), i.e., ORF1, ORF2, ORF3, ORF4, ORF5 and ORF6. The additional reading frame encoded protein ORF5 is attributed to life cycle of rat HEV. The ORFF5 protein's function remains undetermined. Therefore, it is of interest to analyze the ORF5 protein for its physiochemical properties, primary structure, secondary structure, tertiary structure and functional characteristics using bioinformatics tools. Analysis of the ORF5 protein revealed it as highly unstable, hydrophilic with basic pI. The ORF5 protein consisted mostly of Arg, Pro, Ser, Leu and Gly. The 3D structural homology model of the ORF5 protein generated showed mixed α/β structural fold with predominance of coils. Structural analysis revealed the presence of clefts, pores and a tunnel. This data will help in the sequence, structure and functional annotation of ORF5.
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Affiliation(s)
- Zoya Shafat
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Anwar Ahmed
- Centre of Excellence in Biotechnology Research, College of Science, King Saud University, Riyadh, Saudi Arabia
| | - Mohammad K Parvez
- Department of Pharmacognosy, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia
| | - Shama Parveen
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
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Induction of Hepatitis E Virus Anti-ORF3 Antibodies from Systemic Administration of a Muscle-Specific Adeno-Associated Virus (AAV) Vector. Viruses 2022; 14:v14020266. [PMID: 35215859 PMCID: PMC8878420 DOI: 10.3390/v14020266] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2021] [Revised: 01/23/2022] [Accepted: 01/24/2022] [Indexed: 01/25/2023] Open
Abstract
The hepatitis E virus (HEV) is a major global health problem, leading to large outbreaks in the developing world and chronic infections in the developed world. HEV is a non-enveloped virus, which circulates in the blood in a quasi-enveloped form. The quasi-envelope protects HEV particles from neutralising anti-capsid antibodies in the serum; however, most vaccine approaches are designed to induce an immune response against the HEV capsid. In this study, we explored systemic in vivo administration of a novel synthetic and myotropic Adeno-associated virus vector (AAVMYO3) to express the small HEV phosphoprotein ORF3 (found on quasi-enveloped HEV) in the musculature of mice, resulting in the robust and dose-dependent formation of anti-ORF3 antibodies. Neutralisation assays using the serum of ORF3 AAV-transduced mice showed a modest inhibitory effect on the infection of quasi-enveloped HEV in vivo, comparable to previously characterised anti-ORF3 antibodies used as a control. The novel AAVMYO3 capsid used in this study can serve as a versatile platform for the continued development of vector-based vaccines against HEV and other infectious agents, which could complement traditional vaccines akin to the current positive experience with SARS-CoV-2.
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Cheung CKM, Wong SH, Law AWH, Law MF. Transfusion-transmitted hepatitis E: What we know so far? World J Gastroenterol 2022; 28:47-75. [PMID: 35125819 PMCID: PMC8793017 DOI: 10.3748/wjg.v28.i1.47] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/10/2021] [Revised: 07/16/2021] [Accepted: 12/22/2021] [Indexed: 02/06/2023] Open
Abstract
Hepatitis E virus (HEV) is a major cause of viral hepatitis globally. There is growing concern about transfusion-transmitted HEV (TT-HEV) as an emerging global health problem. HEV can potentially result in chronic infection in immunocompromised patients, leading to a higher risk of liver cirrhosis and even death. Between 0.0013% and 0.281% of asymptomatic blood donors around the world have HEV viremia, and 0.27% to 60.5% have anti-HEV immunoglobulin G. HEV is infectious even at very low blood concentrations of the virus. Immunosuppressed patients who develop persistent hepatitis E infection should have their immunosuppressant regimen reduced; ribavirin may be considered as treatment. Pegylated interferon can be considered in those who are refractory or intolerant to ribavirin. Sofosbuvir, a nucleotide analog, showed modest antiviral activity in some clinical studies but sustained viral response was not achieved. Therefore, rescue treatment remains an unmet need. The need for HEV screening of all blood donations remains controversial. Universal screening has been adopted in some countries after consideration of risk and resource availability. Various pathogen reduction methods have also been proposed to reduce the risk of TT-HEV. Future studies are needed to define the incidence of transmission through transfusion, their clinical features, outcomes and prognosis.
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Affiliation(s)
| | - Sunny Hei Wong
- Institute of Digestive Disease and Department of Medicine and Therapeutics, the Chinese University of Hong Kong, Hong Kong 852, China
- Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore 639798, Singapore
| | | | - Man Fai Law
- Medicine and Therapeutics, Prince of Wales Hospital, Hong Kong 852, China
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Gordeychuk I, Kyuregyan K, Kondrashova A, Bayurova E, Gulyaev S, Gulyaeva T, Potemkin I, Karlsen A, Isaeva O, Belyakova A, Lyashenko A, Sorokin A, Chumakov A, Morozov I, Isaguliants M, Ishmukhametov A, Mikhailov M. Immunization with recombinant ORF2 p551 protein protects common marmosets (Callithrix jacchus) against homologous and heterologous hepatitis E virus challenge. Vaccine 2022; 40:89-99. [PMID: 34836660 DOI: 10.1016/j.vaccine.2021.11.042] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2021] [Revised: 10/18/2021] [Accepted: 11/14/2021] [Indexed: 12/23/2022]
Abstract
BACKGROUND Hepatitis E virus (HEV) is a major causative agent of acute hepatitis worldwide, prompting continuous HEV vaccine efforts. Vaccine development is hampered by the lack of convenient animal models susceptible to infection with different HEV genotypes. We produced recombinant open reading frame 2 protein (pORF2; p551) of HEV genotype (GT) 3 and assessed its immunogenicity and protectivity against HEV challenge in common marmosets (Callithrix jacchus, CM). METHODS p551 with consensus sequence corresponding to amino acid residues 110-660 of HEV GT3 pORF2 was expressed in E. coli and purified by affinity chromatography. CMs were immunized intramuscularly with 20 μg of p551 VLPs with alum adjuvant (n = 4) or adjuvant alone (n = 2) at weeks 0, 3, 7 and 19. At week 27, p551-immunized and control animals were challenged with HEV GT1 or GT3 and thereafter longitudinally screened for markers of liver function, anti-HEV IgG and HEV RNA in feces and sera. RESULTS Purified p551 formed VLPs with particle size of 27.71 ± 2.42 nm. Two immunizations with p551 induced anti-HEV IgG mean titer of 1:1810. Immunized CMs challenged with homologous and heterologous HEV genotype did not develop HEV infection during the follow-up. Control CMs infected with both HEV GT1 and GT3 demonstrated signs of HEV infection with virus shedding and elevation of the levels of liver enzymes. High levels of anti-HEV IgG persisted in vaccinated CMs and control CMs that resolved HEV infection, for up to two years post challenge. CONCLUSIONS CMs are shown to be a convenient laboratory animal model susceptible to infection with HEV GT1 and GT3. Immunization with HEV GT3 ORF2/p551 triggers potent anti-HEV antibody response protecting CMs from homologous and heterologous HEV challenge. This advances p551 in VLPs as a prototype vaccine against HEV.
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Affiliation(s)
- Ilya Gordeychuk
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, Moscow 127994, Russia.
| | - Karen Kyuregyan
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; I.I. Mechnikov Research Institute of Vaccines and Sera, Moscow 105064, Russia; Russian Medical Academy of Continuous Professional Education, Moscow 125993, Russia.
| | - Alla Kondrashova
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, Moscow 127994, Russia
| | - Ekaterina Bayurova
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Stanislav Gulyaev
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Tatiana Gulyaeva
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Ilya Potemkin
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; I.I. Mechnikov Research Institute of Vaccines and Sera, Moscow 105064, Russia; Russian Medical Academy of Continuous Professional Education, Moscow 125993, Russia.
| | - Anastasia Karlsen
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; I.I. Mechnikov Research Institute of Vaccines and Sera, Moscow 105064, Russia; Russian Medical Academy of Continuous Professional Education, Moscow 125993, Russia; N.F. Gamaleya Federal Research Center for Epidemiology & Microbiology, Moscow 123098, Russia
| | - Olga Isaeva
- I.I. Mechnikov Research Institute of Vaccines and Sera, Moscow 105064, Russia; Russian Medical Academy of Continuous Professional Education, Moscow 125993, Russia.
| | - Alla Belyakova
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Anna Lyashenko
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Alexey Sorokin
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia
| | - Alexey Chumakov
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, Moscow 127994, Russia
| | - Igor Morozov
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia.
| | - Maria Isaguliants
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; N.F. Gamaleya Federal Research Center for Epidemiology & Microbiology, Moscow 123098, Russia; Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, 17177 Stockholm, Sweden.
| | - Aydar Ishmukhametov
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, Moscow 108819, Russia; Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, Moscow 127994, Russia.
| | - Mikhail Mikhailov
- I.I. Mechnikov Research Institute of Vaccines and Sera, Moscow 105064, Russia; Russian Medical Academy of Continuous Professional Education, Moscow 125993, Russia.
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The Capsid (ORF2) Protein of Hepatitis E Virus in Feces Is C-Terminally Truncated. Pathogens 2021; 11:pathogens11010024. [PMID: 35055972 PMCID: PMC8779013 DOI: 10.3390/pathogens11010024] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2021] [Revised: 12/21/2021] [Accepted: 12/24/2021] [Indexed: 11/17/2022] Open
Abstract
The hepatitis E virus (HEV) is a causative agent of hepatitis E. HEV virions in circulating blood and culture media are quasi-enveloped, while those in feces are nonenveloped. The capsid (ORF2) protein associated with an enveloped HEV virion is reported to comprise the translation product of leucine 14/methionine 16 to 660 (C-terminal end). However, the nature of the ORF2 protein associated with fecal HEV remains unclear. In the present study, we compared the molecular size of the ORF2 protein among fecal HEV, cell-culture-generated HEV (HEVcc), and detergent-treated protease-digested HEVcc. The ORF2 proteins associated with fecal HEV were C-terminally truncated and showed the same size as those of the detergent-treated protease-digested HEVcc virions (60 kDa), in contrast to those of the HEVcc (68 kDa). The structure prediction of the ORF2 protein (in line with previous studies) demonstrated that the C-terminal region (54 amino acids) of an ORF2 protein is in flux, suggesting that proteases target this region. The nonenveloped nondigested HEV structure prediction indicates that the C-terminal region of the ORF2 protein moves to the surface of the virion and is unnecessary for HEV infection. Our findings clarify the maturation of nonenveloped HEV and will be useful for studies on the HEV lifecycle.
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Shafat Z, Hamza A, Deeba F, Parvez MK, Parveen S. Molecular insights into the Y-domain of hepatitis E virus using computational analyses. BENI-SUEF UNIVERSITY JOURNAL OF BASIC AND APPLIED SCIENCES 2021; 10:76. [DOI: 10.1186/s43088-021-00154-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2021] [Accepted: 10/03/2021] [Indexed: 11/11/2022] Open
Abstract
Abstract
Background
Hepatitis E virus (HEV) of the family Hepeviridae is a major causative agent of acute hepatitis in developing countries. The Y-domain is derived from multi-domain non-structural polyprotein encoded by open reading frame 1 (ORF1). Previous studies have demonstrated the essentiality of Y-domain sequences in HEV life cycle; however, its function remains completely unexplored. The following study was thus conceptualized to examine the detailed computational investigation for the putative Y-domain to estimate its phylogenetic assessment, physiochemical properties, structural and functional characteristics using in silico analyses.
Results
The phylogenetic assessment of Y-domain with a vast range of hosts indicated that the protein was very well conserved throughout the course of evolution. The Y-domain was found to be unstable, hydrophilic and basic in nature with high thermostability value. Structural analysis of Y-domain revealed mixed α/β structural fold of the protein having higher percentage of alpha-helices. The three-dimensional (3D) protein model generated through homology modelling revealed the presence of clefts, tunnels and pore. Gene ontology analysis predicted Y-domain protein’s involvement in several binding and catalytic activities as well as significant biological processes. Mutations in the conserved amino acids of the Y-domain suggested that it may stabilize or de-stabilize the protein structure that might affect its structure–function relationship.
Conclusions
This theoretical study will facilitate towards deciphering the role of unexplored Y-domain, thereby providing better understanding towards the pathogenesis of HEV infection.
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Shafat Z, Hamza A, Islam A, Al-Dosari MS, Parvez MK, Parveen S. Structural exploration of Y-domain reveals its essentiality in HEV pathogenesis. Protein Expr Purif 2021; 187:105947. [PMID: 34314826 DOI: 10.1016/j.pep.2021.105947] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2021] [Revised: 07/08/2021] [Accepted: 07/21/2021] [Indexed: 12/19/2022]
Abstract
Hepatitis E virus (HEV) is a major causative agent of hepatitis E infections across the globe. Although the essentiality of HEV nonstructural polyprotein (pORF1) putative Y-domain (Yd) has been established in viral pathogenesis, its structural-functional role remains elusive. The current research discusses the novel exploration on Yd protein expression, purification, biophysical characterization and structure-based docking analysis. The codon optimized synthetic gene and optimized expression parameters i.e., 5 h induction with 0.25 mM IPTG at 37 °C, resulted in efficient production of Yd protein (~40 kDa) in E. coli BL21(DE3) cells. Majority of the recombinant Yd (rYd) protein expressed as inclusion bodies was solubilized in 0.5% N-lauroylsarcosine and purified using Ni-NTA chromatography. Circular dichroism (CD) and UV visible absorption spectroscopic studies on Yd revealed both secondary and tertiary structure stability in alkaline range (pH 8.0-10.0), suggesting correlation with its physiological activity. Thus, loss in structure at low pH perhaps play crucial role in cytoplasmic-membrane interaction. The biophysical data were in good agreement with insilico structural analyses, which suggested mixed α/β fold, non-random and basic nature of Yd protein. Furthermore, due to Yd protein essentiality in HEV replication and pathogenesis, it was considered as a template for docking and drug-likeness analyses. The 3D modeling of Yd protein and structure-based screening and drug-likeness of inhibitory compounds, including established antiviral drugs led to the identification of top nine promising candidates. Nonetheless, in vitro studies on the predicted interaction of Yd with intracellular-membrane towards establishing replication-complexes as well as validations of the proposed therapeutic agents are warranted.
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Affiliation(s)
- Zoya Shafat
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Abu Hamza
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Asimul Islam
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India
| | - Mohammed S Al-Dosari
- Department of Pharmacognosy, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia
| | - Mohammad K Parvez
- Department of Pharmacognosy, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia.
| | - Shama Parveen
- Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India.
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Lienhard J, Vonlanthen-Specker I, Sidler X, Bachofen C. Screening of Swiss Pig Herds for Hepatitis E Virus: A Pilot Study. Animals (Basel) 2021; 11:3050. [PMID: 34827782 PMCID: PMC8614339 DOI: 10.3390/ani11113050] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2021] [Revised: 10/20/2021] [Accepted: 10/21/2021] [Indexed: 12/14/2022] Open
Abstract
Hepatitis E virus (HEV) is an important cause of acute hepatitis in humans worldwide. In industrialised countries, most infections are caused by the zoonotic genotype 3. The main reservoir was found in pigs, with fattening pigs as the main shedders. The aim of this study was to establish a screening tool to detect HEV in pig farms. HEV-positive samples were sequenced using Sanger sequencing. First, different sample materials, including floor swabs, slurry, dust swabs and faeces were tested for HEV. Floor swabs turned out to give the best results and, in the form of sock swabs, were used for the screening of Swiss pig herds. A total of 138 pig farms were tested, with a focus on fattening pigs. Overall, 81 farms (58.8%) were HEV positive. Most sequences belonged to subtype 3h, in which they formed a specific cluster (Swiss cluster). In addition, subtype 3l and two unassigned sequences were detected. As a conclusion, sock swabs were found to be a helpful tool to screen pig herds for HEV and establish a sequence collection that may enable molecular epidemiology and support outbreak investigation and prevention.
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Affiliation(s)
- Julia Lienhard
- Institute of Virology, Vetsuisse Faculty, University of Zurich, 8057 Zurich, Switzerland; (J.L.); (I.V.-S.)
| | | | - Xaver Sidler
- Division of Swine Medicine, Department of Farm Animals, Vetsuisse Faculty, University of Zurich, 8057 Zurich, Switzerland;
| | - Claudia Bachofen
- Institute of Virology, Vetsuisse Faculty, University of Zurich, 8057 Zurich, Switzerland; (J.L.); (I.V.-S.)
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