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Supekar R, Sarkar J, Chakrabarti P, Biswas S. Diagnostic challenges due to hepatitis B virus surface antigen mutations outside the major hydrophilic region. Arch Virol 2025; 170:71. [PMID: 40063291 DOI: 10.1007/s00705-025-06256-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2024] [Accepted: 01/29/2025] [Indexed: 03/29/2025]
Abstract
The number of observed cases of occult hepatitis B virus infection (OBI) in eastern India has been increasing. Here, S gene mutations were identified in apparently healthy individuals with OBI, and the S protein variants from these patients were characterized in vitro. Plasma samples from 217 healthy blood donors were collected from three different regions in eastern India and screened for hepatitis B virus (HBV) infection using a nucleic acid amplification test and immunoassays for serological markers. S protein variants found in positive plasma samples were characterized using a liver cell line. Twenty-nine of the 217 plasma samples tested, were positive for HBV DNA and were negative for hepatitis B surface antigen (HBsAg) and antibody to HBV core antigen (anti-HBc). Sequencing of the HBV S gene revealed a novel S protein mutation (L173H) in an area outside the major hydrophilic region. Known OBI-associated mutations (S34L, P178R), a mutation resulting in a stop codon at position 196, associated with lamivudine-resistance, the substitution I81T, and a dual mutation (G145A and Q101H) were also identified. S proteins containing these mutations, produced by transfection of human hepatoma (Huh7) cells with recombinant plasmids, were undetectable or gave significantly weaker signals than the wild-type control, despite similar levels of S mRNA production for the mutant and wild-type plasmids. The OBI cases in this study were unexpectedly seronegative. In vitro analysis revealed that the mutations identified here caused the virus to evade immunodetection using commercial immunoassays, thereby rendering a large portion of the population "silently" infected with HBV.
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Affiliation(s)
- Ruchi Supekar
- Infectious Diseases and Immunology Division, Council of Scientific and Industrial Research-Indian Institute of Chemical Biology (CSIR-IICB), 4, Raja S.C. Mullick Road, Kolkata, West Bengal, 700032, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Jit Sarkar
- Infectious Diseases and Immunology Division, Council of Scientific and Industrial Research-Indian Institute of Chemical Biology (CSIR-IICB), 4, Raja S.C. Mullick Road, Kolkata, West Bengal, 700032, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
- Community Health Program, SWANIRVAR, North 24 Parganas, West Bengal, India
| | - Partha Chakrabarti
- Infectious Diseases and Immunology Division, Council of Scientific and Industrial Research-Indian Institute of Chemical Biology (CSIR-IICB), 4, Raja S.C. Mullick Road, Kolkata, West Bengal, 700032, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Subhajit Biswas
- Infectious Diseases and Immunology Division, Council of Scientific and Industrial Research-Indian Institute of Chemical Biology (CSIR-IICB), 4, Raja S.C. Mullick Road, Kolkata, West Bengal, 700032, India.
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India.
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2
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McCoullough LC, Sadauskas T, Sozzi V, Mak KY, Mason H, Littlejohn M, Revill PA. The in vitro replication phenotype of hepatitis B virus (HBV) splice variants Sp3 and Sp9 and their impact on wild-type HBV replication. J Virol 2024; 98:e0153823. [PMID: 38501924 PMCID: PMC11019940 DOI: 10.1128/jvi.01538-23] [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: 10/02/2023] [Accepted: 02/22/2024] [Indexed: 03/20/2024] Open
Abstract
Prior to nuclear export, the hepatitis B virus (HBV) pregenomic RNA may be spliced by the host cell spliceosome to form shorter RNA sequences known as splice variants. Due to deletions in the open reading frames, splice variants may encode novel fusion proteins. Although not essential for HBV replication, the role of splice variants and their novel fusion proteins largely remains unknown. Some splice variants and their encoded novel fusion proteins have been shown to impair or promote wild-type HBV replication in vitro, and although splice variants Sp3 and Sp9 are two of the most common splice variants identified to date, their in vitro replication phenotype and their impact on wild-type HBV replication are unclear. Here, we utilize greater than genome-length Sp3 and Sp9 constructs to investigate their replication phenotype in vitro, and their impact on wild-type HBV replication. We show that Sp3 and Sp9 were incapable of autonomous replication, which was rescued by providing the polymerase and core proteins in trans. Furthermore, we showed that Sp3 had no impact on wild-type HBV replication, whereas Sp9 strongly reduced wild-type HBV replication in co-transfection experiments. Knocking out Sp9 novel precore-surface and core-surface fusion protein partially restored replication, suggesting that these proteins contributed to suppression of wild-type HBV replication, providing further insights into factors regulating HBV replication in vitro. IMPORTANCE The role of hepatitis B virus (HBV) splice variants in HBV replication and pathogenesis currently remains largely unknown. However, HBV splice variants have been associated with the development of hepatocellular carcinoma, suggesting a role in HBV pathogenesis. Several in vitro co-transfection studies have shown that different splice variants have varying impacts on wild-type HBV replication, perhaps contributing to viral persistence. Furthermore, all splice variants are predicted to produce novel fusion proteins. Sp1 hepatitis B splice protein contributes to liver disease progression and apoptosis; however, the function of other HBV splice variant novel fusion proteins remains largely unknown. We show that Sp9 markedly impairs HBV replication in a cell culture co-transfection model, mediated by expression of Sp9 novel fusion proteins. In contrast, Sp3 had no effect on wild-type HBV replication. Together, these studies provide further insights into viral factors contributing to regulation of HBV replication.
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Affiliation(s)
- Laura C. McCoullough
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Tomas Sadauskas
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Vitina Sozzi
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Kai Yan Mak
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Hugh Mason
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Margaret Littlejohn
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
- Department of Infectious Diseases, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
| | - Peter A. Revill
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
- Department of Infectious Diseases, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia
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3
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Giraud G, El Achi K, Zoulim F, Testoni B. Co-Transcriptional Regulation of HBV Replication: RNA Quality Also Matters. Viruses 2024; 16:615. [PMID: 38675956 PMCID: PMC11053573 DOI: 10.3390/v16040615] [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: 03/25/2024] [Revised: 04/12/2024] [Accepted: 04/15/2024] [Indexed: 04/28/2024] Open
Abstract
Chronic hepatitis B (CHB) virus infection is a major public health burden and the leading cause of hepatocellular carcinoma. Despite the efficacy of current treatments, hepatitis B virus (HBV) cannot be fully eradicated due to the persistence of its minichromosome, or covalently closed circular DNA (cccDNA). The HBV community is investing large human and financial resources to develop new therapeutic strategies that either silence or ideally degrade cccDNA, to cure HBV completely or functionally. cccDNA transcription is considered to be the key step for HBV replication. Transcription not only influences the levels of viral RNA produced, but also directly impacts their quality, generating multiple variants. Growing evidence advocates for the role of the co-transcriptional regulation of HBV RNAs during CHB and viral replication, paving the way for the development of novel therapies targeting these processes. This review focuses on the mechanisms controlling the different co-transcriptional processes that HBV RNAs undergo, and their contribution to both viral replication and HBV-induced liver pathogenesis.
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Affiliation(s)
- Guillaume Giraud
- INSERM U1052, CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Université Claude Bernard Lyon 1, 69008 Lyon, France (F.Z.)
- The Lyon Hepatology Institute EVEREST, 69003 Lyon, France
| | - Khadija El Achi
- INSERM U1052, CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Université Claude Bernard Lyon 1, 69008 Lyon, France (F.Z.)
| | - Fabien Zoulim
- INSERM U1052, CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Université Claude Bernard Lyon 1, 69008 Lyon, France (F.Z.)
- The Lyon Hepatology Institute EVEREST, 69003 Lyon, France
- Hospices Civils de Lyon, Hôpital Croix Rousse, Service d’Hépato-Gastroentérologie, 69004 Lyon, France
| | - Barbara Testoni
- INSERM U1052, CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, Université Claude Bernard Lyon 1, 69008 Lyon, France (F.Z.)
- The Lyon Hepatology Institute EVEREST, 69003 Lyon, France
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Im YR, Jagdish R, Leith D, Kim JU, Yoshida K, Majid A, Ge Y, Ndow G, Shimakawa Y, Lemoine M. Prevalence of occult hepatitis B virus infection in adults: a systematic review and meta-analysis. Lancet Gastroenterol Hepatol 2022; 7:932-942. [PMID: 35961359 PMCID: PMC9630161 DOI: 10.1016/s2468-1253(22)00201-1] [Citation(s) in RCA: 19] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/05/2022] [Revised: 06/14/2022] [Accepted: 06/15/2022] [Indexed: 12/03/2022]
Abstract
Background Despite growing concerns about transmissibility and clinical impact, occult hepatitis B virus (HBV) infection has received little attention in the hepatitis elimination agenda. We aimed to estimate the prevalence of occult HBV infection at a global and regional scale and in specific populations. Methods For this systematic review and meta-analysis, we searched the MEDLINE, Embase, Global Health, and Web of Science databases for articles published in any language between Jan 1, 2010, and Aug 14, 2019. We included original articles and conference abstracts of any study design that reported the proportion of HBsAg-negative adults (aged ≥18 years) who are positive for HBV DNA (ie, people with occult HBV infection). The prevalence of occult HBV infection was pooled, using the DerSimonian-Laird random-effects model, in the general population and specific groups defined by the type of study participants (blood donors; other low-risk populations; high-risk populations; and people with advanced chronic liver disease), and stratified by HBV endemicity in each country. We also assessed the performance of anti-HBc as an alternative biomarker to detect occult HBV infection. The study was registered with PROSPERO, CRD42019115490. Findings 305 of 3962 articles were eligible, allowing a meta-analysis of 140 521 993 individuals tested for HBV DNA. Overall, only two studies evaluated occult HBV infection in the general population, precluding unbiased global and regional estimates of occult HBV infection prevalence. In blood donors, occult HBV infection prevalence mirrored HBV endemicity: 0·06% (95% CI 0·00–0·26) in low-endemicity countries, 0·12% (0·04–0·23) in intermediate-endemicity countries, and 0·98% (0·44–1·72), in high-endemicity countries (p=0·0012). In high-risk groups, occult HBV infection prevalence was substantial, irrespective of endemicity: 5·5% (95% CI 2·9–8·7) in low-endemicity countries, 5·2% (2·5–8·6) in intermediate-endemicity countries, and 12·0% (3·4–24·7) in high-endemicity countries. The pooled sensitivity of anti-HBc to identify occult HBV infection was 77% (95% CI 62–88) and its specificity was 76% (68–83). Interpretation A substantial proportion of people carry occult HBV infection, especially among high-risk groups across the globe and people living in highly endemic countries. Occult HBV infection should be part of the global viral hepatitis elimination strategy. Funding None.
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5
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Saitta C, Pollicino T, Raimondo G. Occult Hepatitis B Virus Infection: An Update. Viruses 2022; 14:v14071504. [PMID: 35891484 PMCID: PMC9318873 DOI: 10.3390/v14071504] [Citation(s) in RCA: 47] [Impact Index Per Article: 15.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2022] [Revised: 06/28/2022] [Accepted: 06/29/2022] [Indexed: 11/16/2022] Open
Abstract
Occult hepatitis B virus (HBV) infection (OBI) refers to a condition in which replication-competent viral DNA is present in the liver (with detectable or undetectable HBV DNA in the serum) of individuals testing negative for the HBV surface antigen (HBsAg). In this peculiar phase of HBV infection, the covalently closed circular DNA (cccDNA) is in a low state of replication. Many advances have been made in clarifying the mechanisms involved in such a suppression of viral activity, which seems to be mainly related to the host's immune control and epigenetic factors. OBI is diffused worldwide, but its prevalence is highly variable among patient populations. This depends on different geographic areas, risk factors for parenteral infections, and assays used for HBsAg and HBV DNA detection. OBI has an impact in several clinical contexts: (a) it can be transmitted, causing a classic form of hepatitis B, through blood transfusion or liver transplantation; (b) it may reactivate in the case of immunosuppression, leading to the possible development of even fulminant hepatitis; (c) it may accelerate the progression of chronic liver disease due to different causes toward cirrhosis; (d) it maintains the pro-oncogenic properties of the "overt" infection, favoring the development of hepatocellular carcinoma.
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Affiliation(s)
- Carlo Saitta
- Division of Medicine and Hepatology, University Hospital of Messina, 98124 Messina, Italy;
- Department of Clinical and Experimental Medicine, University of Messina, 98124 Messina, Italy
| | - Teresa Pollicino
- Department of Human Pathology, University Hospital of Messina, 98124 Messina, Italy;
| | - Giovanni Raimondo
- Division of Medicine and Hepatology, University Hospital of Messina, 98124 Messina, Italy;
- Department of Clinical and Experimental Medicine, University of Messina, 98124 Messina, Italy
- Correspondence: ; Tel.: +39-(0)-902212392
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6
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Gherlan GS. Occult hepatitis B — the result of the host immune response interaction with different genomic expressions of the virus. World J Clin Cases 2022; 10:5518-5530. [PMID: 35979101 PMCID: PMC9258381 DOI: 10.12998/wjcc.v10.i17.5518] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/05/2022] [Revised: 01/30/2022] [Accepted: 04/03/2022] [Indexed: 02/06/2023] Open
Abstract
With over 40 years of history, occult hepatitis B infection (OBI) continues to remain an important and challenging public health problem. Defined as the presence of replication-competent hepatitis B virus (HBV) DNA (i.e., episomal HBV covalently closed circular DNA) in the liver and/or HBV DNA in the blood of people who test negative for hepatitis B surface antigen (HBsAg) in currently available assays, OBI is currently diagnosed using polymerase chain reaction (PCR) and real-time PCR assays. However, all efforts should be made to exclude a false negative HBsAg in order to completely follow the definition of OBI. In recent years, significant advances have been made in understanding the HBV lifecycle and the molecular mechanisms that lead to the persistence of the virus in the occult form. These factors are mainly related to the host immune system and, to a smaller proportion, to the virus. Both innate and adaptive immune responses are important in HBV infection management, and epigenetic changes driven by host mechanisms (acetylation, methylation, and microRNA implication) are added to such actions. Although greater genetic variability in the S gene of HBV isolated from OBIs was found compared with overt infection, the mechanisms of OBI are not mainly viral mutations.
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Affiliation(s)
- George Sebastian Gherlan
- Department of Infectious Diseases, “Carol Davila” University of Medicine and Pharmacy, Bucharest 030303, Romania
- Department of Infectious Diseases, “Dr. Victor Babes” Hospital of Infectious and Tropical Diseases, Bucharest 030303, Romania
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7
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Kremsdorf D, Lekbaby B, Bablon P, Sotty J, Augustin J, Schnuriger A, Pol J, Soussan P. Alternative splicing of viral transcripts: the dark side of HBV. Gut 2021; 70:2373-2382. [PMID: 34535538 DOI: 10.1136/gutjnl-2021-324554] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/02/2021] [Accepted: 09/06/2021] [Indexed: 02/06/2023]
Abstract
Regulation of alternative splicing is one of the most efficient mechanisms to enlarge the proteomic diversity in eukaryotic organisms. Many viruses hijack the splicing machinery following infection to accomplish their replication cycle. Regarding the HBV, numerous reports have described alternative splicing events of the long viral transcript (pregenomic RNA), which also acts as a template for viral genome replication. Alternative splicing of HBV pregenomic RNAs allows the synthesis of at least 20 spliced variants. In addition, almost all these spliced forms give rise to defective particles, detected in the blood of infected patients. HBV-spliced RNAs have long been unconsidered, probably due to their uneasy detection in comparison to unspliced forms as well as for their dispensable role during viral replication. However, recent data highlighted the relevance of these HBV-spliced variants through (1) the trans-regulation of the alternative splicing of viral transcripts along the course of liver disease; (2) the ability to generate defective particle formation, putative biomarker of the liver disease progression; (3) modulation of viral replication; and (4) their intrinsic propensity to encode for novel viral proteins involved in liver pathogenesis and immune response. Altogether, tricky regulation of HBV alternative splicing may contribute to modulate multiple viral and cellular processes all along the course of HBV-related liver disease.
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Affiliation(s)
- Dina Kremsdorf
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France
| | - Bouchra Lekbaby
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France
| | - Pierre Bablon
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France
| | - Jules Sotty
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France
| | - Jérémy Augustin
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France
| | - Aurélie Schnuriger
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France.,Assistance Publique - Hôpitaux de Paris, Département de Virologie, GHU Paris-Est, Paris, France
| | - Jonathan Pol
- Institut National de la Santé et de la Recherche Médicale U1138, Centre de Recherche des Cordeliers, Université de Paris, Sorbonne Université, Paris, France.,Metabolomics ann Cell Biology Platforms, Gustave Roussy Cancer Campus, Villejuif, France
| | - Patrick Soussan
- Institut National de la Santé et de la Recherche Médicale U938, Centre de Recherche de Saint Antoine, Sorbonne Université-Faculté Saint Antoine, Paris, France .,Assistance Publique - Hôpitaux de Paris, Département de Virologie, GHU Paris-Est, Paris, France
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8
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Olagbenro M, Anderson M, Gaseitsiwe S, Powell EA, Gededzha MP, Selabe SG, Blackard JT. In silico analysis of mutations associated with occult hepatitis B virus (HBV) infection in South Africa. Arch Virol 2021; 166:3075-3084. [PMID: 34468889 PMCID: PMC11930061 DOI: 10.1007/s00705-021-05196-7] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2020] [Accepted: 06/09/2021] [Indexed: 01/02/2023]
Abstract
Occult hepatitis B virus (OBI) infection is defined by the presence of viral DNA in the liver and/or serum in absence of hepatitis B surface antigen (HBsAg). While multiple studies have identified mutations that are associated with OBI, only a small portion of these mutations have been functionally characterized in vitro. Using complementary in silico approaches, the effects of OBI-associated amino acid mutations on HBV protein function in HBV/HIV-positive ART-naïve South Africans were evaluated. Two OBI-associated mutations in the PreS1 region, one in the PreS2 region, and seven in the surface region of subgenotype A1 sequences were identified as deleterious. In subgenotype A2 sequences, 11 OBI-associated mutations in the PreS1 region, seven in the PreS2 region, and 31 in the surface region were identified as deleterious. In the polymerase region, 14 OBI-associated mutations in subgenotype A1 and 71 OBI-associated mutations in subgenotype A2 were identified as deleterious. This study utilized in silico approaches to characterize the likely impact of OBI-associated mutations on viral function, thereby identifying and prioritizing candidates and reducing the significant cost associated with functional studies that are essential for mechanistic studies of the OBI phenotype.
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Affiliation(s)
- Matthew Olagbenro
- Division of Digestive Diseases, University of Cincinnati College of Medicine, ML 0595, Albert Sabin Way, Cincinnati, OH, 45267-0595, USA
| | | | | | - Eleanor A Powell
- Division of Digestive Diseases, University of Cincinnati College of Medicine, ML 0595, Albert Sabin Way, Cincinnati, OH, 45267-0595, USA
- Department of Pathology and Laboratory Medicine, University of Cincinnati College of Medicine, Cincinnati, OH, USA
| | - Maemu P Gededzha
- Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of Witwatersrand and National Health Laboratory Service, Johannesburg, South Africa
- HIV and Hepatitis Research Unit, Department of Virology, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | - Selokela G Selabe
- HIV and Hepatitis Research Unit, Department of Virology, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | - Jason T Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, ML 0595, Albert Sabin Way, Cincinnati, OH, 45267-0595, USA.
- HIV and Hepatitis Research Unit, Department of Virology, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa.
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9
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Ghosh S, Chakraborty A, Banerjee S. Persistence of Hepatitis B Virus Infection: A Multi-Faceted Player for Hepatocarcinogenesis. Front Microbiol 2021; 12:678537. [PMID: 34526974 PMCID: PMC8435854 DOI: 10.3389/fmicb.2021.678537] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2021] [Accepted: 07/06/2021] [Indexed: 12/14/2022] Open
Abstract
Hepatitis B virus (HBV) infection has a multi-dimensional effect on the host, which not only alters the dynamics of immune response but also persists in the hepatocytes to predispose oncogenic factors. The virus exists in multiple forms of which the nuclear localized covalently closed circular DNA (cccDNA) is the most stable and the primary reason for viral persistence even after clearance of surface antigen and viral DNA. The second reason is the existence of pregenomic RNA (pgRNA) containing virion particles. On the other hand, the integration of the viral genome in the host chromosome also leads to persistent production of viral proteins along with the chromosomal instabilities. The interferon treatment or administration of nucleot(s)ide analogs leads to reduction in the viral DNA load, but the pgRNA and surface antigen clearance are a slow process and complete loss of serological HBsAg is rare. The prolonged exposure of immune cells to the viral antigens, particularly HBs antigen, in the blood circulation results in T-cell exhaustion, which disrupts immune clearance of the virus and virus-infected cells. In addition, it predisposes immune-tolerant microenvironment, which facilitates the tumor progression. Thus cccDNA, pgRNA, and HBsAg along with the viral DNA could be the therapeutic targets in the early disease stages that may improve the quality of life of chronic hepatitis B patients by impeding the progression of the disease toward hepatocellular carcinoma.
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Affiliation(s)
| | | | - Soma Banerjee
- Centre for Liver Research, School of Digestive and Liver Diseases, Institute of Post Graduate Medical Education and Research, Kolkata, India
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10
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Ye X, Zhao Y, Li R, Li T, Zheng X, Xiong W, Zeng J, Xu M, Chen L. High Frequency Occult Hepatitis B Virus Infection Detected in Non-Resolved Donations Suggests the Requirement of Anti-HBc Test in Blood Donors in Southern China. Front Immunol 2021; 12:699217. [PMID: 34394093 PMCID: PMC8355616 DOI: 10.3389/fimmu.2021.699217] [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: 04/26/2021] [Accepted: 07/12/2021] [Indexed: 02/04/2023] Open
Abstract
Background Most Chinese Blood Centers adopted mini pool (MP) nucleic acid testing (NAT) for HBV screening due to high cost of Individual donation (ID) NAT, and different proportions of MP-reactive but ID-non-reactive donations (MP+/ID-, defined as non-resolved donations) have been observed during daily donor screening process. Some of these non-resolved donations are occult HBV infections (OBIs), which pose potential risk of HBV transmission if they are not deferred. This study is aimed to further analyze these non-resolved donations. Methods The non-resolved plasma samples were further analyzed by serological tests and various HBV DNA amplification assays including quantitative PCR (qPCR) and nested PCR amplifying the basic core and pre-core promoter regions (BCP/PC; 295 base pairs) and HBsAg (S) region (496 base pairs). Molecular characterizations of HBV DNA+ non-resolved samples were determined by sequencing analysis. Results Of 17,226 MPs from 103,356 seronegative blood donations, 98 MPs were detected reactive for HBV. Fifty-six out of these 98 (57.1%) reactive MPs were resolved as HBV DNA+, but the remaining 42 pools (42.9%, 252 donations) were left non-resolved with a high rate (53.2%) of anti-HBc+. Surprisingly, among 42 non-resolved MPs, 17 contained one donation identified as OBIs by alternative NAT assays. Sequence analysis on HBV DNAs extracted from these OBI donations showed some key mutations in the S region that may lead to failure in HBsAg detection and vaccine escape. Conclusion A total of 53.2% of the non-resolved donations were anti-HBc+, and OBIs were identified in 40.5% of these non-resolved pools. Therefore, non-resolved donations with anti-HBc+ might pose potential risk for HBV transmission. Our present analysis indicates that anti-HBc testing in non-resolved donations should be used to identify OBIs in order to further increase blood safety in China.
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Affiliation(s)
- Xianlin Ye
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Yu Zhao
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Ran Li
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Tong Li
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Xin Zheng
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Wen Xiong
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Jinfeng Zeng
- Department of Laboratory, Shenzhen Blood Center, Shenzhen, China
| | - Min Xu
- Provincial Key Laboratory for Transfusion-Transmitted Infectious Diseases, Institute of Blood Transfusion, Chinese Academy of Medical Sciences (CAMS) and Peking Union Medical College (PUMC), Chengdu, China
| | - Limin Chen
- Provincial Key Laboratory for Transfusion-Transmitted Infectious Diseases, Institute of Blood Transfusion, Chinese Academy of Medical Sciences (CAMS) and Peking Union Medical College (PUMC), Chengdu, China.,The Joint Laboratory on Transfusion-Transmitted Diseases (TTD) Between Institute of Blood Transfusion, Nanning Blood Center, Chinese Academy of Medical Sciences and Nanning Blood Center, Nanning, China.,Toronto General Research Institute, University Health Network, University of Toronto, Toronto, ON, Canada
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11
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Liu Y, Veeraraghavan V, Pinkerton M, Fu J, Douglas MW, George J, Tu T. Viral Biomarkers for Hepatitis B Virus-Related Hepatocellular Carcinoma Occurrence and Recurrence. Front Microbiol 2021; 12:665201. [PMID: 34194408 PMCID: PMC8236856 DOI: 10.3389/fmicb.2021.665201] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2021] [Accepted: 05/06/2021] [Indexed: 12/24/2022] Open
Abstract
Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide and the fourth leading cause of cancer-related death. The most common risk factor for developing HCC is chronic infection with hepatitis B virus (HBV). Early stages of HBV-related HCC (HBV-HCC) are generally asymptomatic. Moreover, while serum alpha-fetoprotein (AFP) and abdominal ultrasound are widely used to screen for HCC, they have poor sensitivity. Thus, HBV-HCC is frequently diagnosed at an advanced stage, in which there are limited treatment options and high mortality rates. Serum biomarkers with high sensitivity and specificity are crucial for earlier diagnosis of HCC and improving survival rates. As viral-host interactions are key determinants of pathogenesis, viral biomarkers may add greater diagnostic power for HCC than host biomarkers alone. In this review, we summarize recent research on using virus-derived biomarkers for predicting HCC occurrence and recurrence; including circulating viral DNA, RNA transcripts, and viral proteins. Combining these viral biomarkers with AFP and abdominal ultrasound could improve sensitivity and specificity of early diagnosis, increasing the survival of patients with HBV-HCC. In the future, as the mechanisms that drive HBV-HCC to become clearer, new biomarkers may be identified which can further improve early diagnosis of HBV-HCC.
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Affiliation(s)
- Yuanyuan Liu
- Department of Infectious Diseases, The Affiliated Xi'an Central Hospital of Xi'an Jiaotong University, Xi'an, China.,Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia
| | - Vaishnavi Veeraraghavan
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia.,School of Medical Science, The University of Sydney, Camperdown, NSW, Australia
| | - Monica Pinkerton
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia.,School of Medical Science, The University of Sydney, Camperdown, NSW, Australia
| | - Jianjun Fu
- Department of Infectious Diseases, The Affiliated Xi'an Central Hospital of Xi'an Jiaotong University, Xi'an, China
| | - Mark W Douglas
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia.,Marie Bashir Institute for Infectious Diseases and Biosecurity, University of Sydney, Sydney, NSW, Australia.,Centre for Infectious Diseases and Microbiology, Westmead Hospital, Sydney, NSW, Australia
| | - Jacob George
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia
| | - Thomas Tu
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW, Australia.,Marie Bashir Institute for Infectious Diseases and Biosecurity, University of Sydney, Sydney, NSW, Australia
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12
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Rajaby R, Zhou Y, Meng Y, Zeng X, Li G, Wu P, Sung WK. SurVirus: a repeat-aware virus integration caller. Nucleic Acids Res 2021; 49:e33. [PMID: 33444454 PMCID: PMC8034624 DOI: 10.1093/nar/gkaa1237] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2020] [Revised: 12/01/2020] [Accepted: 01/12/2021] [Indexed: 01/01/2023] Open
Abstract
A significant portion of human cancers are due to viruses integrating into human genomes. Therefore, accurately predicting virus integrations can help uncover the mechanisms that lead to many devastating diseases. Virus integrations can be called by analysing second generation high-throughput sequencing datasets. Unfortunately, existing methods fail to report a significant portion of integrations, while predicting a large number of false positives. We observe that the inaccuracy is caused by incorrect alignment of reads in repetitive regions. False alignments create false positives, while missing alignments create false negatives. This paper proposes SurVirus, an improved virus integration caller that corrects the alignment of reads which are crucial for the discovery of integrations. We use publicly available datasets to show that existing methods predict hundreds of thousands of false positives; SurVirus, on the other hand, is significantly more precise while it also detects many novel integrations previously missed by other tools, most of which are in repetitive regions. We validate a subset of these novel integrations, and find that the majority are correct. Using SurVirus, we find that HPV and HBV integrations are enriched in LINE and Satellite regions which had been overlooked, as well as discover recurrent HBV and HPV breakpoints in human genome-virus fusion transcripts.
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Affiliation(s)
- Ramesh Rajaby
- School of Computing, National University of Singapore, 13 Computing Drive, 117417, Singapore.,NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, 117456, Singapore
| | - Yi Zhou
- Agricultural Bioinformatics Key Laboratory of Hubei Province, Hubei Engineering Technology Research Center of Agricultural Big Data, College of Informatics, Huazhong Agricultural University, Wuhan 430070, China
| | - Yifan Meng
- Department of Gynecologic Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.,Cancer Biology Research Center (Key laboratory of the ministry of education), Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Xi Zeng
- Agricultural Bioinformatics Key Laboratory of Hubei Province, Hubei Engineering Technology Research Center of Agricultural Big Data, College of Informatics, Huazhong Agricultural University, Wuhan 430070, China
| | - Guoliang Li
- Agricultural Bioinformatics Key Laboratory of Hubei Province, Hubei Engineering Technology Research Center of Agricultural Big Data, College of Informatics, Huazhong Agricultural University, Wuhan 430070, China
| | - Peng Wu
- Department of Gynecologic Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.,Cancer Biology Research Center (Key laboratory of the ministry of education), Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Wing-Kin Sung
- School of Computing, National University of Singapore, 13 Computing Drive, 117417, Singapore.,Agricultural Bioinformatics Key Laboratory of Hubei Province, Hubei Engineering Technology Research Center of Agricultural Big Data, College of Informatics, Huazhong Agricultural University, Wuhan 430070, China.,Genome Institute of Singapore, 60 Biopolis Street, Genome 138672, Singapore
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13
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Lim CS, Sozzi V, Littlejohn M, Yuen LK, Warner N, Betz-Stablein B, Luciani F, Revill PA, Brown CM. Quantitative analysis of the splice variants expressed by the major hepatitis B virus genotypes. Microb Genom 2021; 7:mgen000492. [PMID: 33439114 PMCID: PMC8115900 DOI: 10.1099/mgen.0.000492] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2020] [Accepted: 11/23/2020] [Indexed: 12/13/2022] Open
Abstract
Hepatitis B virus (HBV) is a major human pathogen that causes liver diseases. The main HBV RNAs are unspliced transcripts that encode the key viral proteins. Recent studies have shown that some of the HBV spliced transcript isoforms are predictive of liver cancer, yet the roles of these spliced transcripts remain elusive. Furthermore, there are nine major HBV genotypes common in different regions of the world, these genotypes may express different spliced transcript isoforms. To systematically study the HBV splice variants, we transfected human hepatoma cells, Huh7, with four HBV genotypes (A2, B2, C2 and D3), followed by deep RNA-sequencing. We found that 13-28 % of HBV RNAs were splice variants, which were reproducibly detected across independent biological replicates. These comprised 6 novel and 10 previously identified splice variants. In particular, a novel, singly spliced transcript was detected in genotypes A2 and D3 at high levels. The biological relevance of these splice variants was supported by their identification in HBV-positive liver biopsy and serum samples, and in HBV-infected primary human hepatocytes. Interestingly the levels of HBV splice variants varied across the genotypes, but the spliced pregenomic RNA SP1 and SP9 were the two most abundant splice variants. Counterintuitively, these singly spliced SP1 and SP9 variants had a suboptimal 5' splice site, supporting the idea that splicing of HBV RNAs is tightly controlled by the viral post-transcriptional regulatory RNA element.
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Affiliation(s)
- Chun Shen Lim
- Department of Biochemistry, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand
| | - Vitina Sozzi
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Margaret Littlejohn
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Lilly K.W. Yuen
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Nadia Warner
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Brigid Betz-Stablein
- Systems Medicine, School of Medical Sciences, Faculty of Medicine, University of New South Wales, Sydney, New South Wales, Australia
- Present address: Dermatology Research Centre, Diamantina Institute, University of Queensland, Brisbane, Queensland, Australia
| | - Fabio Luciani
- Systems Medicine, School of Medical Sciences, Faculty of Medicine, University of New South Wales, Sydney, New South Wales, Australia
| | - Peter A. Revill
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
- Department of Microbiology and Immunology, University of Melbourne, Melbourne, Victoria, Australia
| | - Chris M. Brown
- Department of Biochemistry, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand
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14
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The evolution and clinical impact of hepatitis B virus genome diversity. Nat Rev Gastroenterol Hepatol 2020; 17:618-634. [PMID: 32467580 DOI: 10.1038/s41575-020-0296-6] [Citation(s) in RCA: 101] [Impact Index Per Article: 20.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 03/20/2020] [Indexed: 02/06/2023]
Abstract
The global burden of hepatitis B virus (HBV) is enormous, with 257 million persons chronically infected, resulting in more than 880,000 deaths per year worldwide. HBV exists as nine different genotypes, which differ in disease progression, natural history and response to therapy. HBV is an ancient virus, with the latest reports greatly expanding the host range of the Hepadnaviridae (to include fish and reptiles) and casting new light on the origins and evolution of this viral family. Although there is an effective preventive vaccine, there is no cure for chronic hepatitis B, largely owing to the persistence of a viral minichromosome that is not targeted by current therapies. HBV persistence is also facilitated through aberrant host immune responses, possibly due to the diverse intra-host viral populations that can respond to host-mounted and therapeutic selection pressures. This Review summarizes current knowledge on the influence of HBV diversity on disease progression and treatment response and the potential effect on new HBV therapies in the pipeline. The mechanisms by which HBV diversity can occur both within the individual host and at a population level are also discussed.
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15
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Choga WT, Anderson M, Zumbika E, Phinius BB, Mbangiwa T, Bhebhe LN, Baruti K, Kimathi PO, Seatla KK, Musonda RM, Bell TG, Moyo S, Blackard JT, Gaseitsiwe S. In Silico Prediction of Human Leukocytes Antigen (HLA) Class II Binding Hepatitis B Virus (HBV) Peptides in Botswana. Viruses 2020; 12:E731. [PMID: 32640609 PMCID: PMC7412261 DOI: 10.3390/v12070731] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2020] [Accepted: 07/03/2020] [Indexed: 02/07/2023] Open
Abstract
Hepatitis B virus (HBV) is the primary cause of liver-related malignancies worldwide, and there is no effective cure for chronic HBV infection (CHB) currently. Strong immunological responses induced by T cells are associated with HBV clearance during acute infection; however, the repertoire of epitopes (epi) presented by major histocompatibility complexes (MHCs) to elicit these responses in various African populations is not well understood. In silico approaches were used to map and investigate 15-mers HBV peptides restricted to 9 HLA class II alleles with high population coverage in Botswana. Sequences from 44 HBV genotype A and 48 genotype D surface genes (PreS/S) from Botswana were used. Of the 1819 epi bindings predicted, 20.2% were strong binders (SB), and none of the putative epi bind to all the 9 alleles suggesting that multi-epitope, genotype-based, population-based vaccines will be more effective against HBV infections as opposed to previously proposed broad potency epitope-vaccines which were assumed to work for all alleles. In total, there were 297 unique epi predicted from the 3 proteins and amongst, S regions had the highest number of epi (n = 186). Epitope-densities (Depi) between genotypes A and D were similar. A number of mutations that hindered HLA-peptide binding were observed. We also identified antigenic and genotype-specific peptides with characteristics that are well suited for the development of sensitive diagnostic kits. This study identified candidate peptides that can be used for developing multi-epitope vaccines and highly sensitive diagnostic kits against HBV infection in an African population. Our results suggest that viral variability may hinder HBV peptide-MHC binding, required to initiate a cascade of immunological responses against infection.
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Affiliation(s)
- Wonderful Tatenda Choga
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Division of Human Genetics, Department of Pathology, Faculty of Health Sciences, University of Cape Town, Cape Town 7925, South Africa
| | - Motswedi Anderson
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
| | - Edward Zumbika
- Department of Applied Biology and Biochemistry, Faculty of Applied Sciences, National University of Science and Technology, Bulawayo 0000, Zimbabwe;
| | - Bonolo B. Phinius
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
| | - Tshepiso Mbangiwa
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Division of Human Genetics, Department of Pathology, Faculty of Health Sciences, University of Cape Town, Cape Town 7925, South Africa
| | - Lynnette N. Bhebhe
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
| | - Kabo Baruti
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Department of Biological Sciences, Faculty of Science, University of Botswana, Gaborone 0000, Botswana
| | | | - Kaelo K. Seatla
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Department of Medical Laboratory Sciences, Faculty of Health Sciences, University of Botswana, Gaborone 0000, Botswana
| | - Rosemary M. Musonda
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA
| | - Trevor Graham Bell
- Independent Researcher, P.O. Box 497, Wits, Johannesburg 2050, South Africa;
| | - Sikhulile Moyo
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA
| | - Jason T. Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA;
| | - Simani Gaseitsiwe
- Research Laboratory, Botswana Harvard AIDS Institute Partnership, Gaborone 0000, Botswana; (W.T.C.); (M.A.); (B.B.P.); (T.M.); (L.N.B.); (K.B.); (K.K.S.); (R.M.M.); (S.M.)
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA
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16
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Elalfy H, Besheer T, Elhammady D, El Mesery A, Shaltout SW, Abd El-Maksoud M, Amin AI, Bekhit AN, Abd El Aziz M, El-Bendary M. Pathological characterization of occult hepatitis B virus infection in hepatitis C virus-associated or non-alcoholic steatohepatitis-related hepatocellular carcinoma. World J Meta-Anal 2020; 8:67-77. [DOI: 10.13105/wjma.v8.i2.67] [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: 11/12/2019] [Revised: 01/08/2020] [Accepted: 04/09/2020] [Indexed: 02/06/2023] Open
Abstract
Occult hepatitis B virus (HBV) infection, by definition, is a state in which infection with this virus does not manifest with the conventional diagnostic laboratory criteria reserved for the obvious form of HBV infection. As a result, occult HBV infection is commonly a surprise finding discovered accidently during the evaluation of other apparent liver diseases, such as hepatitis C virus (HCV) infection or non-alcoholic fatty liver disease and, more importantly, their evolution into life-threatening hepatocellular carcinoma. As infection with HCV and occult HBV is rarely considered when assessing these more obvious conditions, and in an attempt to offer a better understanding of this phenomenon, this study attempted to shed some light onto the uniqueness of occult HBV infection by addressing the natural history of HBV and HCV infections, as well as non-alcoholic fatty liver disease. This was carried out by taking into account the exclusive integration process undertaken by the HBV genome into infected host hepatocytes, with consideration given to conditions which afford reactivation of the occult infection and stress on the molecular mechanisms that underlie occult HBV infection. Finally, the clinical outcome of occult HBV infection and its relation to hepatocellular carcinoma is analyzed.
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Affiliation(s)
- Hatem Elalfy
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Tarek Besheer
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Dina Elhammady
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Ahmed El Mesery
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Shaker Wagih Shaltout
- Tropical Medicine Department, Faculty of Medicine, Port Said University, Port Said 42511, Egypt
| | - Mohamed Abd El-Maksoud
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Ahmed I Amin
- Internal Medicine Department, Faculty of Medicine, Port Said University, Port Said 42511, Egypt
| | - Ahmed Nasr Bekhit
- Tropical Medicine Department, Zagazig General Hospital, Zagazig 44511, Egypt
| | - Mahmoud Abd El Aziz
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
| | - Mahmoud El-Bendary
- Endemic Medicine Department, Faculty of Medicine, Mansoura University, Mansoura 35516, Egypt
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17
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Bollinger RC, Thio CL, Sulkowski MS, McKenzie-White J, Thomas DL, Flexner C. Addressing the global burden of hepatitis B virus while developing long-acting injectables for the prevention and treatment of HIV. Lancet HIV 2019; 7:e443-e448. [PMID: 31870675 DOI: 10.1016/s2352-3018(19)30342-x] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2019] [Revised: 08/05/2019] [Accepted: 08/13/2019] [Indexed: 12/23/2022]
Abstract
The first long-acting formulations of HIV drugs are undergoing regulatory review for use in maintenance of viral suppression in people with HIV. Although these novel drug formulations could contribute greatly to HIV treatment and prevention efforts, their lack of activity against hepatitis B virus (HBV) could limit their global impact, particularly in populations with high burdens of both HIV and HBV. An urgent need for greater investment in research and development of long-acting drugs with dual activity against HIV and HBV exists. Access to long-acting HIV drug formulations with dual activity against HBV would be transformative and have a great impact on efforts to prevent, treat, and eradicate both of these important global epidemics.
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Affiliation(s)
| | - Chloe L Thio
- School of Medicine, Johns Hopkins University, Baltimore, MD, USA
| | - Mark S Sulkowski
- School of Medicine, Johns Hopkins University, Baltimore, MD, USA
| | | | - David L Thomas
- School of Medicine, Johns Hopkins University, Baltimore, MD, USA
| | - Charles Flexner
- School of Medicine, Johns Hopkins University, Baltimore, MD, USA
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18
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Raimondo G, Locarnini S, Pollicino T, Levrero M, Zoulim F, Lok AS. Update of the statements on biology and clinical impact of occult hepatitis B virus infection. J Hepatol 2019; 71:397-408. [PMID: 31004683 DOI: 10.1016/j.jhep.2019.03.034] [Citation(s) in RCA: 335] [Impact Index Per Article: 55.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/21/2019] [Revised: 03/20/2019] [Accepted: 03/28/2019] [Indexed: 02/06/2023]
Abstract
In October 2018 a large number of international experts with complementary expertise came together in Taormina to participate in a workshop on occult hepatitis B virus infection (OBI). The objectives of the workshop were to review the existing knowledge on OBI, to identify issues that require further investigation, to highlight both existing controversies and newly emerging perspectives, and ultimately to update the statements previously agreed in 2008. This paper represents the output from the workshop.
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Affiliation(s)
- Giovanni Raimondo
- Division of Clinical and Molecular Hepatology, University of Messina, Messina, Italy; Department of Clinical and Experimental Medicine, University of Messina, Messina, Italy.
| | - Stephen Locarnini
- Victorian Infectious Diseases Reference Laboratory at the Doherty Institute, Melbourne, Victoria, Australia
| | - Teresa Pollicino
- Division of Clinical and Molecular Hepatology, University of Messina, Messina, Italy; Department of Human Pathology, University of Messina, Messina, Italy
| | - Massimo Levrero
- Cancer Research Center of Lyon, INSERM U1052, Hospices Civils de Lyon, Lyon University, Lyon, France
| | - Fabien Zoulim
- Cancer Research Center of Lyon, INSERM U1052, Hospices Civils de Lyon, Lyon University, Lyon, France
| | - Anna S Lok
- Division of Gastroenterology and Hepatology, University of Michigan, Ann Arbor, MI, USA
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19
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Molecular Characterization of Near Full-Length Genomes of Hepatitis B Virus Isolated from Predominantly HIV Infected Individuals in Botswana. Genes (Basel) 2018; 9:genes9090453. [PMID: 30205537 PMCID: PMC6162474 DOI: 10.3390/genes9090453] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2018] [Revised: 08/27/2018] [Accepted: 09/03/2018] [Indexed: 12/13/2022] Open
Abstract
The World Health Organization plans to eliminate hepatitis B and C Infections by 2030. Therefore, there is a need to study and understand hepatitis B virus (HBV) epidemiology and viral evolution further, including evaluating occult (HBsAg-negative) HBV infection (OBI), given that such infections are frequently undiagnosed and rarely treated. We aimed to molecularly characterize HBV genomes from 108 individuals co-infected with human immunodeficiency virus (HIV) and chronic hepatitis B (CHB) or OBI identified from previous HIV studies conducted in Botswana from 2009 to 2012. Full-length (3.2 kb) and nearly full-length (~3 kb) genomes were amplified by nested polymerase chain reaction (PCR). Sequences from OBI participants were compared to sequences from CHB participants and GenBank references to identify OBI-unique mutations. HBV genomes from 50 (25 CHB and 25 OBI) individuals were successfully genotyped. Among OBI participants, subgenotype A1 was identified in 12 (48%), D3 in 12 (48%), and E in 1 (4%). A similar genotype distribution was observed in CHB participants. Whole HBV genome sequences from Botswana, representing OBI and CHB, were compared for the first time. There were 43 OBI-unique mutations, of which 26 were novel. Future studies using larger sample sizes and functional analysis of OBI-unique mutations are warranted.
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20
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Anderson M, Choga WT, Moyo S, Bell TG, Mbangiwa T, Phinius BB, Bhebhe L, Sebunya TK, Makhema J, Marlink R, Kramvis A, Essex M, Musonda RM, Blackard JT, Gaseitsiwe S. In Silico Analysis of Hepatitis B Virus Occult Associated Mutations in Botswana Using a Novel Algorithm. Genes (Basel) 2018; 9:genes9090420. [PMID: 30134551 PMCID: PMC6162659 DOI: 10.3390/genes9090420] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2018] [Accepted: 08/16/2018] [Indexed: 02/06/2023] Open
Abstract
Occult hepatitis B infections (OBI) represent a reservoir of undiagnosed and untreated hepatitis B virus (HBV), hence the need to identify mutations that lead to this phenotype. Functionally characterizing these mutations by in vitro studies is time-consuming and expensive. To bridge this gap, in silico approaches, which predict the effect of amino acid (aa) variants on HBV protein function, are necessary. We developed an algorithm for determining the relevance of OBI-associated mutations using in silico approaches. A 3 kb fragment of subgenotypes A1 and D3 from 24 chronic HBV-infected (CHB) and 24 OBI participants was analyzed. To develop and validate the algorithm, the effects of 68 previously characterized occult-associated mutations were determined using three computational tools: PolyPhen2, SNAP2, and PROVEAN. The percentage of deleterious mutations (with impact on protein function) predicted were 52 (76.5%) by PolyPhen2, 55 (80.9%) by SNAP2, and 65 (95.6%) by PROVEAN. At least two tools correctly predicted 59 (86.8%) mutations as deleterious. To identify OBI-associated mutations exclusive to Botswana, study sequences were compared to CHB sequences from GenBank. Of the 43 OBI-associated mutations identified, 26 (60.5%) were predicted by at least two tools to have an impact on protein function. To our knowledge, this is the first study to use in silico approaches to determine the impact of OBI-associated mutations, thereby identifying potential candidates for functional analysis to facilitate mechanistic studies of the OBI phenotype.
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Affiliation(s)
- Motswedi Anderson
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Faculty of Science, Department of Biological Sciences, University of Botswana, Gaborone, Botswana.
| | | | - Sikhulile Moyo
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
| | - Trevor Graham Bell
- Hepatitis Virus Diversity Research Unit (HVDRU), Faculty of Health Sciences, Department of Internal Medicine, School of Clinical Medicine, University of the Witwatersrand, Johannesburg 2050, South Africa.
| | - Tshepiso Mbangiwa
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Faculty of Allied Health Sciences, University of Botswana, Gaborone, Botswana.
| | - Bonolo B Phinius
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
| | - Lynette Bhebhe
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
| | - Theresa K Sebunya
- Faculty of Science, Department of Biological Sciences, University of Botswana, Gaborone, Botswana.
| | - Joseph Makhema
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
| | - Richard Marlink
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
- Rutgers Global Health Institute, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ 08854, USA.
| | - Anna Kramvis
- Hepatitis Virus Diversity Research Unit (HVDRU), Faculty of Health Sciences, Department of Internal Medicine, School of Clinical Medicine, University of the Witwatersrand, Johannesburg 2050, South Africa.
| | - Max Essex
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
| | | | - Jason T Blackard
- College of Medicine, University of Cincinnati, Cincinnati, OH 45267, USA.
| | - Simani Gaseitsiwe
- Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana.
- Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
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21
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Kostaki EG, Karamitros T, Stefanou G, Mamais I, Angelis K, Hatzakis A, Kramvis A, Paraskevis D. Unravelling the history of hepatitis B virus genotypes A and D infection using a full-genome phylogenetic and phylogeographic approach. eLife 2018; 7:36709. [PMID: 30082021 PMCID: PMC6118819 DOI: 10.7554/elife.36709] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2018] [Accepted: 07/28/2018] [Indexed: 12/17/2022] Open
Abstract
Hepatitis B virus (HBV) infection constitutes a global public health problem. In order to establish how HBV was disseminated across different geographic regions, we estimated the levels of regional clustering for genotypes D and A. We used 916 HBV-D and 493 HBV-A full-length sequences to reconstruct their global phylogeny. Phylogeographic analysis was conducted by the reconstruction of ancestral states using the criterion of parsimony. The putative origin of genotype D was in North Africa/Middle East. HBV-D sequences form low levels of regional clustering for the Middle East and Southern Europe. In contrast, HBV-A sequences form two major clusters, the first including sequences mostly from sub-Saharan Africa, and the second including sequences mostly from Western and Central Europe. Conclusion: We observed considerable differences in the global dissemination patterns of HBV-D and HBV-A and different levels of monophyletic clustering in relation to the regions of prevalence of each genotype.
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Affiliation(s)
- Evangelia-Georgia Kostaki
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
| | - Timokratis Karamitros
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
- Department of ZoologyUniversity of OxfordOxfordUnited Kingdom
| | - Garyfallia Stefanou
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
| | - Ioannis Mamais
- Department of Health Sciences, School of SciencesEuropean University of CyprusNicosiaCyprus
| | - Konstantinos Angelis
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
| | - Angelos Hatzakis
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
| | - Anna Kramvis
- Hepatitis Virus Diversity Research Unit, Department of Internal Medicine, Faculty of Health ScienceUniversity of the WitwatersrandJohannesburgSouth Africa
| | - Dimitrios Paraskevis
- Department of Hygiene, Epidemiology and Medical Statistics, Medical SchoolNational and Kapodistrian University of AthensAthensGreece
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22
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Blackard JT, Sherman KE. Hepatitis B virus (HBV) reactivation-The potential role of direct-acting agents for hepatitis C virus (HCV). Rev Med Virol 2018; 28:e1984. [PMID: 29761585 PMCID: PMC6233878 DOI: 10.1002/rmv.1984] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2017] [Revised: 04/04/2018] [Accepted: 04/05/2018] [Indexed: 12/16/2022]
Abstract
Hepatitis C virus (HCV) is known to inhibit hepatitis B virus (HBV) replication in patients with HBV/HCV coinfection. Reactivation of HBV in patients treated for HCV with direct-acting agents (DAAs) has emerged recently as an important clinical consideration. A growing number of case reports and case series support the association between new HCV treatments and HBV reactivation. Yet, very little is known about the specific viral characteristics that facilitate reactivation as functional characterization of the reactivated HBV has been conducted only rarely. This review provides the most recent data on HBV reactivation in the context of DAA initiation and highlights the existing viral genomic data from reactivating viruses. Current functional studies of HBV reactivation are largely limited by the retrospective identification of cases, no standardization of genomic regions that are studied with respect to HBV reactivation, and the lack of inclusion of nonreactivating controls to establish specific viral mutations that are associated with HBV reactivation. Importantly, none of these sequencing studies included cases of HBV reactivation after initiation of DAAs. While new HCV treatments have revolutionized care for HCV infected patients, HBV reactivation will likely increase in frequency, as DAAs are more commonly prescribed. Pretreatment determination of HBV status and thoughtful management of HBV coinfections will be necessary and lead to improved patient safety and yield optimal treatment results.
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Affiliation(s)
- Jason T Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
| | - Kenneth E Sherman
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
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23
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Hatazawa Y, Yano Y, Okada R, Tanahashi T, Hayashi H, Hirano H, Minami A, Kawano Y, Tanaka M, Fukumoto T, Murakami Y, Yoshida M, Hayashi Y. Quasispecies variant of pre-S/S gene in HBV-related hepatocellular carcinoma with HBs antigen positive and occult infection. Infect Agent Cancer 2018; 13:7. [PMID: 29434654 PMCID: PMC5797373 DOI: 10.1186/s13027-018-0179-4] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2017] [Accepted: 01/26/2018] [Indexed: 12/15/2022] Open
Abstract
Background Hepatocellular carcinoma (HCC) can develop in patients who are negative for the hepatitis B surface antigen (HBsAg) in serum but positive for hepatitis B virus (HBV) DNA in the liver, referred to as occult HBV infection (OBI). Previous reports showed that HBV variants in OBI-related HCC are different from those in HBsAg-positive HCC. In the present study, HBV quasispecies based on the pre-S/S gene in OBI-related HCC patients were examined by high throughput sequencing and compared with those in HBsAg-positive HCC. Methods Nineteen tissue samples (9 OBI-related and 10 HBsAg-positive non-cancerous tissues) were collected at the time of surgery at Kobe University Hospital. The quasispecies with more than 1% variation in the pre-S/S region were isolated and analysed by ultra-deep sequencing. Results There were no significant differences in the major HBV populations, which exhibit more than 20% variation within the entire pre-S/S region, between OBI-related HCC and HBsAg-positive HCC. However, the prevalences of major populations with pre-S2 region mutations and of minor populations with polymerized human serum albumin-binding domain mutations were significantly higher in OBI-related HCC than in HBsAg-positive HCC. Moreover, the major variant populations associated with the B-cell epitope, located within the pre-S1 region, and the a determinant domain, located in the S region, were detected frequently in HBsAg-positive HCC. The minor populations of variants harbouring the W4R, L30S, Q118R/Stop, N123D and S124F/P mutations in the pre-S region and the L21F/S and L42F/S mutations in the S region were detected more frequently in OBI-related HCC than in HBsAg-positive HCC. Conclusions Ultra-deep sequencing revealed that the B-cell epitope domain in the pre-S1 region and alpha determinant domain in the S region were variable in HBsAg-positive HCC, although the quasispecies associated with the pre-S2 region were highly prevalent in OBI-related HCC. Trial registration Ref: R000034382/UMIN000030113; Retrospectively registered 25 November 2017.
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Affiliation(s)
- Yuri Hatazawa
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Yoshihiko Yano
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan.,2Division of Molecular Medicine & Medical Genetics, Department of Pathology, Kobe University Graduate School of Medicine, Kobe, Japan
| | - Rina Okada
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Toshihito Tanahashi
- Department of Internal Medicine, Tokushima Prefectural Naruto Hospital, Tokushima, Japan
| | - Hiroki Hayashi
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Hirotaka Hirano
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Akihiro Minami
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Yuki Kawano
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Motofumi Tanaka
- 4Division of Hepato-Biliary-Pancreatic Surgery, Department of Surgery, Kobe University Graduate School of Medicine, Kobe, Japan
| | - Takumi Fukumoto
- 4Division of Hepato-Biliary-Pancreatic Surgery, Department of Surgery, Kobe University Graduate School of Medicine, Kobe, Japan
| | - Yoshiki Murakami
- 5Department of Hepatology, Osaka City University Graduate School of Medicine, Osaka, Japan
| | - Masaru Yoshida
- 1Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo 650-0017 Japan
| | - Yoshitake Hayashi
- 2Division of Molecular Medicine & Medical Genetics, Department of Pathology, Kobe University Graduate School of Medicine, Kobe, Japan
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24
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Zhou T, Block T, Liu F, Kondratowicz AS, Sun L, Rawat S, Branson J, Guo F, Steuer HM, Liang H, Bailey L, Moore C, Wang X, Cuconatti A, Gao M, Lee ACH, Harasym T, Chiu T, Gotchev D, Dorsey B, Rijnbrand R, Sofia MJ. HBsAg mRNA degradation induced by a dihydroquinolizinone compound depends on the HBV posttranscriptional regulatory element. Antiviral Res 2017; 149:191-201. [PMID: 29133129 DOI: 10.1016/j.antiviral.2017.11.009] [Citation(s) in RCA: 40] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2017] [Revised: 11/01/2017] [Accepted: 11/07/2017] [Indexed: 12/23/2022]
Abstract
In pursuit of novel therapeutics targeting the hepatitis B virus (HBV) infection, we evaluated a dihydroquinolizinone compound (DHQ-1) that in the nanomolar range reduced the production of virion and surface protein (HBsAg) in tissue culture. This compound also showed broad HBV genotype coverage, but was inactive against a panel of DNA and RNA viruses of other species. Oral administration of DHQ-1 in the AAV-HBV mouse model resulted in a significant reduction of serum HBsAg as soon as 4 days following the commencement of treatment. Reduction of HBV markers in both in vitro and in vivo experiments was related to the reduced amount of viral RNA including pre-genomic RNA (pgRNA) and 2.4/2.1 kb HBsAg mRNA. Nuclear run-on and subcellular fractionation experiments indicated that DHQ-1 mediated HBV RNA reduction was the result of accelerated viral RNA degradation in the nucleus, rather than the consequence of inhibition of transcription initiation. Through mutagenesis of HBsAg gene sequences, we found induction of HBsAg mRNA decay by DHQ-1 required the presence of the HBV posttranscriptional regulatory element (HPRE), with a 109 nucleotides sequence within the central region of the HPRE alpha sub-element being the most critical. Taken together, the current study shows that a small molecule can reduce the overall levels of HBV RNA, especially the HBsAg mRNA, and viral surface proteins. This may shed light on the development of a new class of HBV therapeutics.
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Affiliation(s)
- Tianlun Zhou
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States.
| | - Timothy Block
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States
| | - Fei Liu
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Andrew S Kondratowicz
- Arbutus BioPharma, 100 - 8900 Glenlyon Parkway, Burnaby, British Columbia V5J 5J8, Canada
| | - Liren Sun
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States
| | - Siddhartha Rawat
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States
| | - Jeffrey Branson
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States
| | - Fang Guo
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | | | - Hongyan Liang
- Baruch S. Blumberg Institute, Department of Translational Medicine, Doylestown, PA 18902, United States
| | - Lauren Bailey
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Chris Moore
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Xiaohe Wang
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Andy Cuconatti
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Min Gao
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Amy C H Lee
- Arbutus BioPharma, 100 - 8900 Glenlyon Parkway, Burnaby, British Columbia V5J 5J8, Canada
| | - Troy Harasym
- Arbutus BioPharma, 100 - 8900 Glenlyon Parkway, Burnaby, British Columbia V5J 5J8, Canada
| | - Tim Chiu
- Arbutus BioPharma, 100 - 8900 Glenlyon Parkway, Burnaby, British Columbia V5J 5J8, Canada
| | - Dimitar Gotchev
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Bruce Dorsey
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Rene Rijnbrand
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States
| | - Michael J Sofia
- Arbutus BioPharma, 701 Veterans Circle, Warminster, PA 18974, United States.
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25
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Kim MH, Kang SY, Lee WI. Occult HBV among Anti-HBc Alone: Mutation Analysis of an HBV Surface Gene and Pre-S Gene. Yonsei Med J 2017; 58:557-563. [PMID: 28332361 PMCID: PMC5368141 DOI: 10.3349/ymj.2017.58.3.557] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/29/2016] [Revised: 12/23/2016] [Accepted: 01/26/2017] [Indexed: 12/14/2022] Open
Abstract
PURPOSE The aim of this study is to investigate the molecular characteristics of occult hepatitis B virus (HBV) infection in 'anti-HBc alone' subjects. MATERIALS AND METHODS Twenty-four patients with 'anti-HBc alone' and 20 control patients diagnosed with HBV were analyzed regarding S and pre-S gene mutations. All specimens were analyzed for HBs Ag, anti-HBc, and anti-HBs. For specimens with an anti-HBc alone, quantitative analysis of HBV DNA, as well as sequencing and mutation analysis of S and pre-S genes, were performed. RESULTS A total 24 were analyzed for the S gene, and 14 were analyzed for the pre-S gene through sequencing. A total of 20 control patients were analyzed for S and pre-S gene simultaneously. Nineteen point mutations of the major hydrophilic region were found in six of 24 patients. Among them, three mutations, S114T, P127S/T, M133T, were detected in common. Only one mutation was found in five subjects of the control group; this mutation was not found in the occult HBV infection group, however. Pre-S mutations were detected in 10 patients, and mutations of site aa58-aa100 were detected in 9 patients. A mutation on D114E was simultaneously detected. Although five mutations from the control group were found at the same location (aa58-aa100), no mutations of occult HBV infection were detected. CONCLUSION The prevalence of occult HBV infection is not low among 'anti-HBc alone' subjects. Variable mutations in the S gene and pre-S gene were associated with the occurrence of occult HBV infection. Further larger scale studies are required to determine the significance of newly detected mutations.
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Affiliation(s)
- Myeong Hee Kim
- Department of Laboratory Medicine, Kyung Hee University School of Medicine and Kyung Hee University Hospital at Gangdong, Seoul, Korea
| | - So Young Kang
- Department of Laboratory Medicine, Kyung Hee University School of Medicine and Kyung Hee University Hospital at Gangdong, Seoul, Korea.
| | - Woo In Lee
- Department of Laboratory Medicine, Kyung Hee University School of Medicine and Kyung Hee University Hospital at Gangdong, Seoul, Korea
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26
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Hossain MG, Ueda K. Investigation of a Novel Hepatitis B Virus Surface Antigen (HBsAg) Escape Mutant Affecting Immunogenicity. PLoS One 2017; 12:e0167871. [PMID: 28045894 PMCID: PMC5207502 DOI: 10.1371/journal.pone.0167871] [Citation(s) in RCA: 33] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2016] [Accepted: 11/21/2016] [Indexed: 02/07/2023] Open
Abstract
Mutation in the hepatitis B virus surface antigen (HBsAg) may affect the efficiency of diagnostic immunoassays or success of vaccinations using HBsAg. Thus, antigenicity and immunogenicity analyses of the mutated HBsAg are necessary to develop novel diagnostic tools and efficient vaccinations. Here, the in vitro antigenicity of three wild-type HBsAg open reading frames (ORFs) (adr4, W1S [subtype adr] and W3S [subtype adr]) isolated from clinically infected patients and nineteen synthesized single/double/multiple amino acid-substituted mutants were tested with commercial ELISA kits. Immunofluorescence staining of transfected cells and Western blot analysis confirmed that these ORFs were expressed at comparable levels in HEK-293 cells. W1S and adr4 were clearly detected, whereas W3S could not be detected. Using the same commercial immunoassay kit, we found that the single mutants, K120P and D123T, were marginally reactive, whereas W3S-aW1S and the double mutant, K120P/D123T, exhibited antigenicity roughly equivalent to the wild-type wako1S. On the other hand, the single mutants of W1S, P120K and T123D, significantly impaired the reactivity, while W1S-aW3S and the double mutant of W1S, P120K/T123D, resulted in a complete loss of antigenicity. In addition, ELISA revealed reduced HBs antigenicity of two mutants, W1S N146G and W1S Q129R/G145R. These commercial ELISA-based antigenic reactivities of HBsAg were also strongly correlated with the predicted Ai alterations of affected amino acids due to the specific mutation. In conclusion, this study showed for the first time that lysine (K120) and aspartate (D123) simultaneously affected HBsAg antigenicity, leading to diagnostic failure. These findings will improve diagnostic assays and vaccine development.
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Affiliation(s)
- Md. Golzar Hossain
- Division of Virology, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, Suita, Osaka, Japan
| | - Keiji Ueda
- Division of Virology, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, Suita, Osaka, Japan
- * E-mail:
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27
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Makvandi M. Update on occult hepatitis B virus infection. World J Gastroenterol 2016; 22:8720-8734. [PMID: 27818588 PMCID: PMC5075547 DOI: 10.3748/wjg.v22.i39.8720] [Citation(s) in RCA: 105] [Impact Index Per Article: 11.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/28/2016] [Revised: 06/13/2016] [Accepted: 07/20/2016] [Indexed: 02/06/2023] Open
Abstract
The event of mutations in the surface antigen gene of hepatitis B virus (HBV) results in undetectable hepatitis B surface antigen with positive/negative anti-hepatitis B core (anti-HBc) antibody status in serum and this phenomenon is named occult hepatitis B infection (OBI). The presence of anti-HBc antibody in serum is an important key for OBI tracking, although about 20% of OBI cases are negative for anti-HBc antibody. The diagnosis of OBI is mainly based on polymerase chain reaction (PCR) and real-time PCR assays. However, real-time PCR is a more reliable method than PCR. OBI is a great issue for the public health problem and a challenge for the clinical entity worldwide. The persistence of OBI may lead to the development of cirrhosis and hepatocellular carcinoma. With regard to OBI complications, the screening of HBV DNA by the highly sensitive molecular means should be implemented for: (1) patients with a previous history of chronic or acute HBV infection; (2) patients co-infected with hepatitis C virus/human immunodeficiency virus; (3) patients undergoing chemotherapy or anti-CD20 therapy; (4) recipients of organ transplant; (5) blood donors; (6) organ transplant donors; (7) thalassemia and hemophilia patients; (8) health care workers; (9) patients with liver related disease (cryptogenic); (10) hemodialysis patients; (11) patients undergoing lamivudine or interferon therapy; and (12) children in time of HBV vaccination especially in highly endemic areas of HBV. Active HBV vaccination should be implemented for the close relatives of patients who are negative for OBI markers. Thus, the goal of this review is to evaluate the rate of OBI with a focus on status of high risk groups in different regions of the world.
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28
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Hudu SA, Harmal NS, Saeed MI, Alshrari AS, Malik YA, Niazlin MT, Hassan R, Sekawi Z. Molecular and serological detection of occult hepatitis B virus among healthy hepatitis B surface antigen-negative blood donors in Malaysia. Afr Health Sci 2016; 16:677-683. [PMID: 27917199 DOI: 10.4314/ahs.v16i3.6] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
BACKGROUND Occult hepatitis B infections are becoming a major global threat, but the available data on its prevalence in various parts of the world are often divergent. OBJECTIVE This study aimed to detect occult hepatitis B virus in hepatitis B surface antigen-negative serum using anti-HBc as a marker of previous infection. PATIENT AND METHODS A total of 1000 randomly selected hepatitis B surface antigen-negative sera from blood donors were tested for hepatitis B core antibody and hepatitis B surface antibody using an ELISA and nested polymerase chain reaction was done using primers specific to the surface gene (S-gene). RESULTS Of the 1000 samples 55 (5.5%) were found to be reactive, of which 87.3% (48/55) were positive for hepatitis B surface antibody, indicating immunity as a result of previous infection however, that does not exclude active infection with escaped mutant HBV. Nested PCR results showed the presence of hepatitis B viral DNA in all the 55 samples that were positive for core protein, which is in agreement with the hepatitis B surface antibody result. CONCLUSION This study reveals the 5.5% prevalence of occult hepatitis B among Malaysian blood donors as well as the reliability of using hepatitis B core antibody in screening for occult hepatitis B infection in low endemic, low socioeconomic settings.
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Affiliation(s)
- Shuaibu A Hudu
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia; Medical Microbiology and Parasitology, Faculty of Basic Medical Sciences, College of Health Sciences, Usmanu Danfodiyo University Sokoto, 840232 Sokoto State, Nigeria
| | - Nabil S Harmal
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia; Department of Medical Microbiology, Faculty of Medicine and Health Sciences, Sana'a University, Sana'a, Yemen
| | - Mohammed I Saeed
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia
| | - Ahmad S Alshrari
- Department of Basic Health Sciences, Faculty of Pharmacy, Northern Border Universiti, 91911 Rafha, Saudi Arabia
| | - Yasmin A Malik
- Department of Clinical Science, Faculty of Medicine and Health Sciences, Universiti Tunku Abdul Rahman, Malaysia
| | - Mohd T Niazlin
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia
| | - Roshida Hassan
- National Blood Centre Malaysia, Jalan Tun Razak Kuala Lumpur, 504000 Malaysia
| | - Zamberi Sekawi
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia
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29
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Zhu HL, Li X, Li J, Zhang ZH. Genetic variation of occult hepatitis B virus infection. World J Gastroenterol 2016; 22:3531-3546. [PMID: 27053845 PMCID: PMC4814639 DOI: 10.3748/wjg.v22.i13.3531] [Citation(s) in RCA: 60] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/29/2015] [Revised: 12/13/2015] [Accepted: 12/30/2015] [Indexed: 02/06/2023] Open
Abstract
Occult hepatitis B virus infection (OBI), characterized as the persistence of hepatitis B virus (HBV) surface antigen (HBsAg) seronegativity and low viral load in blood or liver, is a special form of HBV infection. OBI may be related mainly to mutations in the HBV genome, although the underlying mechanism of it remains to be clarified. Mutations especially within the immunodominant “α” determinant of S protein are “hot spots” that could contribute to the occurrence of OBI via affecting antigenicity and immunogenicity of HBsAg or replication and secretion of virion. Clinical reports account for a large proportion of previous studies on OBI, while functional analyses, especially those based on full-length HBV genome, are rare.
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30
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Lim CS, Brown CM. Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element. RNA Biol 2016; 13:743-7. [PMID: 27031749 DOI: 10.1080/15476286.2016.1166330] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel.
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Affiliation(s)
- Chun Shen Lim
- a Biochemistry and Genetics Otago , University of Otago , Dunedin , New Zealand
| | - Chris M Brown
- a Biochemistry and Genetics Otago , University of Otago , Dunedin , New Zealand
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Powell EA, Boyce CL, Gededzha MP, Selabe SG, Mphahlele MJ, Blackard JT. Functional analysis of 'a' determinant mutations associated with occult HBV in HIV-positive South Africans. J Gen Virol 2016; 97:1615-1624. [PMID: 27031988 DOI: 10.1099/jgv.0.000469] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Occult hepatitis B is defined by the presence of hepatitis B virus (HBV) DNA in the absence of hepatitis B surface antigen (HBsAg). Occult HBV is associated with the development of hepatocellular carcinoma, reactivation during immune suppression, and virus transmission. Viral mutations contribute significantly to the occult HBV phenotype. Mutations in the 'a' determinant of HBsAg are of particular interest, as these mutations are associated with immune escape, vaccine escape and diagnostic failure. We examined the effects of selected occult HBV-associated mutations identified in a population of HIV-positive South Africans on HBsAg production in vitro. Mutations were inserted into two different chronic HBV backbones and transfected into a hepatocyte-derived cell line. HBsAg levels were quantified by enzyme-linked immunosorbent assay (ELISA), while the detectability of mutant HBsAg was determined using an HA-tagged HBsAg expression system. Of the seven mutations analysed, four (S132P, C138Y, N146D and C147Y) resulted in decreased HBsAg expression in one viral background but not in the second viral background. One mutation (N146D) led to a decrease in HBsAg detected as compared to HA-tag, indicating that this mutation compromises the ability of the ELISA to detect HBsAg. The contribution of occult-associated mutations to the HBsAg-negative phenotype of occult HBV cannot be determined adequately by testing the effect of the mutation in a single viral background, and rigorous analysis of these mutations is required.
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Affiliation(s)
| | | | | | | | - M Jeffrey Mphahlele
- Sefako Makgatho Health Sciences University; South Africa Medical Research Council
| | - Jason T Blackard
- Division of Digestive Diseases ML 0595 231 Albert Sabin Way, University of Cincinnati, University of Cincinnati College of Medicine, USA
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Sarin SK, Kumar M, Lau GK, Abbas Z, Chan HLY, Chen CJ, Chen DS, Chen HL, Chen PJ, Chien RN, Dokmeci AK, Gane E, Hou JL, Jafri W, Jia J, Kim JH, Lai CL, Lee HC, Lim SG, Liu CJ, Locarnini S, Al Mahtab M, Mohamed R, Omata M, Park J, Piratvisuth T, Sharma BC, Sollano J, Wang FS, Wei L, Yuen MF, Zheng SS, Kao JH. Asian-Pacific clinical practice guidelines on the management of hepatitis B: a 2015 update. Hepatol Int 2016; 10:1-98. [PMID: 26563120 PMCID: PMC4722087 DOI: 10.1007/s12072-015-9675-4] [Citation(s) in RCA: 1883] [Impact Index Per Article: 209.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/20/2015] [Accepted: 09/14/2015] [Indexed: 02/06/2023]
Abstract
Worldwide, some 240 million people have chronic hepatitis B virus (HBV), with the highest rates of infection in Africa and Asia. Our understanding of the natural history of HBV infection and the potential for therapy of the resultant disease is continuously improving. New data have become available since the previous APASL guidelines for management of HBV infection were published in 2012. The objective of this manuscript is to update the recommendations for the optimal management of chronic HBV infection. The 2015 guidelines were developed by a panel of Asian experts chosen by the APASL. The clinical practice guidelines are based on evidence from existing publications or, if evidence was unavailable, on the experts' personal experience and opinion after deliberations. Manuscripts and abstracts of important meetings published through January 2015 have been evaluated. This guideline covers the full spectrum of care of patients infected with hepatitis B, including new terminology, natural history, screening, vaccination, counseling, diagnosis, assessment of the stage of liver disease, the indications, timing, choice and duration of single or combination of antiviral drugs, screening for HCC, management in special situations like childhood, pregnancy, coinfections, renal impairment and pre- and post-liver transplant, and policy guidelines. However, areas of uncertainty still exist, and clinicians, patients, and public health authorities must therefore continue to make choices on the basis of the evolving evidence. The final clinical practice guidelines and recommendations are presented here, along with the relevant background information.
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Affiliation(s)
- S K Sarin
- Department of Hepatology, Institute of Liver and Biliary Sciences, New Delhi, India.
| | - M Kumar
- Department of Hepatology, Institute of Liver and Biliary Sciences, New Delhi, India
| | - G K Lau
- Division of Gastroenterology and Hepatology, Humanity and Health Medical Centre, Hong Kong SAR, China
- The Institute of Translational Hepatology, Beijing, China
| | - Z Abbas
- Department of Hepatogastroenterlogy, Sindh Institute of Urology and Transplantation, Karachi, Pakistan
| | - H L Y Chan
- Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong, China
| | - C J Chen
- Genomics Research Center, Academia Sinica, National Taiwan University, Taipei, Taiwan
| | - D S Chen
- Department of Internal Medicine, National Taiwan University College of Medicine, Taipei, Taiwan
| | - H L Chen
- Graduate Institute of Clinical Medicine, National Taiwan University College of Medicine, Taipei, Taiwan
| | - P J Chen
- Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan
| | - R N Chien
- Liver Research Unit, Chang Gung Memorial Hospital and University, Chilung, Taiwan
| | - A K Dokmeci
- Department of Gastroenterology, Ankara University School of Medicine, Ankara, Turkey
| | - Ed Gane
- New Zealand Liver Transplant Unit, Auckland City Hospital, Auckland, New Zealand
| | - J L Hou
- Department of Infectious Diseases and Hepatology Unit, Nanfang Hospital, Guangzhou, China
| | - W Jafri
- Department of Medicine, Aga Khan University, Karachi, Pakistan
| | - J Jia
- Beijing Friendship Hospital, Capital Medical University, Beijing, China
| | | | - C L Lai
- Department of Medicine, University of Hong Kong, Hong Kong, China
| | - H C Lee
- Internal Medicine Asan Medical Center, Seoul, Korea
| | - S G Lim
- Division of Gastroenterology and Hepatology, National University Health System, Singapore, Singapore
| | - C J Liu
- Graduate Institute of Clinical Medicine, National Taiwan University College of Medicine, Taipei, Taiwan
| | - S Locarnini
- Research and Molecular Development, Victorian Infectious Diseases Reference Laboratory, Melbourne, Australia
| | - M Al Mahtab
- Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh
| | - R Mohamed
- Department of Medicine, Faculty of Medicine, University Malaya, Kuala Lumpur, Malaysia
| | - M Omata
- Yamanashi Hospitals (Central and Kita) Organization, 1-1-1 Fujimi, Kofu-shi, Yamanashi, 400-8506, Japan
| | - J Park
- Department of Internal Medicine, Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, Korea
| | - T Piratvisuth
- NKC Institute of Gastroenterology and Hepatology, Prince of Songkla University, Songkhla, Thailand
| | - B C Sharma
- Department of Gastroenterology, G.B. Pant Hospital, New Delhi, India
| | - J Sollano
- Department of Medicine, University of Santo Tomas, Manila, Philippines
| | - F S Wang
- Treatment and Research Center for Infectious Diseases, Beijing 302 Hospital, Beijing, China
| | - L Wei
- Peking University Hepatology Institute, Beijing, China
| | - M F Yuen
- Division of Gastroenterology and Hepatology, Department of Medicine, University of Hong Kong, Pofulam, Hong Kong
| | - S S Zheng
- Department of Hepatobiliary and Pancreatic Surgery, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, 310003, Zhejiang Province, China
| | - J H Kao
- Graduate Institute of Clinical Medicine and Hepatitis Research Center, National Taiwan University College of Medicine, National Taiwan University Hospital, Taipei, Taiwan
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Spreafico M, Berzuini A, Foglieni B, Candotti D, Raffaele L, Guarnori I, Colli A, Maldini FF, Allain JP, Prati D. Poor efficacy of nucleic acid testing in identifying occult HBV infection and consequences for safety of blood supply in Italy. J Hepatol 2015; 63:1068-76. [PMID: 26116791 DOI: 10.1016/j.jhep.2015.06.016] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/04/2015] [Revised: 04/30/2015] [Accepted: 06/16/2015] [Indexed: 01/04/2023]
Abstract
BACKGROUND & AIMS In Italy, DNA screening of blood donations for hepatitis B virus (HBV) was introduced to prevent the transmission of window period and occult HBV infection. Anti-HBc screening is not recommended in order to avoid shortage of the blood supply. To contain costs, donor samples are generally pooled before testing. We evaluated the safety of this national policy using a prospective repository of donors/recipient pairs. METHODS We used highly sensitive nucleic acid testing (NAT) assays to test repository and follow-up samples from donors who were initially classified as negative by minipool NAT assays (6-MP), but were later found to carry occult HBV DNA. When available, we also analysed recipients' pre- and post-transfusion samples, collected in the context of a repository financed by the European Commission (the BOTIA project). RESULTS Between 2008 and 2011 6-MP NAT assays identified 18 carriers of occult HBV infection among 12,695 donors; 28 samples from previous donations were available from 13 of these carriers. Highly sensitive HBV DNA detection methods showed that 6-MP HBV DNA screening failed to identify 14/28 (50%) viraemic donations, that were released for transfusion. HBV marker testing of such blood product recipients revealed two cases of transfusion transmitted HBV infection, documented by donor-recipient sequence identity. CONCLUSIONS Viraemic blood donations from occult HBV infection carriers remain undetected by current minipool HBV DNA screening, and transfusion transmission of HBV continues to occur in susceptible patients. More effective individual HBV DNA screening and/or tests for antibodies to HBV core antigen should be considered to improve blood safety.
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Affiliation(s)
- Marta Spreafico
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy.
| | - Alessandra Berzuini
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | - Barbara Foglieni
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | - Daniel Candotti
- Institut National de la Transfusion Sanguine (INTS), Département d'études des agents transmissibles, F-75015 Paris, France
| | - Livia Raffaele
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | - Irene Guarnori
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | - Agostino Colli
- Department of General Medicine, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | - Francesco Fumagalli Maldini
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy
| | | | - Daniele Prati
- Department of Transfusion Medicine and Hematology, Azienda Ospedaliera della Provincia di Lecco, Alessandro Manzoni Hospital, Lecco, Italy.
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Pondé RAA. Molecular mechanisms underlying HBsAg negativity in occult HBV infection. Eur J Clin Microbiol Infect Dis 2015; 34:1709-31. [PMID: 26105620 DOI: 10.1007/s10096-015-2422-x] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2015] [Accepted: 06/03/2015] [Indexed: 02/06/2023]
Abstract
Although genomic detection is considered the gold standard test on HBV infection identification, the HBsAg investigation is still the most frequent clinical laboratory request to diagnose HBV infection in activity. However, the non-detection of HBsAg in the bloodstream of chronic or acutely infected individuals has been a phenomenon often observed in clinical practice, despite the high sensitivity and specificity of screening assays standardized commercially and adopted in routine. The expansion of knowledge about the hepatitis B virus biology (replication/life cycle, genetic variability/mutability/heterogeneity), their biochemical and immunological properties (antigenicity and immunogenicity), in turn, has allowed to elucidate some mechanisms that may explain the occurrence of this phenomenon. Therefore, the negativity for HBsAg during the acute or chronic infection course may become a fragile or at least questionable result. This manuscript discusses some mechanisms that could explain the negativity for HBsAg in a serological profile of individuals with HBV infection in activity, or factors that could compromise its detection in the bloodstream during HBV infection.
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Affiliation(s)
- R A A Pondé
- Laboratory of Human Virology, Institute of Tropical Pathology and Public Health, Federal University of Goiás, Goiânia, Goiás, Brazil,
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Tandoi F, Caviglia GP, Pittaluga F, Abate ML, Smedile A, Romagnoli R, Salizzoni M. Prediction of occult hepatitis B virus infection in liver transplant donors through hepatitis B virus blood markers. Dig Liver Dis 2014; 46:1020-4. [PMID: 25201211 DOI: 10.1016/j.dld.2014.07.172] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/05/2014] [Revised: 07/25/2014] [Accepted: 07/26/2014] [Indexed: 12/11/2022]
Abstract
BACKGROUND Occult hepatitis B virus infection is defined as detectable HBV-DNA in liver of HBsAg-negative individuals, with or without detectable serum HBV-DNA. In deceased liver donors, results of tissue analysis cannot be obtained prior to allocation for liver transplantation. AIMS we investigated prevalence and predictability of occult hepatitis B using blood markers of viral exposure/infection in deceased liver donors. METHODS In 50 consecutive HBsAg-negative/anti-HBc-positive and 20 age-matched HBsAg-negative/anti-HBc-negative donors, a nested-PCR assay was employed in liver biopsies for diagnosis of occult hepatitis B according to Taormina criteria. All donors were characterized for plasma HBV-DNA and serum anti-HBs/anti-HBe. RESULTS In liver tissue, occult hepatitis B was present in 30/50 anti-HBc-positive (60%) and in 0/20 anti-HBc-negative donors (p<0.0001). All anti-HBc-positive donors with detectable HBV-DNA in plasma (n=5) or anti-HBs>1,000 mIU/mL (n=5) eventually showed occult infection, i.e, 10/30 occult hepatitis B-positive donors which could have been identified prior to transplantation. In the remaining 40 anti-HBc-positive donors, probability of occult infection was 62% for anti-HBe-positive and/or anti-HBs ≥ 58 mIU/mL; 29% for anti-HBe-negative and anti-HBs<58 mIU/mL. CONCLUSIONS In deceased donors, combining anti-HBc with other blood markers of hepatitis B exposure/infection allows to predict occult hepatitis B with certainty and speed in one third of cases. These findings might help refine the allocation of livers from anti-HBc-positive donors.
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Affiliation(s)
- Francesco Tandoi
- Liver Transplant Center, General Surgery Unit, Department of Surgical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
| | - Gian Paolo Caviglia
- Laboratory of Digestive and Liver Physiopathology, Department of Medical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
| | - Fabrizia Pittaluga
- Laboratory of Microbiology and Virology, Department of Laboratory Medicine, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
| | - Maria Lorena Abate
- Laboratory of Digestive and Liver Physiopathology, Department of Medical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
| | - Antonina Smedile
- Laboratory of Digestive and Liver Physiopathology, Department of Medical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
| | - Renato Romagnoli
- Liver Transplant Center, General Surgery Unit, Department of Surgical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy.
| | - Mauro Salizzoni
- Liver Transplant Center, General Surgery Unit, Department of Surgical Sciences, A.O.U. Città della Salute e della Scienza, Molinette Hospital, University of Turin, Turin, Italy
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Powell EA, Gededzha MP, Rentz M, Rakgole NJ, Selabe SG, Seleise TA, Mphahlele MJ, Blackard JT. Mutations associated with occult hepatitis B in HIV-positive South Africans. J Med Virol 2014; 87:388-400. [PMID: 25164924 DOI: 10.1002/jmv.24057] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 07/27/2014] [Indexed: 12/16/2022]
Abstract
Occult hepatitis B is characterized by the absence of hepatitis B surface antigen (HBsAg) but the presence of HBV DNA. Because diagnosis of hepatitis B virus (HBV) typically includes HBsAg detection, occult HBV remains largely undiagnosed. Occult HBV is associated with increased risk of hepatocellular carcinoma, reactivation to chronic HBV during immune suppression, and transmission during blood transfusion and liver transplant. The mechanisms leading to occult HBV infection are unclear, although viral mutations are likely a significant factor. In this study, sera from 394 HIV-positive South Africans were tested for HBV DNA and HBsAg. For patients with detectable HBV DNA, the overlapping surface and polymerase open reading frames (ORFs) were sequenced. Occult-associated mutations-those mutations found exclusively in individuals with occult HBV infection but not in individuals with chronic HBV infection from the same cohort or GenBank references-were identified. Ninety patients (22.8%) had detectable HBV DNA. Of these, 37 had detectable HBsAg, while 53 lacked detectable surface antigen. The surface and polymerase ORFs were cloned successfully for 19 patients with chronic HBV and 30 patients with occult HBV. In total, 235 occult-associated mutations were identified. Ten occult-associated mutations were identified in more than one patient. Additionally, 15 amino acid positions had two distinct occult-associated mutations at the same residue. Occult-associated mutations were common and present in all regions of the surface and polymerase ORFs. Further study is underway to determine the effects of these mutations on viral replication and surface antigen expression in vitro.
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Affiliation(s)
- Eleanor A Powell
- Division of Digestive Disease, University of Cincinnati College of Medicine, Cincinnati, Ohio
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Huang FY, Wong DKH, Seto WK, Zhang AY, Lee CK, Lin CK, Fung J, Lai CL, Yuen MF. Sequence variations of full-length hepatitis B virus genomes in Chinese patients with HBsAg-negative hepatitis B infection. PLoS One 2014; 9:e99028. [PMID: 24901840 PMCID: PMC4047052 DOI: 10.1371/journal.pone.0099028] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2014] [Accepted: 05/09/2014] [Indexed: 12/16/2022] Open
Abstract
BACKGROUND The underlying mechanism of HBsAg-negative hepatitis B virus (HBV) infection is notoriously difficult to elucidate because of the extremely low DNA levels which define the condition. We used a highly efficient amplification method to overcome this obstacle and achieved our aim which was to identify specific mutations or sequence variations associated with this entity. METHODS A total of 185 sera and 60 liver biopsies from HBsAg-negative, HBV DNA-positive subjects or known chronic hepatitis B (CHB) subjects with HBsAg seroclearance were amplified by rolling circle amplification followed by full-length HBV genome sequencing. Eleven HBsAg-positive CHB subjects were included as controls. The effects of pivotal mutations identified on regulatory regions on promoter activities were analyzed. RESULTS 22 and 11 full-length HBV genomes were amplified from HBsAg-negative and control subjects respectively. HBV genotype C was the dominant strain. A higher mutation frequency was observed in HBsAg-negative subjects than controls, irrespective of genotype. The nucleotide diversity over the entire HBV genome was significantly higher in HBsAg-negative subjects compared with controls (p = 0.008) and compared with 49 reference sequences from CHB patients (p = 0.025). In addition, HBsAg-negative subjects had significantly higher amino acid substitutions in the four viral genes than controls (all p<0.001). Many mutations were uniquely found in HBsAg-negative subjects, including deletions in promoter regions (13.6%), abolishment of pre-S2/S start codon (18.2%), disruption of pre-S2/S mRNA splicing site (4.5%), nucleotide duplications (9.1%), and missense mutations in "α" determinant region, contributing to defects in HBsAg production. CONCLUSIONS These data suggest an accumulation of multiple mutations constraining viral transcriptional activities contribute to HBsAg-negativity in HBV infection.
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Affiliation(s)
- Fung-Yu Huang
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - Danny Ka-Ho Wong
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
- State Key Laboratory for Liver Research, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - Wai-Kay Seto
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - An-Ye Zhang
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - Cheuk-Kwong Lee
- Hong Kong Red Cross Blood Transfusion Service, Hospital Authority, Hong Kong, China
| | - Che-Kit Lin
- Hong Kong Red Cross Blood Transfusion Service, Hospital Authority, Hong Kong, China
| | - James Fung
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
- State Key Laboratory for Liver Research, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - Ching-Lung Lai
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
- State Key Laboratory for Liver Research, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
| | - Man-Fung Yuen
- Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
- State Key Laboratory for Liver Research, The University of Hong Kong, Queen Mary Hospital, Hong Kong, China
- * E-mail:
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Besharat S, Katoonizadeh A, Moradi A. Potential mutations associated with occult hepatitis B virus status. HEPATITIS MONTHLY 2014; 14:e15275. [PMID: 24829588 PMCID: PMC4013497 DOI: 10.5812/hepatmon.15275] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/06/2013] [Revised: 01/14/2014] [Accepted: 02/20/2014] [Indexed: 12/11/2022]
Abstract
CONTEXT Occult hepatitis B virus (HBV) status (OHBS) is simply defined as the presence of HBV DNA in the liver (with or without detectable HBV DNA in the serum), in the absence of serum HBV surface antigen (HBsAg). Importance of OHBS is mostly clinical, related to its possible role in spreading through blood transfusion and liver transplantation; causing classic forms of HBV. Mechanisms underlying this entity are poorly defined. Several possibilities have been suggested, with major classification into two groups: defective host immune response and viral replication activity through mutations of HBV DNA sequence. Mutations are extensively investigated in all four overlapping open reading frames (ORFs) of HBV genome, to define their possible role in the pathogenesis of OHBS. Some of these mutations like S-escape mutants could not be detected by the routine available assays, making them difficult to diagnosis. Therefore, trying to detect this covert condition could be more helpful for defining better preventive and therapeutic strategies. EVIDENCE ACQUISITION In the present study we provided an in-depth review of the most important new data available on different mutations in HBV genome of patients with OHBS, which may play a role in the pathogenesis of OHBS. The data were collected through reviewing the full-text articles, identified by the PubMed search, using the following keywords and their different combinations: occult hepatitis B, HBV genome, "a" determinant, HBV open reading frames, S mutations, X mutations, P mutations and C mutations. RESULTS Variants within the major hydrophilic region (MHR) of the HBsAg, deletions in the pre-S1region, codon stop in the S open reading frames (ORF), sporadic non common mutations, some mutations affecting the posttranslational production of HBV proteins in the S ORF like deletion mutations, mutations in start codon and nucleotide changes in the X ORF, deletion and point mutations in P ORF and sometimes, nucleotide substitution in the C ORF are among the assumed mutations detected to have a role in OHBS appearance. CONCLUSIONS Studies mostly lacked a control group and the whole-length HBV sequencing was scant with conflicting results, suggesting that OHBS is often a result of multiple mechanisms. Additional studies on full-length HBV genomes from occult and non-occult HBV cases may shed more light on the interplay between different mechanisms involved in the pathogenesis of OHBS.
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Affiliation(s)
- Sima Besharat
- Liver and Pancreatobiliary Diseases Research Center, Digestive Disease Research Institute, Tehran University of Medical Sciences, Tehran, IR Iran
- Golestan Research Center of Gastroentrology and Hepatology, Golestan University of Medical Sciences, Gorgan, IR Iran
| | - Aezam Katoonizadeh
- Liver and Pancreatobiliary Diseases Research Center, Digestive Disease Research Institute, Tehran University of Medical Sciences, Tehran, IR Iran
| | - Abdolvahab Moradi
- Golestan Research Center of Gastroentrology and Hepatology, Golestan University of Medical Sciences, Gorgan, IR Iran
- Corresponding Author: Abdolvahab Moradi, Golestan Research Center of Gastroentrology and Hepatology, Golestan University of Medical Sciences, Gorgan, IR Iran. Tel: +98-1712340835; +98-9111772107, Fax: +98-1712369210, E-mail:
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Occult HBV infection: a faceless enemy in liver cancer development. Viruses 2014; 6:1590-611. [PMID: 24717680 PMCID: PMC4014712 DOI: 10.3390/v6041590] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2014] [Revised: 03/13/2014] [Accepted: 03/20/2014] [Indexed: 12/12/2022] Open
Abstract
The hepatitis B virus (HBV) represents a worldwide public health problem; the virus is present in one third of the global population. However, this rate may in fact be higher due to occult hepatitis B virus infection (OBI). This condition is characterized by the presence of the viral genome in the liver of individuals sero-negative for the virus surface antigen (HBsAg). The causes of the absence of HBsAg in serum are unknown, however, mutations have been identified that produce variants not recognized by current immunoassays. Epigenetic and immunological host mechanisms also appear to be involved in HBsAg suppression. Current evidence suggests that OBI maintains its carcinogenic potential, favoring the progression of fibrosis and cirrhosis of the liver. In common with open HBV infection, OBI can contribute to the establishment of hepatocellular carcinoma. Epidemiological data regarding the global prevalence of OBI vary due to the use of detection methods of different sensitivity and specificity. In Latin America, which is considered an area of low prevalence for HBV, diagnostic screening methods using gene amplification tests for confirmation of OBI are not conducted. This prevents determination of the actual prevalence of OBI, highlighting the need for the implementation of cutting edge technology in epidemiological surveillance systems.
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Support of the infectivity of hepatitis delta virus particles by the envelope proteins of different genotypes of hepatitis B virus. J Virol 2014; 88:6255-67. [PMID: 24648462 DOI: 10.1128/jvi.00346-14] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
UNLABELLED This study examined how the envelope proteins of 25 variants of hepatitis B virus (HBV) genotypes A to I support hepatitis delta virus (HDV) infectivity. The assembled virions bore the same HDV ribonucleoprotein and differed only by the HBV variant-specific envelope proteins coating the particles. The total HDV yields varied within a 122-fold range. A residue Y (position 374) in the HDV binding site was identified as critical for HDV assembly. Virions that bound antibodies, which recognize the region that includes the HBV matrix domain and predominantly but not exclusively immunoprecipitate the PreS1-containing virions, were termed PreS1*-HDVs. Using in vitro infection of primary human hepatocytes (PHH), we measured the specific infectivity (SI), which is the number of HDV genomes/cell produced by infection and normalized by the PreS1*-MOI, which is the multiplicity of infection that reflects the number of PreS1*-HDVs per cell in the inoculum used. The SI values varied within a 160-fold range and indicated a probable HBV genotype-specific trend of D > B > E > A in supporting HDV infectivity. Three variants, of genotypes B, C, and D, supported the highest SI values. We also determined the normalized index (NI) of infected PHH, which is the percentage of HDV-infected hepatocytes normalized by the PreS1*-MOI. Comparison of the SI and NI values revealed that, while a particular HBV variant may facilitate the infection of a relatively significant fraction of PHH, it may not always result in a considerable number of genomes that initiated replication after entry. The potential implications of these findings are discussed in the context of the mechanism of attachment/entry of HBV and HDV. IMPORTANCE The study advances the understanding of the mechanisms of (i) attachment and entry of HDV and HBV and (ii) transmission of HDV infection/disease.
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Huang X, Hollinger FB. Occult hepatitis B virus infection and hepatocellular carcinoma: a systematic review. J Viral Hepat 2014; 21:153-62. [PMID: 24438677 DOI: 10.1111/jvh.12222] [Citation(s) in RCA: 56] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/13/2013] [Accepted: 12/08/2013] [Indexed: 12/13/2022]
Abstract
Occult hepatitis B (OHB) infection has been reported to play an important role in the development of hepatocellular carcinoma (HCC). In this systematic review, a significantly higher prevalence of OHB was observed in patients with HCC in the presence or absence of HCV infection when compared with control populations without HCC. Correspondingly, among adequately designed prospective studies, the cumulative probability of developing HCC was significantly greater among patients with OHB than among HBV DNA-negative patients in the presence or absence of HCV infection. Study design, inclusion criteria, treatment options, methodology and potential confounding variables were evaluated, and immunopathogenic mechanisms that could be involved in OHB as a risk factor in HCC were reviewed. From this analysis, we conclude that although OHB is an independent risk factor in HCC development in anti-HCV-negative patients, a synergistic or additive role in the occurrence of HCC in HCV-coinfected patients is more problematic due to the HCC risk attributable to HCV alone, especially in patients with advanced fibrosis and cirrhosis.
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Affiliation(s)
- X Huang
- Department of Blood Transfusion, The General Hospital of Jinan Military Command, Jinan, China
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Oshima Y, Tsukamoto H, Tojo A. Association of hepatitis B with antirheumatic drugs: a case–control study. Mod Rheumatol 2014. [DOI: 10.3109/s10165-012-0709-7] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
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Development of a highly sensitive bioluminescent enzyme immunoassay for hepatitis B virus surface antigen capable of detecting divergent mutants. CLINICAL AND VACCINE IMMUNOLOGY : CVI 2013; 20:1255-65. [PMID: 23761660 DOI: 10.1128/cvi.00186-13] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Hepatitis B virus (HBV) infections are sometimes overlooked when using commercial kits to measure hepatitis B virus surface antigen (HBsAg) due to their low sensitivities and reactivities to mutant strains of various genotypes. We developed an ultrasensitive bioluminescent enzyme immunoassay (BLEIA) for HBsAg using firefly luciferase, which is adaptable to a variety of HBsAg mutants, by combining four monoclonal antibodies with a polyclonal antibody against HBsAg. The measurement of seroconversion panels showed trace amounts of HBsAg during the early infection phase by the BLEIA because of its high sensitivity of 5 mIU/ml. The BLEIA detected HBsAg as early as did PCR in five of seven series and from 2.1 to 9.4 days earlier than commercial immunoassay methods. During the late infection phase, the BLEIA successfully detected HBsAg even 40 days after the disappearance of HBV DNA and the emergence of antibodies against HBsAg. The HBsAg BLEIA successfully detected all 13 recombinant HBsAg and 45 types of HBsAg mutants with various mutations within amino acids 90 to 164 in the S gene product. Some specimens had higher values determined by the BLEIA than those by a commercial chemiluminescent immunoassay; this suggests that such discrepancies were caused by the dissociation of preS1/preS2 peptides from the particle surface. With its highly sensitive detection of low-titer HBsAg, including various mutants, the HBsAg BLEIA is considered to be useful for the early diagnosis and prevention of HBV infection because of the shorter window of infection prior to detection, which facilitates early prediction of recurrence in HBV-infected individuals.
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Specific amino acid substitutions in the S protein prevent its excretion in vitro and may contribute to occult hepatitis B virus infection. J Virol 2013; 87:7882-92. [PMID: 23658444 DOI: 10.1128/jvi.00710-13] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Occult hepatitis B virus (HBV) infection (OBI) is defined as low plasma level of HBV DNA with undetectable HBV surface antigen (HBsAg) outside the preseroconversion window period. The mechanisms leading to OBI remain largely unknown. The potential role of specific amino acid substitutions in the S protein from OBI in HBsAg production and excretion was examined in vitro. HBsAg was quantified in culture supernatants and cell extracts of HuH-7 cells transiently transfected with plasmids containing the S gene of eight HBsAg(+) controls and 18 OBI clones. The intracellular (IC)/extracellular (EC) HBsAg production ratio was ∼1.0 for the majority of controls. Three IC/EC HBsAg patterns were observed in OBI strains clones: pattern 1, an IC/EC ratio of 1.0, was found in 5/18 OBI clones, pattern 2, detectable IC but low or undetectable EC HBsAg (IC/EC, 7.0 to 800), was found in 6/18 OBIs, and pattern 3, low or undetectable IC and EC HBsAg, was found in 7/18 clones. Intracellular immunofluorescence staining showed that in pattern 2, HBsAg was concentrated around the nucleus, suggesting retention in the endoplasmic reticulum. The substitution M75T, Y100S, or P178R was present in 4/6 pattern 2 OBI clones. Site-directed-mutagenesis-corrected mutations reversed HBsAg excretion to pattern 1 and, when introduced into a control clone, induced pattern 2 except for Y100S. In a control and several OBIs, variants of a given quasispecies expressed HBsAg according to different patterns. However, the P178R substitution present in all cloned sequences of two OBI strains may contribute significantly to the OBI phenotype.
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Blackard JT, Martin CM, Sengupta S, Forrester J. Limited infection with occult hepatitis B virus in drug users in the USA. Hepatol Res 2013; 43:413-7. [PMID: 22909008 PMCID: PMC3505246 DOI: 10.1111/j.1872-034x.2012.01072.x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
AIM Occult HBV infection (O-HBV) is defined as low level HBV replication in the absence of detectable circulating HBV surface antigen. O-HBV has been implicated in HBV reactivation, advanced liver fibrosis and cirrhosis, reduced interferon response rates, elevated liver enzyme levels, and the development of hepatocellular carcinoma. However, the prevalence of O-HBV has not been clearly established in certain at-risk populations, such as injection drug users. METHODS Therefore, the current pilot study examined the prevalence of O-HBV in a prospective cohort designed to assess the role of injection and non-injection drug use (IDU) on HIV-associated comorbidities. RESULTS Utilizing two distinct real-time polymerase chain reaction assays, HBV DNA was not detected in 99 participants examined. CONCLUSION This finding is in contrast to other data from US IDU cohorts and suggests that the prevalence of O-HBV infection is very specific to the cohort studied, is sensitive to other confounding variables such as hepatitis C virus and/or HIV serostatus, and should not be generalized across risk groups or distinct cohorts.
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Affiliation(s)
- Jason T. Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH
,Address requests for reprints to: Jason Blackard, PhD, Division of Digestive Diseases, University of Cincinnati College of Medicine, ML 0595, 231 Albert Sabin Way, Cincinnati, OH 45267, Phone: (513) 558-4389, Fax: (513) 558-1744,
| | - Christina M. Martin
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH
| | - Satarupa Sengupta
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH
| | - Janet Forrester
- Department of Public Health & Community Medicine, Tufts University School of Medicine, Boston, MA
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Huang CC, Kuo TM, Yeh CT, Hu CP, Chen YL, Tsai YL, Chen ML, Chou YC, Chang C. One single nucleotide difference alters the differential expression of spliced RNAs between HBV genotypes A and D. Virus Res 2013; 174:18-26. [PMID: 23501362 DOI: 10.1016/j.virusres.2013.02.004] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2012] [Revised: 02/08/2013] [Accepted: 02/08/2013] [Indexed: 12/13/2022]
Abstract
Hepatitis B virus (HBV) is generally classified into eight genotypes (A to H) based on genomic sequence divergence. The sequence variation among the different HBV genotypes suggests that the spliced RNAs should be different from genotype to genotype. However, the cis-acting element involved in the modulation of the distinct expression profiles of spliced HBV RNAs remains unidentified. Moreover, the biological role of splicing in the life cycle of HBV is not yet understood. In this study, spliced RNAs generated from genotypes A and D were carefully characterized in transfected HepG2 cells. The species and frequency of the spliced RNAs were dramatically different in the two genotypes. Of note, a population of multiply spliced RNAs with intron 2067-2350 excision was identified in HBV genotype A-transfected HepG2 cells, but not in genotype D transfected HepG2 cells. Further, we found a single nucleotide difference (2335) located within the polypyrimidine tract of the splice acceptor site 2350 between the two genotypes, and a single base substitution at 2335 was able to convert the splicing pattern of genotype D (or genotype A) to that of genotype A (or genotype D). These findings suggest that different unique splice sites may be preferentially used in different HBV genotypes resulting in distinct populations of spliced RNAs. The possible significance of the distinct spliced RNAs generated from the different HBV genotypes in HBV infection is discussed.
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Affiliation(s)
- Chien-Chiao Huang
- Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan.
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Martin CM, Welge JA, Rouster SD, Shata MT, Sherman KE, Blackard JT. Mutations associated with occult hepatitis B virus infection result in decreased surface antigen expression in vitro. J Viral Hepat 2012; 19:716-23. [PMID: 22967103 PMCID: PMC3442934 DOI: 10.1111/j.1365-2893.2012.01595.x] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Occult hepatitis B virus (HBV) infection is characterized by the absence of detectable hepatitis B surface antigen (HBsAg) in the serum, despite detectable HBV DNA. Investigations of the mechanisms underlying the development of occult HBV infection are lacking in the current literature, although viral mutations in the surface region, resulting in decreased HBsAg expression or secretion, represent one potential mechanism. Wild-type HBsAg expression vectors were constructed from genotype-matched chronic HBV sequences. Site-directed mutagenesis was then utilized to introduce three genotype A mutations - M103I, K122R and G145A - associated with occult HBV infection in vivo, alone and in combination, into the wild-type HBsAg vectors. Transfection of Huh7 and HepG2 cell lines was performed, and cell culture supernatants and cell lysates were collected over 7 days to assess the effects of these mutations on extracellular and intracellular HBsAg levels. The G145A mutation resulted in significantly decreased extracellular and intracellular HBsAg expression in vitro. The most pronounced reduction in HBsAg expression was observed when all three mutations were present. The mutations evaluated in vitro in the current study resulted in decreased HBsAg expression and potentially increased hepatic retention and/or decreased hepatic secretion of synthesized HBsAg, which could explain the lack of HBsAg detection that is characteristic of occult HBV infection in vivo.
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Affiliation(s)
- Christina M. Martin
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267
| | - Jeffrey A. Welge
- Departments of Psychiatry and Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267
| | - Susan D. Rouster
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267
| | - Mohamed Tarek Shata
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267
| | - Kenneth E. Sherman
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267
| | - Jason T. Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267, Address requests for reprints to: Jason Blackard, PhD Division of Digestive Diseases University of Cincinnati College of Medicine ML 0595, 231 Albert Sabin Way Cincinnati, OH 45267 Phone: (513) 558-4389 Fax: (513) 558-1744
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Oshima Y, Tsukamoto H, Tojo A. Association of hepatitis B with antirheumatic drugs: a case-control study. Mod Rheumatol 2012; 23:694-704. [PMID: 22802011 DOI: 10.1007/s10165-012-0709-7] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2011] [Accepted: 06/19/2012] [Indexed: 12/12/2022]
Abstract
BACKGROUND Though concern of hepatitis B virus (HBV) reactivation by antirheumatic agents has limited therapeutic opportunities in HBV-infected rheumatoid arthritis (RA) patients, the relative risks (RR) among such agents have not been clarified. OBJECTIVE We compared the reporting of antirheumatic-agent-associated hepatitis B. PATIENTS We assessed 92 hepatitis B cases and 98,069 controls from a population of 98,161 RA patients registered into the US Food and Drug Administration's (FDA's) adverse event database between 2004 and 2010. MEASUREMENTS A reporting odds ratio (ROR), a signal suggesting a risk for hepatitis B among antirheumatic agents, was measured. RESULTS Treatment with corticosteroids [ROR 2.3 (95% confidence interval 1.3-4.0)], methotrexate [4.9 (3.9-6.0)], rituximab [7.2 (5.3-9.9)], tacrolimus [4.2 (1.5-11.9)], or reporting from Japan [2.2 (1.1-4.2)] were associated with higher signal, whereas adalimumab had a lower ROR [0.2 (0.1-0.4)]. LIMITATIONS There are known limitations of spontaneous reporting, such as underreporting, the Weber effect, reporting bias, indication bias, and limited clinical information such as HBV status. CONCLUSIONS Adalimumab's low reporting rate is most likely be due to notoriety. However, the possibility that adalimumab might suppress reactivation of HBV cannot be denied. Until the possibility is clarified in well-designed clinical studies, physicians should use adalimumab cautiously in patients with HBV.
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Affiliation(s)
- Yasuo Oshima
- The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
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Hepatitis B virus DNA splicing in Lebanese blood donors and genotype A to E strains: implications for hepatitis B virus DNA quantification and infectivity. J Clin Microbiol 2012; 50:3159-67. [PMID: 22785194 DOI: 10.1128/jcm.01251-12] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Hepatitis B virus (HBV) is one of the major viruses transmissible by blood that causes chronic infection in immunocompromised individuals. The study of 61 HBV carrier blood donors from Lebanon revealed multiple patterns of spliced HBV DNA. HBV DNA splicing was examined and quantified in samples of five genotypes and in seroconversion panels. The Lebanese sample median viral load was 1.5 ×10(2) IU/ml. All strains were genotype D, serotype ayw; 35 clustered as subgenotype D1 and 7 clustered as subgenotype D2. Three splice variants (SP1, SP1A, and Pol/S) were observed in 12 high-viral-load samples. Twenty samples of each genotype, A to E, were tested for the presence of HBV spliced DNA and SP1-specific splice variant. An unspliced HBV genome was dominant, but 100% of strains with a viral load of ≥10(5) copies/ml contained various proportions of spliced DNA. SP1 was detected in 56/100 (56%) samples in levels that correlated with the overall viral load. HBV DNA quantification with S (unspliced) and X (total DNA) regions provided different levels of viral load, with the difference corresponding to spliced DNA. During the highly infectious window period, the SP1 variant became detectable shortly after the hepatitis B surface antigen (HBsAg), suggesting a correlation between the initiation of splicing and the production of detectable levels of HBsAg. The quantification of HBV DNA with primers located outside and inside the spliced region might provide different estimations of viral load and differentiate between infectious and defective viral genomes. The role of splicing neoproteins in HBV replication and interaction with the host remains to be determined.
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