An <scp>ORF</scp>1‐rearranged hepatitis <scp>E</scp> virus derived from a chronically infected patient efficiently replicates in cell culture

Johne, Reimar; Reetz, Jochen; Ulrich, Rainer G.; Machnowska, Patrycja; Sachsenröder, Jana; Nickel, Peter; Hofmann, Jörg

doi:10.1111/jvh.12157

Cited by 102 publications

(138 citation statements)

References 36 publications

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“…Similar to 4 late resurgences of HEV RNA levels in vitro demonstrated here, others found productive in vitro infections with insertions in the ORF1 region after 5 to 6 weeks of culture of chronic HEV gt3 sera, but not of acute-phase sera (26)(27)(28)(29).…”

Section: Discussionmentioning

confidence: 60%

Hepatitis E Virus (HEV) Genotype 3 Infection of Human Liver Chimeric Mice as a Model for Chronic HEV Infection

Garde

Pas

Net

et al. 2016

J Virol

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Genotype 3 (gt3) hepatitis E virus (HEV) infections are emerging in Western countries. Immunosuppressed patients are at risk of chronic HEV infection and progressive liver damage, but no adequate model system currently mimics this disease course. Here we explore the possibilities of in vivo HEV studies in a human liver chimeric mouse model (uPA ؉/؉ Nod-SCID-IL2R␥ ؊/؊ ) next to the A549 cell culture system, using HEV RNA-positive EDTA-plasma, feces, or liver biopsy specimens from 8 immunocompromised patients with chronic gt3 HEV. HEV from feces-or liver-derived inocula showed clear virus propagation within 2 weeks after inoculation onto A549 cells, compared to slow or no HEV propagation of HEV RNA-positive, EDTA-plasma samples. These in vitro HEV infectivity differences were mirrored in human-liver chimeric mice after intravenous (i.v.) inoculation of selected samples. HEV RNA levels of up to 8 log IU HEV RNA/gram were consistently present in 100% of chimeric mouse livers from week 2 to week 14 after inoculation with human feces-or liver-derived HEV. Feces and bile of infected mice contained moderate to large amounts of HEV RNA, while HEV viremia was low and inconsistently detected. Mouse-passaged HEV could subsequently be propagated for up to 100 days in vitro. In contrast, cell culture-derived or seronegative EDTA-plasma-derived HEV was not infectious in inoculated animals. In conclusion, the infectivity of feces-derived human HEV is higher than that of EDTA-plasma-derived HEV both in vitro and in vivo. Persistent HEV gt3 infections in chimeric mice show preferential viral shedding toward mouse bile and feces, paralleling the course of infection in humans. H epatitis E virus (HEV) is a nonenveloped positive-sense single-stranded RNA virus of the genus Orthohepevirus and family Hepeviridae (1). Four major HEV genotypes infecting humans have been described so far. Genotype 1 (gt1) and gt2 strains are isolated only from humans, whereas gt3 and gt4 strains are considered zoonotic viruses, present in both humans and several other species like pigs and wild game. HEV is spread through the oral-fecal route via contaminated water in developing countries or, among other routes, via direct contact with animals or the consumption of undercooked meat in industrialized countries. In immunocompetent individuals, HEV infection is mainly self-limiting and often asymptomatic and thus remains largely underdiagnosed. HEV infections in immunosuppressed patients, such as solid-organ transplant recipients, often persist and can progress quickly to liver fibrosis and cirrhosis (2-4).HEV gt3 infections are emerging in Western countries, including France, the United Kingdom, and the Netherlands (5-7). Despite an overall decrease in anti-HEV seroprevalence from 1996 to 2011, young adult blood donors demonstrated higher seroprevalences from 2000 to 2011 (8, 9). In addition, HEV RNA-positive blood donations were reported to increase in the Netherlands since 2012 (10). Although the exact source of this HEV gt3 infection is unknown,...

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Section: Discussionmentioning

confidence: 60%

Hepatitis E Virus (HEV) Genotype 3 Infection of Human Liver Chimeric Mice as a Model for Chronic HEV Infection

Garde

Pas

Net

et al. 2016

J Virol

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“…The HVR has been proposed to play a role in HEV persistence in immunocompromised individuals (22). Additionally, insertions and rearrangements within the ORF1 protein have given an enhanced ability to replicate in cell culture (23,24).…”

mentioning

confidence: 99%

The Lysine Residues within the Human Ribosomal Protein S17 Sequence Naturally Inserted into the Viral Nonstructural Protein of a Unique Strain of Hepatitis E Virus Are Important for Enhanced Virus Replication

Kenney

Meng

2015

J Virol

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Hepatitis E virus (HEV) is an important but extremely understudied human pathogen. Due largely to the lack of an efficient cell culture system for HEV, the molecular mechanisms of HEV replication and pathogenesis are poorly understood. Recently, a unique genotype 3 strain of HEV recovered from a chronically infected patient was adapted for growth in HepG2C3A human hepatoma cells. The adaptation of the Kernow C-1 P6 HEV to propagate in HepG2C3A cells selected for a rare virus recombinant that contains an insertion of a 171-nucleotide sequence encoding amino acids 21 to 76 of the human ribosomal protein S17 (RPS17) within the hypervariable region (HVR) of the HEV ORF1 protein. When the RPS17 insertion was placed into a strain of genotype 1 HEV which infects only humans, it expanded the host range of the virus, allowing it to infect cell lines from multiple animal species, including cow, dog, cat, chicken, and hamster. In this study, we utilized forward and reverse genetics to attempt to define which aspects of the RPS17 insertion allow for the ability of the Kernow C-1 P6 HEV to adapt in cell culture and allow for expanded host tropism. We demonstrate that the RPS17 sequence insertion in HEV bestows novel nuclear/nucleolar trafficking capabilities to the ORF1 protein of Kernow P6 HEV and that lysine residues within the RPS17 insertion, but not nuclear localization of the ORF1 protein, correlate with the enhanced replication of the HEV Kernow C-1 P6 strain. The results from this study have important implications for understanding the mechanism of cross-species infection and replication of HEV. IMPORTANCEHEV is an important pathogen worldwide. The virus causes high mortality (up to 30%) in pregnant women and has been recognized to cause chronic hepatitis in immunocompromised populations. The life cycle of HEV has been understudied due to a lack of sufficient cell culture systems in which to propagate the virus. Recently, insertions and rearrangements of the hypervariable region (HVR) within the HEV genome, allowing for cell culture adaptation and expansion of the host range, have been reported. We utilized these cell culture-adapted HEV strains to assess how the HVR may be involved in virus replication and host range. We provide evidence that insertion of the RPS17 sequence in HEV likely confers nuclear trafficking capabilities to the nonstructural protein of the virus and that lysine residues within the RPS17 insertion are important for enhanced replication of the virus. These data will help to elucidate the mechanism of cross-species infection of HEV in the future. Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis in many parts of the world (1, 2). Major outbreaks and epidemics are most common in developing countries, where a lack of proper sanitation conditions leads to waterborne spread of the disease. In industrialized countries, HEV infection is typically sporadic and endemic. Recently, chronic HEV infection has become an important clinical problem in immunosuppressed individual...

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“…In this study, two additional HEV-positive serum samples from patients acutely infected with a virus of unknown genotype and a wild boar liver sample infected with HEV 3i genome FJ705359 were also inoculated onto A549 cells, and a persistent infection was obtained only with the serum sample of the chronically infected patient. Interestingly, a complex insertion within the hypervariable region of the genome was identified, as in two other cell culture-adapted HEV 3a strains that also originated from chronically infected patients (14). These three insertions differed by their sequences but had similar lengths and positions in the HEV genome, and such insertions were absent from the genome described here.…”

Section: Discussionmentioning

confidence: 89%

“…Nevertheless, present analyses revealed that approximately two-thirds of HEV 3i sequences from humans were from known chronically infected kidney transplant recipients. In addition, Johne et al recently obtained a persistently infected cell line after inoculation with the serum sample of a kidney transplant recipient chronically infected with HEV 3i (genome KC618402) (14). In this study, two additional HEV-positive serum samples from patients acutely infected with a virus of unknown genotype and a wild boar liver sample infected with HEV 3i genome FJ705359 were also inoculated onto A549 cells, and a persistent infection was obtained only with the serum sample of the chronically infected patient.…”

Section: Discussionmentioning

confidence: 99%

“…Using short HEV ORF2 fragments may therefore generate misclassification. HEV 3i has been identified in five countries (Austria, Argentina, Bolivia, Germany, and Uruguay) and from various hosts, including humans (3,14,17), wild boars (12,13), and swine (11,17). HEV sequences from strains wbGER27 (FJ705359) and BB02 (FJ998008), which were recovered from wild boars in Germany, were fully sequenced and considered the first HEV 3i genomes (12,13).…”

Section: Discussionmentioning

confidence: 99%

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Hepatitis E Virus of Subtype 3i in Chronically Infected Kidney Transplant Recipients in Southeastern France

et al. 2014

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e Hepatitis E virus (HEV) is a leading cause of waterborne acute hepatitis in developing countries. In Europe, HEV causes a zoonotic disease and is hyperendemic in southern France. Four HEV genotypes (1 to 4) have been defined, and the most used classification divides them into 24 subtypes. Autochthonous European HEV strains belong in majority to genotype 3. Subtypes 3c, 3f, and 3e are representative of the HEV diversity in France. HEV causes chronic hepatitis in solid-organ transplant recipients in Europe, and viral characteristics associated with chronicity are poorly documented. We sequenced 343-nucleotide-long HEV genomic fragments from the serum of eight chronically infected kidney transplant recipients and a near-full-length genome in one case. We identified in four patients (50%) HEV of subtype 3i, not described previously in France. If shorter genomic fragments were used in phylogenetic analyses, these HEV sequences were clustered with open reading frame 2 (ORF2) fragments labeled as subtype 3c. At least five of the eight HEV 3i sequences recovered from humans in our phylogenetic analyses were from chronically infected kidney transplant recipients. These data show that the description of the prevalence and geographical distribution of HEV subtypes may be partially inaccurate and that criteria for classification as 3i and 3c should be clarified. Extended molecular virology analyses are required to improve knowledge of HEV epidemiology and determinants of chronic HEV infection.

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An ORF1‐rearranged hepatitis E virus derived from a chronically infected patient efficiently replicates in cell culture

Cited by 102 publications

References 36 publications

Hepatitis E Virus (HEV) Genotype 3 Infection of Human Liver Chimeric Mice as a Model for Chronic HEV Infection

Hepatitis E Virus (HEV) Genotype 3 Infection of Human Liver Chimeric Mice as a Model for Chronic HEV Infection

The Lysine Residues within the Human Ribosomal Protein S17 Sequence Naturally Inserted into the Viral Nonstructural Protein of a Unique Strain of Hepatitis E Virus Are Important for Enhanced Virus Replication

Hepatitis E Virus of Subtype 3i in Chronically Infected Kidney Transplant Recipients in Southeastern France

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