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Advanced sequencing approaches detected insertions of viral and human origin in the viral genome of chronic hepatitis E virus patients.

ABSTRACT: The awareness of hepatitis E virus (HEV) increased significantly in the last decade due to its unexpectedly high prevalence in high-income countries. There, infections with HEV-genotype 3 (HEV-3) are predominant which can progress to chronicity in immunocompromised individuals. Persistent infection and antiviral therapy can select HEV-3 variants; however, the spectrum and occurrence of HEV-3 variants is underreported. To gain in-depth insights into the viral population and to perform detailed characterization of viral genomes, we used a new approach combining long-range PCR with next-generation and third-generation sequencing which allowed near full-length sequencing of HEV-3 genomes. Furthermore, we developed a targeted ultra-deep sequencing approach to assess the dynamics of clinically relevant mutations in the RdRp-region and to detect insertions in the HVR-domain in the HEV genomes. Using this new approach, we not only identified several insertions of human (AHNAK, RPL18) and viral origin (RdRp-derived) in the HVR-region isolated from an exemplary sample but detected a variant containing two different insertions simultaneously (AHNAK- and RdRp-derived). This finding is the first HEV-variant recognized as such showing various insertions in the HVR-domain. Thus, this molecular approach will add incrementally to our current knowledge of the HEV-genome organization and pathogenesis in chronic hepatitis E.

SUBMITTER: Papp CP

PROVIDER: S-EPMC8811047 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

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Advanced sequencing approaches detected insertions of viral and human origin in the viral genome of chronic hepatitis E virus patients.

Papp C-Patrick CP Biedermann Paula P Harms Dominik D Wang Bo B Kebelmann Marianne M Choi Mira M Helmuth Johannes J Corman Victor M VM Thürmer Andrea A Altmann Britta B Klink Patrycja P Hofmann Jörg J Bock C-Thomas CT

Scientific reports 20220202 1

The awareness of hepatitis E virus (HEV) increased significantly in the last decade due to its unexpectedly high prevalence in high-income countries. There, infections with HEV-genotype 3 (HEV-3) are predominant which can progress to chronicity in immunocompromised individuals. Persistent infection and antiviral therapy can select HEV-3 variants; however, the spectrum and occurrence of HEV-3 variants is underreported. To gain in-depth insights into the viral population and to perform detailed ch ...[more]

PMID: 35110582

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Project description:Persistent antigen exposure during chronic hepatitis B infection leads to exhausted immune responses, thus impeding viral control. In recent years, immunometabolism opens new therapeutic possibilities for the modulation of immune responses. Herein, we investigated the immunomodulatory effect of L-carnitine (L-Cn) on immune cells in chronic HBV infection. In this study, 141 treatment-naïve patients with chronic HBV infection, 38 patients who achieved HBsAg loss following antiviral treatment, and 47 patients who suffered from HBV-related HCC from real-life clinical practice were recruited. The plasma L-Cn levels were measured by ELISA. RNA sequencing was conducted to define the transcriptional profiles of peripheral blood mononuclear cells after L-Cn stimulation. In vitro assays were performed to assess the effect of L-Cn on immune cells; the frequencies and function of immune cells were analyzed by flow cytometry. We found that compared with patients with HBsAg loss, patients with HBsAg positivity and patients who suffered from HBV-related HCC had higher levels of L-Cn, and the plasma levels of L-Cn in the HBeAg-positive chronic hepatitis patients who had elevated ALT were significantly higher than that of HBeAg-negative chronic infection and HBsAg loss groups. Moreover, a positive correlation between plasma levels of L-Cn and HBsAg levels was found. Additionally, RNA sequencing analysis demonstrated that L-Cn altered the transcriptional profiles related to immune response. In vitro assays revealed that L-Cn suppressed the proliferation of and IFN-γ production by CD4+ and CD8+ T cells. It also down-regulated the proliferation and IgG production of B cells. Notably, L-Cn enhanced IL-10 secretion from regulatory T cells and up-regulated the expression of inhibitory receptors on T cells. Moreover, a variant in CPT2 (rs1799821) was confirmed to be associated with L-Cn levels as well as complete response in CHB patients following Peg-IFNα antiviral therapy. Taken together, the immunosuppressive properties of L-Cn may hinder the control of HBV in chronic HBV infection, implicating that L-Cn manipulation might influence the prognosis of patients with HBV infection.

Project description:Recombinant strains of hepatitis E virus (HEV) with insertions of human genomic fragments or HEV sequence duplications in the sequence encoding the polyproline region (PPR) were previously described in chronically infected patients. Such genomic rearrangements confer a replicative advantage in vitro but little is known about their frequency, location, or origin. As the sequences of only a few virus genomes are available, we analyzed the complete genomes of 114 HEV genotype 3 strains from immunocompromised (n = 85) and immunocompetent (n = 29) patients using the single molecular real-time sequencing method to determine the frequency, location, and origin of inserted genomic fragments, plus the proportions of variants with genomic rearrangements in each virus quasispecies. We also examined the amino acid compositions and post-translational modifications conferred by these rearrangements by comparing them to sequences without human gene insertions or HEV gene duplications. We found genomic rearrangements in 7/114 (6.1%) complete genome sequences (4 HEV-3f, 1 HEV-3e, 1 HEV-3 h, and 1 HEV-3chi-new), all from immunocompromised patients, and 3/7 were found at the acute phase of infection. Six of the seven patients harbored virus-host recombinant variants, including one patient with two different recombinant variants. We also detected three recombinant variants with genome duplications of the PPR or PPR + X domains in a single patient. All the genomic rearrangements (seven human fragment insertions of varying origins and three HEV genome duplications) occurred in the PPR. The sequences with genomic rearrangements had specific characteristics: increased net load (p < 0.001) and more ubiquitination (p < 0.001), phosphorylation (p < 0.001), and acetylation (p < 0.001) sites. The human fragment insertions and HEV genome duplications had slightly different characteristics. We believe this is the first description of HEV strains with genomic rearrangements in patients at the acute phase of infection; perhaps these strains are directly transmitted. Clearly, genomic rearrangements produce a greater net load with duplications and insertions having different features. Further studies are needed to clarify the mechanisms by which such modifications influence HEV replication.

Dataset Information

Advanced sequencing approaches detected insertions of viral and human origin in the viral genome of chronic hepatitis E virus patients.

Publications

Advanced sequencing approaches detected insertions of viral and human origin in the viral genome of chronic hepatitis E virus patients.

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