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The Capital Region of Denmark - a part of Copenhagen University Hospital
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Productive Homologous and Non-homologous Recombination of Hepatitis C Virus in Cell Culture

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  1. CO-HEP; Copenhagen Hepatitis C Program

    Project: Types of projects

  1. The fading boundaries between patient and environmental routes of triazole resistance selection in Aspergillus fumigatus

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  2. Hepatitis C virus cell-cell transmission and resistance to direct-acting antiviral agents

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  3. JC polyomavirus infection is strongly controlled by human leucocyte antigen class II variants

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  4. Association study of common genetic variants and HIV-1 acquisition in 6,300 infected cases and 7,200 controls

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  1. Ribavirin inhibition of cell-culture infectious hepatitis C genotype 1-3 viruses is strain-dependent

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  2. Genome Sequence of an Unknown Subtype of Hepatitis C Virus Genotype 6: Another Piece for the Taxonomic Puzzle

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  3. Virus Adaptation and Selection Following Challenge of Animals Vaccinated against Classical Swine Fever Virus

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  4. HCV p7 as a novel vaccine-target inducing multifunctional CD4+ and CD8+ T-cells targeting liver cells expressing the viral antigen

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Genetic recombination is an important mechanism for increasing diversity of RNA viruses, and constitutes a viral escape mechanism to host immune responses and to treatment with antiviral compounds. Although rare, epidemiologically important hepatitis C virus (HCV) recombinants have been reported. In addition, recombination is an important regulatory mechanism of cytopathogenicity for the related pestiviruses. Here we describe recombination of HCV RNA in cell culture leading to production of infectious virus. Initially, hepatoma cells were co-transfected with a replicating JFH1ΔE1E2 genome (genotype 2a) lacking functional envelope genes and strain J6 (2a), which has functional envelope genes but does not replicate in culture. After an initial decrease in the number of HCV positive cells, infection spread after 13-36 days. Sequencing of recovered viruses revealed non-homologous recombinants with J6 sequence from the 5' end to the NS2-NS3 region followed by JFH1 sequence from Core to the 3' end. These recombinants carried duplicated sequence of up to 2400 nucleotides. HCV replication was not required for recombination, as recombinants were observed in most experiments even when two replication incompetent genomes were co-transfected. Reverse genetic studies verified the viability of representative recombinants. After serial passage, subsequent recombination events reducing or eliminating the duplicated region were observed for some but not all recombinants. Furthermore, we found that inter-genotypic recombination could occur, but at a lower frequency than intra-genotypic recombination. Productive recombination of attenuated HCV genomes depended on expression of all HCV proteins and tolerated duplicated sequence. In general, no strong site specificity was observed. Non-homologous recombination was observed in most cases, while few homologous events were identified. A better understanding of HCV recombination could help identification of natural recombinants and thereby lead to improved therapy. Our findings suggest mechanisms for occurrence of recombinants observed in patients.
Original languageEnglish
JournalP L o S Pathogens (Online)
Volume9
Issue number3
Pages (from-to)e1003228
ISSN1553-7374
DOIs
Publication statusPublished - Mar 2013

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