title: Internal Disequilibria and Phenotypic Diversification during Replication of Hepatitis C Virus in a Noncoevolving Cellular Environment.
abstract: Viral quasispecies evolution upon long-term virus replication in a noncoevolving cellular environment raises relevant general issues, such as the attainment of population equilibrium, compliance with the molecular-clock hypothesis, or stability of the phenotypic profile. Here, we evaluate the adaptation, mutant spectrum dynamics, and phenotypic diversification of hepatitis C virus (HCV) in the course of 200 passages in human hepatoma cells in an experimental design that precluded coevolution of the cells with the virus. Adaptation to the cells was evidenced by increase in progeny production. The rate of accumulation of mutations in the genomic consensus sequence deviated slightly from linearity, and mutant spectrum analyses revealed a complex dynamic of mutational waves, which was sustained beyond passage 100. The virus underwent several phenotypic changes, some of which impacted the virus-host relationship, such as enhanced cell killing, a shift toward higher virion density, and increased shutoff of host cell protein synthesis. Fluctuations in progeny production and failure to reach population equilibrium at the genomic level suggest internal instabilities that anticipate an unpredictable HCV evolution in the complex liver environment.IMPORTANCELong-term virus evolution in an unperturbed cellular environment can reveal features of virus evolution that cannot be explained by comparing natural viral isolates. In the present study, we investigate genetic and phenotypic changes that occur upon prolonged passage of hepatitis C virus (HCV) in human hepatoma cells in an experimental design in which host cell evolutionary change is prevented. Despite replication in a noncoevolving cellular environment, the virus exhibited internal population disequilibria that did not decline with increased adaptation to the host cells. The diversification of phenotypic traits suggests that disequilibria inherent to viral populations may provide a selective advantage to viruses that can be fully exploited in changing environments.
authors.affiliation: Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Elena
authors.initials: E
authors.lastname: Moreno
authors.affiliation: Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Isabel
authors.initials: I
authors.lastname: Gallego
authors.affiliation: Liver Unit, Internal Medicine Hospital Universitari Vall d'Hebron, Vall d'Hebron Institut de Recerca (VHIR), Barcelona, Spain.|Roche Diagnostics, S.L., Sant Cugat del Valles, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Gregori
authors.affiliation: Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Adriana
authors.initials: A
authors.lastname: Lucía-Sanz
authors.affiliation: Liver Unit, Internal Medicine Hospital Universitari Vall d'Hebron, Vall d'Hebron Institut de Recerca (VHIR), Barcelona, Spain.
authors.forename: María Eugenia
authors.initials: ME
authors.lastname: Soria
authors.affiliation: Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Victoria
authors.initials: V
authors.lastname: Castro
authors.affiliation: Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Nathan M
authors.initials: NM
authors.lastname: Beach
authors.affiliation: Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Susanna
authors.initials: S
authors.lastname: Manrubia
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Internal Medicine Hospital Universitari Vall d'Hebron, Vall d'Hebron Institut de Recerca (VHIR), Barcelona, Spain.|Universitat Autonoma de Barcelona, Barcelona, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Quer
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Internal Medicine Hospital Universitari Vall d'Hebron, Vall d'Hebron Institut de Recerca (VHIR), Barcelona, Spain.|Universitat Autonoma de Barcelona, Barcelona, Spain.
authors.forename: Juan Ignacio
authors.initials: JI
authors.lastname: Esteban
authors.affiliation: Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, Rockefeller University, New York, New York, USA.
authors.forename: Charles M
authors.initials: CM
authors.lastname: Rice
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.|Instituto de Parasitología y Biomedicina "López-Neyra" (CSIC), Parque Tecnológico Ciencias de la Salud, Armilla, Granada, Spain.
authors.forename: Jordi
authors.initials: J
authors.lastname: Gómez
authors.affiliation: Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Gastaminza
authors.affiliation: Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain edomingo@cbm.csic.es cperales@cbm.csic.es.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Esteban
authors.initials: E
authors.lastname: Domingo
authors.affiliation: Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain edomingo@cbm.csic.es cperales@cbm.csic.es.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd) del Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Internal Medicine Hospital Universitari Vall d'Hebron, Vall d'Hebron Institut de Recerca (VHIR), Barcelona, Spain.
authors.forename: Celia
authors.initials: C
authors.lastname: Perales
mesh_terms: D000220:Adaptation, Biological / Q000235:genetics; D006528:Carcinoma, Hepatocellular / Q000821:virology*; D004261:DNA Replication; D019143:Evolution, Molecular; D016679:Genome, Viral; D016174:Hepacivirus / Q000145:classification / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D008099:Liver / Q000821:virology; D009154:Mutation; D010641:Phenotype; D012367:RNA, Viral / Q000235:genetics; D014779:Virus Replication
chemical_list: D012367:RNA, Viral
keywords: RNA virus; mutant spectrum; population instability; quasispecies