title: Genetic mapping of the cold-adapted phenotype of B/Ann Arbor/1/66, the master donor virus for live attenuated influenza vaccines (FluMist).
abstract: Cold adapted (ca) B/Ann Arbor/1/66 is the master donor virus for the influenza B (MDV-B) vaccine component of the live attenuated influenza vaccine (FluMist). The six internal genes contributed by MDV-B confer the characteristic cold-adapted (ca), temperature-sensitive (ts) and attenuated (att) phenotypes to the vaccine strains. Previously, it has been determined that the PA and NP segments of MDV-B control the ts phenotype while the att phenotype requires the M segment in addition to PA and NP. Here, we show that the PA, NP and PB2 segments are responsible for the ca phenotype of MDV-B when examined in chicken cell lines. Five loci in three RNA segments, R630 in PB2, M431 in PA and A114, H410 and T509 in NP, are sufficient to allow efficient virus growth at 25 degrees C. Substitution of these five amino acids with wt (wild type) residues completely reverted the MDV-B ca phenotype. Conversely, introduction of these five ca amino acids into B/Yamanashi/166/98 imparted the ca phenotype to this heterologous wt virus. In addition, we also found that the MDV-B M1 gene affected virus replication in chicken cells at 33 and 37 degrees C. Recombinant viruses containing the two MDV-B M1 residues (Q159, V183) replicated less efficiently than those containing wt M1 residues (H159, M183) at 33 and 37 degrees C, implicating the role of the MDV-B M segment to the att phenotype. The complexity of the multigenic signatures controlling the ca, ts and att phenotypes of MDV-B provides the molecular basis for the observed genetic stability of the FluMist vaccines.
authors.affiliation: MedImmune Vaccines, 297 North Bernardo Avenue, Mountain View, CA 94043, USA.
authors.forename: Zhongying
authors.initials: Z
authors.lastname: Chen
authors.forename: Amy
authors.initials: A
authors.lastname: Aspelund
authors.forename: George
authors.initials: G
authors.lastname: Kemble
authors.forename: Hong
authors.initials: H
authors.lastname: Jin
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002642:Chick Embryo; D003080:Cold Temperature; D006801:Humans; D009981:Influenza B virus / Q000145:classification* / Q000235:genetics* / Q000254:growth & development; D007252:Influenza Vaccines; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins; D010641:Phenotype; D012324:RNA-Dependent RNA Polymerase; D011995:Recombination, Genetic; D013696:Temperature; D014613:Vaccines, Attenuated; D014764:Viral Proteins
chemical_list: D007252:Influenza Vaccines; D019590:Nucleocapsid Proteins; C061290:PA protein, influenza viruses; C502893:PB2 protein, influenza virus; D014613:Vaccines, Attenuated; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase