title: Preparation of a standardized, efficacious agricultural H5N3 vaccine by reverse genetics.
abstract: Options for the control of emerging and reemerging H5N1 influenza viruses include improvements in biosecurity and the use of inactivated vaccines. Commercially available H5N2 influenza vaccine prevents disease signs and reduces virus load but does not completely prevent virus shedding after challenge with H5N1 virus. By using reverse genetics, we prepared an H5N3 vaccine whose hemagglutinin is 99.6% homologous to that of A/CK/HK/86.3/02 (H5N1). We used the internal genes of A/PR/8/34 and the H5 of A/Goose/HK/437.4/99 (H5N1) after deletion of basic amino acids from its connecting peptide region. The resulting virus was not lethal to chicken embryos and grew to high HA titers in eggs, allowing preparation of HA protein-standardized vaccine in unconcentrated allantoic fluid. The N3 neuraminidase, derived from A/Duck/Germany/1215/73 (H2N3), permitted discrimination between vaccinated and naturally infected birds. The virus construct failed to replicate in quail and chickens. Similar to parental A/PR/8/34 (H1N1), it replicated in mice and ferrets and spread to the brains of mice; therefore, it should not be used as a live-attenuated vaccine. The H5N3 vaccine, at doses of 1.2 microg HA, induced HI antibodies in chickens and prevented death, signs of disease, and markedly reduced virus shedding after challenge with A/CK/HK/86.3/02 (H5N1) but did not provide sterilizing immunity. Thus, reverse genetics allows the inexpensive preparation of standardized, efficacious H5N3 poultry vaccines that may also reduce the reemergence of H5N1 genotypes.
authors.affiliation: Division of Virology, Department of Infectious Diseases, St. Jude Children's Research Hospital, 332 N. Lauderdale, Memphis, TN 38105, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Liu
authors.forename: John M
authors.initials: JM
authors.lastname: Wood
authors.forename: Trevor
authors.initials: T
authors.lastname: Ellis
authors.forename: Scott
authors.initials: S
authors.lastname: Krauss
authors.forename: Patrick
authors.initials: P
authors.lastname: Seiler
authors.forename: Christie
authors.initials: C
authors.lastname: Johnson
authors.forename: Erich
authors.initials: E
authors.lastname: Hoffmann
authors.forename: Jennifer
authors.initials: J
authors.lastname: Humberd
authors.forename: Diane
authors.initials: D
authors.lastname: Hulse
authors.forename: Yun
authors.initials: Y
authors.lastname: Zhang
authors.forename: Robert G
authors.initials: RG
authors.lastname: Webster
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Perez
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D001483:Base Sequence; D002645:Chickens; D004372:Ducks; D005260:Female; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics / Q000276:immunology; D009980:Influenza A virus / Q000235:genetics / Q000276:immunology*; D007252:Influenza Vaccines / Q000008:administration & dosage / Q000009:adverse effects / Q000235:genetics / Q000592:standards*; D005585:Influenza in Birds / Q000517:prevention & control*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D011201:Poultry Diseases / Q000517:prevention & control*; D011784:Quail; D011995:Recombination, Genetic
chemical_list: D000914:Antibodies, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D007252:Influenza Vaccines