title: Rapid and highly informative diagnostic assay for H5N1 influenza viruses.
abstract: A highly discriminative and information-rich diagnostic assay for H5N1 avian influenza would meet immediate patient care needs and provide valuable information for public health interventions, e.g., tracking of new and more dangerous variants by geographic area as well as avian-to-human or human-to-human transmission. In the present study, we have designed a rapid assay based on multilocus nucleic acid sequencing that focuses on the biologically significant regions of the H5N1 hemagglutinin gene. This allows the prediction of viral strain, clade, receptor binding properties, low- or high-pathogenicity cleavage site and glycosylation status. H5 HA genes were selected from nine known high-pathogenicity avian influenza subtype H5N1 viruses, based on their diversity in biologically significant regions of hemagglutinin and/or their ability to cause infection in humans. We devised a consensus pre-programmed pyrosequencing strategy, which may be used as a faster, more accurate alternative to de novo sequencing. The available data suggest that the assay described here is a reliable, rapid, information-rich and cost-effective approach for definitive diagnosis of H5N1 avian influenza. Knowledge of the predicted functional sequences of the HA will enhance H5N1 avian influenza surveillance efforts.
authors.affiliation: Stanford Genome Technology Center, Stanford University, Palo Alto, California, United States of America. pourmand@stanford.edu
authors.forename: Nader
authors.initials: N
authors.lastname: Pourmand
authors.forename: Lisa
authors.initials: L
authors.lastname: Diamond
authors.forename: Rebecca
authors.initials: R
authors.lastname: Garten
authors.forename: Julianna P
authors.initials: JP
authors.lastname: Erickson
authors.forename: Jochen
authors.initials: J
authors.lastname: Kumm
authors.forename: Ruben O
authors.initials: RO
authors.lastname: Donis
authors.forename: Ronald W
authors.initials: RW
authors.lastname: Davis
mesh_terms: D000818:Animals; D001483:Base Sequence; D001717:Birds; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics; D006801:Humans; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics* / Q000302:isolation & purification* / Q000472:pathogenicity; D005585:Influenza in Birds / Q000821:virology; D007251:Influenza, Human / Q000175:diagnosis* / Q000635:transmission / Q000821:virology*; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D017423:Sequence Analysis, RNA / Q000379:methods*; D014773:Virology / Q000379:methods
chemical_list: D017931:DNA Primers; D004279:DNA, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus