title: Rapid PCR-based molecular pathotyping of H5 and H7 avian influenza viruses.
abstract: While the majority of avian influenza virus (AIV) subtypes are classified as low-pathogenicity avian influenza viruses (LPAIV), the H5 and H7 subtypes have the ability to mutate to highly pathogenic avian influenza viruses (HPAIV) in poultry and therefore are the etiological agents of notifiable AIV (NAIV). It is of great importance to distinguish HPAIV from LPAIV variants during H5/H7 outbreaks and surveillance. To this end, a novel and fast strategy for the molecular pathotyping of H5/H7 AIVs is presented. The differentiation of the characteristic hemagglutinin (HA) protein cleavage sites (CSs) of HPAIVs and LPAIVs is achieved by a novel PCR method where the samples are interrogated for all existing CSs with a 484-plex primer mixture directly targeting the CS region. CSs characteristic for HP or LP H5/H7 viruses are distinguished in a seminested duplex real-time PCR format using plexor fluorogenic primers. Eighty-six laboratory isolates and 60 characterized NAIV-positive clinical specimens from poultry infected with H5/H7 both experimentally and in the field were successfully pathotyped in the validation. The method has the potential to substitute CS sequencing in the HA gene for the determination of the molecular pathotype, thereby providing a rapid means to acquire additional information concerning NAIV outbreaks, which may be critical to their management. The new assay may be extended to the LP/HP differentiation of previously unknown H5/H7 isolates. It may be considered for integration into surveillance and control programs in both domestic and wild bird populations.
authors: Department of Virology, Immunobiology and Parasitology, National Veterinary Institute (SVA), Ulls väg 2B, SE-751 89 Uppsala, Sweden. mikael.leijon@sva.se
authors: Mikael
authors: M
authors: Leijon
authors: Karin
authors: K
authors: Ullman
authors: Susanna
authors: S
authors: Thyselius
authors: Siamak
authors: S
authors: Zohari
authors: Janice C
authors: JC
authors: Pedersen
authors: Amanda
authors: A
authors: Hanna
authors: Sahar
authors: S
authors: Mahmood
authors: Jill
authors: J
authors: Banks
authors: Marek J
authors: MJ
authors: Slomka
authors: Sándor
authors: S
authors: Belák
mesh_terms: D000818:Animals; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics*; D009980:Influenza A virus / Q000145:classification* / Q000302:isolation & purification*; D005585:Influenza in Birds / Q000821:virology*; D058889:Molecular Typing / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011200:Poultry; D014773:Virology / Q000379:methods*
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus