title: Cleavage site stability of Egyptian highly pathogenic avian influenza viruses in backyard chickens during 2009-2011.
abstract: OBJECTIVE
  Two distinguishable subclades of H5N1 (classic and variant strains) are cocirculating among the poultry populations in Egypt despite the intensive vaccination programs. A study to investigate the genetic relationship between avian influenza virus (AIV) isolates from backyard chickens in Sharkia (2009-2011), subclades, and commercially available vaccines was carried out.
  
  
  METHODS
  Forty-eight suspected AIV infected birds were clinically examined and used for virus isolation followed by reverse transcription-polymerase chain reaction. Four H5N1 virus isolates were sequenced and analyzed. The intravenous pathogenicity index (IVPI) of three AIV isolates was determined.
  
  
  RESULTS
  Thirty-four hemagglutinating viral agents (30 AIV subtype H5N1 and 4 Newcastle disease virus) were detected. Both the nucleotide and amino acid sequence identities of four H5N1 virus isolates (SHZA-0412/2009, SHZA-0801/2010, SHMK-1903/2010, and SHAH-1403/2011) were high--98.4-99.7% and 100%, respectively--indicative of their genetic homogeneity. The hemagglutinin cleavage site characterization revealed the presence of multiple basic amino acids (-PQRERRRKKR/GL-) of the highly pathogenic phenotype. These results were supported by IVPI in chickens of 2.69-2.90. The similarity of our isolates with H5N1 AIV vaccine strains (93.9-95.1%) was higher than that with H5N2 strains (77.8-91.9%). The divergence of four sequences with classic and variant lineages is 2-2.7% and 2.3-3%, respectively, with two amino acid substitutions (A249P and N251Y).
  
  
  CONCLUSIONS
  Genetic characterization and IVPI data of backyard H5N1 isolates are indicative of a highly pathogenic avian influenza virus with hemagglutinin cleavage site constancy and two amino acids substitutions with Egyptian classic and variant lineages, suggesting a beginning of antigenic drift.
authors.affiliation: Department of Avian and Rabbit Medicine, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
authors.forename: Reham M
authors.initials: RM
authors.lastname: ElBakrey
authors.affiliation: Department of Avian and Rabbit Medicine, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
authors.forename: Mohammed A
authors.initials: MA
authors.lastname: El Sisi
authors.affiliation: Department of Virology, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
authors.forename: Shimaa M G
authors.initials: SM
authors.lastname: Mansour
authors.affiliation: Department of Zoonoses, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
authors.forename: Heba H
authors.initials: HH
authors.lastname: Ahmed
authors.affiliation: Department of Veterinary and Biomedical Sciences, South Dakota State University, Brookings, SD 57007, USA.
authors.forename: Mrigendra
authors.initials: M
authors.lastname: Rajput
authors.affiliation: Department of Avian and Rabbit Medicine, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt. Electronic address: amalaeid@yahoo.com.
authors.forename: Amal A M
authors.initials: AA
authors.lastname: Eid
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002645:Chickens; D004534:Egypt / Q000453:epidemiology; D016679:Genome, Viral; D042822:Genomic Instability; D005838:Genotype; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics*; D053124:Influenza A Virus, H5N1 Subtype / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D005585:Influenza in Birds / Q000453:epidemiology / Q000473:pathology / Q000821:virology*; D017720:Molecular Epidemiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus
keywords: Backyard chicken; Egypt; H5N1; Hemagglutinin gene; Highly pathogenic avian influenza