title: Alterations in receptor-binding properties of swine influenza viruses of the H1 subtype after isolation in embryonated chicken eggs.
abstract: Alterations of the receptor-binding properties of swine influenza A viruses (SIVs) during their isolation in embryonated chicken eggs have not been well studied. In this study, the receptor-binding properties of classical H1 SIVs isolated solely in eggs or Madin-Darby canine kidney (MDCK) cells were examined. Sequencing analysis revealed substitutions of D190V/N or D225G in the haemagglutinin (HA) proteins in egg isolates, whereas MDCK isolates retained HA genes identical to those of the original viruses present in the clinical samples. Egg isolates with substitution of either D190V/N or D225G had increased haemagglutinating activity for mouse and sheep erythrocytes, but reduced activity for rabbit erythrocytes. Additionally, egg isolates with D225G had increased haemagglutination activity for chicken erythrocytes. A direct binding assay using a sialyl glycopolymer that possessed either a 5-N-acetylneuraminic acid (Neu5Ac) alpha2,6galactose (Gal) or a Neu5Acalpha2,3Gal linkage revealed that the egg isolates used in this study showed higher binding activity to the Neu5Acalpha2,3Gal receptor than MDCK isolates. Increased binding activity of the egg isolates to the Neu5Acalpha2,3Gal receptor was also confirmed by haemagglutination assay with resialylated chicken erythrocytes by Galbeta1,3/4GlcNAcalpha2,3-sialyltransferase. These observations were reinforced by flow-cytometric and N-glycan analyses of the erythrocytes. The alpha2,3-linked sialic acids were expressed predominantly on the surface of mouse and sheep erythrocytes. Chicken erythrocytes expressed Neu5Acalpha2,3Gal more abundantly than Neu5Acalpha2,6Gal, and rabbit erythrocytes expressed both 5-N-glycolylneuraminic acid (Neu5Gc) alpha2,6Gal and Neu5Acalpha2,6Gal. Our results demonstrate clearly that classical H1 SIVs undergo alterations in receptor-binding activity associated with an amino acid substitution in the HA protein during isolation and propagation in embryonated chicken eggs.
authors.affiliation: Thailand-Japan Zoonotic Diseases Collaboration Center, Kasetklang, Chatuchak, Bangkok 10900, Thailand.
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Takemae
authors.forename: Ruttapong
authors.initials: R
authors.lastname: Ruttanapumma
authors.forename: Sujira
authors.initials: S
authors.lastname: Parchariyanon
authors.forename: Shuji
authors.initials: S
authors.lastname: Yoneyama
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Hayashi
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Hiramatsu
authors.forename: Nongluk
authors.initials: N
authors.lastname: Sriwilaijaroen
authors.forename: Yuko
authors.initials: Y
authors.lastname: Uchida
authors.forename: Sachiko
authors.initials: S
authors.lastname: Kondo
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Yagi
authors.forename: Koichi
authors.initials: K
authors.lastname: Kato
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Suzuki
authors.forename: Takehiko
authors.initials: T
authors.lastname: Saito
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002642:Chick Embryo / Q000821:virology; D004285:Dogs; D004912:Erythrocytes / Q000737:chemistry; D006386:Hemagglutination Tests; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000737:chemistry; D053118:Influenza A Virus, H1N1 Subtype / Q000254:growth & development / Q000302:isolation & purification / Q000502:physiology*; D051379:Mice; D019158:N-Acetylneuraminic Acid / Q000097:blood / Q000737:chemistry; D051381:Rats; D011991:Receptors, Virus / Q000502:physiology*; D012756:Sheep; D013552:Swine
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus; D011991:Receptors, Virus; D019158:N-Acetylneuraminic Acid