Hemagglutinin residues of recent human A(H3N2) viruses that affect agglutination of chicken erythrocytes

Abstract Background : The hemagglutination remains crucial in diagnosis of influenza virus and for the antigenic characterization of the hemagglutinin (HA) and neuraminidase. However, the human influenza viruses A(H3N2) isolated recently appear to have lost the ability to agglutinate chicken erythrocytes (ER) (RBC). The molecular determinants of this phenomenon are not known. Methods : Two viruses isolated in Paris were studied, since their ability to agglutinate chicken RBC was observed after serial passages either in Madin Darby canine kidney (MDCK) cells or embryonated hen's eggs. Sequencing analyses and hemadsorption assays were performed to demonstrate the role of amino acid substitutions in the HA gene. Results : Sequencing of the HA gene revealed the presence of either the Leu194Ile or the Val226Ile mutation following the phenotypic change. Hemadsorption assays performed following transfection of COS-1 cells, with plasmids expressing wild-type or mutated HA molecules, showed that the Leu194Ile and Val226Ile mutations were responsible for the ability of the HA to bind chicken RBCs. Conclusion : These findings suggest that a valine at position 226 in the HA molecule, found in recent clinical isolates of human A(H3N2) viruses, could be responsible for their inability to agglutinate chicken erythrocytes.