Effects of egg-adaptation on the receptor-binding properties of human influenza A and B viruses

Propagation of human influenza viruses in embryonated chicken eggs (CE) res ults in the selection of variants with amino acid substitutions near the re ceptor-binding site of the hemagglutinin (HA) molecule. To evaluate the mec hanisms by which these substitutions enable human virus growth in CE, we st udied the binding of 10 human influenza A (H1N1, H3N2) and B strains, isola ted and propagated solely in MDCK cells, and of their egg-adapted counterpa rts to preparations of cellular membranes, gangliosides, sialylglycoprotein s, and sialyloligosaccharides. All egg adapted variants differed from nonad apted strains by increased binding to the plasma membranes of chorio-allant oic (CAM) cells of CE and by the ability to bind to CAM gangliosides. In ad dition, there was no decrease in affinity for inhibitors within allantoic f luid. These findings indicate that growth of human influenza viruses in CE is restricted because of their inefficient binding to receptors on CAM cell s and that gangliosides can play an important role in virus binding and/or penetration. The effects of the ega-adaptation substitutions on the recepto r-binding properties of the viruses include (i) enhancement of virus bindin g to the terminal Sia(alpha 2-3)Gal determinant (substitutions in HA positi ons 190, 225 of H1N1 strains and in position 186 of H3N2 strains); (ii) a d ecrease of steric interference with more distant parts of the Sia(alpha 2-3 Gal)-containing receptors (a loss of glycosylation sites in positions 163 o f H1 HA and 187 of type B HA); and (iii) enhanced ionic interactions with t he negatively charged molecules due to charged substitutions at the tip of the HA [187, 189, 190 (H1), and 145, 156 (H3)]. Concomitantly with enhanced binding to Sia(alpha 2-3)Gal-terminated receptors, all egg-adapted variant s decreased their affinity for equine macroglobulin, a glycoprotein bearing terminal 6'-sialyl(N-acetyllactosamine)-moieties. (C) 1999 Academic Press.