DIFFERENCES IN SIALIC ACID-GALACTOSE LINKAGES IN THE CHICKEN EGG AMNION AND ALLANTOIS INFLUENCE HUMAN INFLUENZA-VIRUS RECEPTOR SPECIFICITY AND VARIANT SELECTION

Human influenza viruses are more efficiently isolated by inoculating p atient samples into the amniotic rather than the allantoic cavity of e mbryonated chicken eggs, This type of cultivation selects virus varian ts with mutations around the hemagglutinin (HA) receptor binding site. To understand the molecular basis of these phenomena, we investigated the abundances of sialic acid (SA) linked to galactose (Gal) by the a lpha-2,3 linkage (SA alpha 2,3Gal) and SA alpha 2,6Gal in egg amniotic and allantoic cells and in Madin-Darby canine kidney (MDCK) cells, Us ing SA-Gal linkage-specific lectins (Maackia amurensis agglutinin spec ific for SA alpha 2,6Gal and Sambucus nigra agglutinin specific for SA alpha 2,3Gal), we found SA alpha 2,3Gal in both allantoic and amnioti c cells and SA alpha 2,6Gal in only the amniotic cells, MDCK; cells co ntained both linkages, To investigate how this difference in abundance s of SA alpha 2,3Gal and SA alpha 2,6Gal in allantoic and amniotic cel ls affects the appearance of host cell variants in eggs, we determined the receptor specificities and HA amino acid sequences of two differe nt patient viruses which were isolated and passaged in the amnion or i n the allantois and which were compared with MDCK cell grown viruses, We found that the viruses maintained high SA alpha 2,6Gal specificitie s when grown in MDCK cells or following up to two amniotic passages; h owever, further passages in either the amnion or allantois resulted in the acquisition of, or a complete shift to, SA alpha 2,3Gal specifici ty, depending on the virus strain. This change in receptor specificity was accompanied by the appearance of variants in the population with Leu-to Gln mutations at position 226 in their HA, These findings sugge st that lack of SA alpha 2,6Gal linkages in the allantois of chicken e ggs is a selective pressure for the appearance of host cell variants w ith altered receptor specificities and amino acid changes at position 226.