The effects of modified sialic acids on mucus and erythrocytes on influenza A virus HA and NA functions

Karen N. Barnard; Brynn K. Alford-Lawrence; David W. Buchholz; Brian R. Wasik; Justin R. LaClair; Hai Yu; Rebekah Honce; Stefan Ruhl; Petar Pajic; Erin K. Daugherity; Xi Chen; Stacey L. Schultz-Cherry; Hector C. Aguilar; Ajit Varki; Colin R. Parrish

To look more broadly at the possible range of modified Sia present in secreted mucus in different animals, we examined saliva from a number of influenza host species, including human, pig, horse, and dog (Fig. 4A,B) . While the proteins in saliva differ from those found in 215 respiratory mucus, they do contain some of the same heavily glycosylated proteins including mucins like MUC5B (49) (50) (51) . Human saliva was similar to the secreted mucus from NHBE and Erythrocytes (red blood cells, RBCs) express high levels of sialylated surface molecules, primarily on glycophorins, and are used in IAV research to study the interactions of HA binding specificity, determining viral titer through the hemagglutination assay, and inhibition of hemagglutination by antibodies (HAI assay) (52, 53). It has long been known that IAV varies in 230 hemagglutination of RBCs from different species, at least in part due to differences in the Sia linkages present. The structures of HAs with Sia bound often suggest that modification of the C4, 5, 7, and/or 9 positions would influence IAV interactions with Sia. We found that chicken and guinea pig RBCs, which are often used to titer IAV virus and as the standard substrate for HAI assays, contained almost exclusively unmodified Neu5Ac, as did those from humans and 235 dogs (Fig. 4C,D) . In contrast, pig, horse, cow, and sheep RBCs contain high proportions of All rights reserved. No reuse allowed without permission.