Qualitative analysis of binding specificity of pathogens to Atlantic salmon mucins.

A. Mucins were isolated by isopycnic density gradient centrifugation, and fractions were collected from the bottom of the tube. Fractions were analyzed for carbohydrate content (glycan), DNA content and density. The glycan peak at fractions 8–11 corresponds to the mucins, and for the experiments presented in Figs 3 and 4, the fractions were pooled based on the glycan peak for each sample. B.-D. Examples of pathogen binding patterns to gradient fractions. B. Low avidity, mucin-specific binding of M. viscosa to an Atlantic salmon skin sample. The binding signal is low but exceeds that of the control and follows the mucin glycan peak (fractions 9–11 in this sample). C. High avidity, mucin-specific binding of A. hydrophila to Atlantic salmon gill sample. The binding follows the mucin glycan peak (fractions 7–10), although avidity appears stronger to a low density glycoform of the main mucin peak (i.e. the binding curve is shifted slightly to the right of the main mucin peak). D. Absence of Y. ruckeri binding to a pyloric cecal sample. The binding signal to the sample is lower than to the non-mucin control and does not follow the mucin glycan peak (fractions 9–11). Binding is expressed as signal/noise luminescence. The horizontal dashed lines denote the binding signal of bacteria to the plastic well. The results of all these qualitative binding analyses are summarized in Table 1. Abbreviations: EU count = europium count; Lum = luminescence, expressed as signal/noise (S/N).