Piglet ileal mucus contains protein and glycolipid (galactosylceramide) receptors specific for Escherichia coli K88 fimbriae

Abstract

The aim of this study was to characterize the Escherichia coli K88-specific receptors in mucus from the small intestines of 35-day-old piglets with the isogenic strains E. coli K-12(pMKO05) (K88+) and E. coli K-12(pMK002) (K88-). These strains differed only in that the latter one cannot produce intact K88 fimbriae because of a deletion in the gene coding for the major fimbrial subunit. Adhesion was studied by incubating 3H-labeled bacteria with crude mucus, pronase-treated whole mucus, mucus fractionated by gel filtration, delipidated mucus, or extracted lipids immobilized in microtiter wells. In addition, E. coli strains were tested for adhesion to glycolipids extracted from mucus by overlaying glycolipid chromatograms with I251-labeled bacteria. The recently reported finding that K88 fimbriae bind to glycoproteins in mucus from the piglet small intestine was confirmed in two ways. Pronase treatment of immobilized mucus reduced adhesion by 82%, and adhesion to delipidated mucus was 14 times greater for the K88+ than for the K88strain. E. coli K88+ adhered to several of the fractions collected after gel filtration of crude mucus, including the void volume (Mr, >250,000). Receptor activity specific for the K88 fimbriae was demonstrated in the lipids extracted from mucus, as the neutral lipids contained six times as much receptor activity as the acidic lipid fraction. Specificity was confirmed by demonstrating that adhesion to the total lipids could be inhibited by pretreatment of the immobilized lipids with K88 fimbriae. Relative to K-12 (K88-), the K-12 (K88+) bacterial cells bound more avidly to galactosylceramide when the neutral lipids were separated on thin-layer chromatography plates. No adhesion to lipids in the acidic fraction separated on thin-layer plates was detected. Relative to adhesion of K-12 (K88-), adhesion of K-12 (K88+) to commercially available galactosylceramide immobilized in microtiter wells confirmed the results with the thin-layer plates. It can be concluded that 35-day-old piglet mucus contains both protein and glycolipid receptors specific for K88 fimbriae, the latter being galactosylceramide.

In vitro inhibition of adhesion of enterotoxigenic Escherichia coli K88 to piglet intestinal mucus by egg-yolk antibodies

“…The adhesion of radioactively labelled E. coli cells to the intestinal mucus from 14-day-old piglets was studied by using a modi¢cation of the methods [11,14,17,23]. All adhesion assays were performed in triplicate in multi-well polystyrene tissue culture plates (Linbro, £at bottom, 1.6 cm; Nunc, polysorp).…”

“…Metcalfe et al [12] and Willemsen and deGraaf [13] have demonstrated that the K88-speci¢c receptor in piglet intestinal mucus is a glycoprotein. Blomberg et al [14] later found that piglet ileal mucus contains protein and glycolipid receptors speci¢c for E. coli K88 ¢mbriae. The amount of K88ab receptor in the mucus was shown to be age related [10,12,13], as is susceptibility to E. coli K88 induced diarrhoea, and it was therefore proposed that the presence of receptors in mucus may play a role in the pathogenesis of K88-bearing strains [11,15].…”

The interaction of F4 fimbriae with porcine enterocytes as analysed by surface plasmon resonance

“…Furthermore, the use of SPR to determine the interaction between adhesins and receptors on eukaryotic cells allows easy screening for blocking agents or receptor analogues to prevent infection. Indeed, one of the strategies to prevent ETEC-induced diarrhoea in both humans and animals, is to prevent attachment of the bacteria to their intestinal receptor and subsequent disease [20,[28][29][30][31][32][33][34].…”

Characterization and purification of porcine small intestinal mucus receptor for Escherichia coli K88ac fimbrial adhesin

“…The presence of both high-and low-a¤nity receptors for the K88 adhesin was ¢rst detected on adhesive brush borders by the use of kinetic binding studies [11]. Since then, putative K88 adhesin receptors have been identi¢ed in both intestinal brush border [12,13] and mucus [7,12,14] preparations. These putative receptors have been reported as being either glycoproteins [7,8,12,13] or glycolipids [14].…”