The use of specific antibodies to demonstrate the glycocalyx and spatial relationships of a K99-, F41- enterotoxigenic strain of Escherichia coli colonizing the ileum of colostrum-deprived calves.

Abstract

Electron microscopy was used to study the interaction between the glycocalyx of enterotoxigenic Escherichia coli strain 210 (09:K30+;K99-;F41-:H-) and the glycocalyx of epithelial cells in then ileum of experimentally infected newborn colostrum-deprived calves. Fixation of tissues in anti-K30 antibody and ruthenium red was used to stabilize the bacterial glycocalyx so that the spatial relationship between the bacteria and the intestinal epithelial cells could be characterized. When strain 210 was grown in vitro and reacted with anti-K30 antibody prior to staining with ruthenium red, the extensive glycocalyx could be clearly visualized surrounding the bacterial cells. By negative staining, an unidentified pilus was also seen. Sections of ileum from infected calves, which were not fixed in antibody nor stained with ruthenium red, revealed attached bacteria which were surrounded by an electron-translucent zone and no visible bacterial glycocalyx. When ruthenium red staining was used, the bacterial glycocalyx partially collapsed during the dehydration steps of fixation, but could be seen as either a fibrous capsule or an electron-dense accretion on the bacterial cell surface. When ileal tissue was reacted for one hour in anti-K30 antibody before staining with ruthenium red, the bacterial glycocalyx was seen as a discrete electron-dense structure up to 1.0microm thick which was in intimate contact with the glycocalyx of the epithelial cells. The importance of the bacterial exopolysaccharide to microcolony formation on the villi could be clearly visualized.