Adherence to human colonocytes of an Escherichia coli strain isolated from severe infantile enteritis: molecular and ultrastructural studies of a fibrillar adhesin

Abstract

Escherichia coli 469-3 (O21:H-) was isolated from a child with severe enteritis. Ultrastructural analysis of the surface of the strain indicated the presence of very fine fimbriae which mediated mannose-resistant hemagglutination of human blood and caused the bacteria to adhere to human epithelial cell lines and to brush borders of isolated human colonic, but not duodenal, enterocytes. A cosmid library of total DNA of the strain, expressed in laboratory strains of E. coli, was screened by a rapid hemadsorption method, and a number of positive clones were identified. Restriction endonuclease fragments specifying mannose-resistant adherence were subcloned from the cosmid DNA of a strongly hemagglutinating clone in a plasmid vector. The identity of the adhesin was confirmed by biochemical, electron-microscopic, and immunological comparisons with the adhesin synthesized by the clinical isolate. It comprised a high-molecular-weight aggregate of a 14,000-dalton subunit protein which bound antiserum raised against the mannose-resistant adhesin of strain 469-3. The adhesin was synthesized by both the clone and the parental strain at growth temperatures above 18 degrees C but by only a fraction of the cells in a pure culture, although all the bacteria which adhered to human cells expressed the protein.