Cloning and characterization of the bundle‐forming pilin gene of enteropathogenic Escherichia coli and its distribution in Salmonella serotypes

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Abstract
Bfp, the structural gene of the major repeating bundle-forming pilus (BFP) subunit, was cloned from the enteroadherent factor (EAF) plasmid of enteropathogenic Escherichia coli (EPEC) strain B171 (O111:NM). The bfp open reading frame encoded a 193-amino-acid protein; comparison of this sequence with the biochemically determined N-terminal amino acid sequence showed that the mature pilin protein is comprised of 180 amino acids, that this sequence is similar to other members of the type IV pilin family, and that it is preceded by a 13-amino-acid signal peptide. Expression of the cloned bfp structural gene in an EPEC strain that had been cured of the EAF plasmid yielded a 21,000 dalton protein that co-migrated with the BFP precursor protein. Thus, other genes, probably carried by the EAF plasmid, are required for the maturation of the bfp product and for the production of extracellular pilus filaments. Use of bfp as a hybridization probe showed that homologous sequences are present in all tested EPEC strains and in 13 of 16 tested Salmonella serotypes. Fifty per cent of these bfp probe-sensitive salmonellae exhibited the localized-adherence (LA) phenotype when incubated with tissue culture cell monolayers, a trait previously associated with EAF plasmid-containing EPEC strains. Scanning electron micrographs of a bfp probe-sensitive, LA-positive Salmonella dublin strain showed that it grows as adherent colonies on infected monolayers and that within these colonies, BFP-like fibres form inter-bacterial linkages. For EAF plasmid-containing EPEC strains and for several Salmonella serotypes, BFP expression may lead to the development of adherent colonies on epithelial surfaces early in the infective process.