Bile salts induce expression of the afimbrial LDA adhesin of atypical enteropathogenic Escherichia coli

Abstract

Atypical enteropathogenic Escherichia coli (aEPEC) strains are frequently implicated in infant diarrhoea in developing countries. Not much is known about the adherence properties of aEPEC; however, it has been shown that these strains can adhere to tissue-cultured cells. A chromosomal region designated the locus for diffuse adherence (LDA) confers aEPEC strain 22 the ability to adhere to culture cells. LDA is an afimbrial adhesin that contains a major subunit, LdaG, whose expression is induced on MacConkey agar at 37 degrees C. We hypothesized that the bile salts found in this culture media induce the expression of LdaG. Strain 22 and the LdaG mutant were grown in Luria-Bertani (LB) media in the presence or absence of bile salts and heat-extracted surface-expressed proteins were separated by SDS-PAGE to determine whether expression of the 25 kDa LdaG protein was induced. Western blot analysis with anti-LdaG confirmed that bile salts enhance LdaG expression at 37 degrees C. Adhesion assays on HeLa cells revealed that adhesion in a diffuse pattern of strain 22 increased in the presence of bile salts. We also confirmed that expression of the localized adherence pattern observed in the ldaG mutant required the presence of a large cryptic plasmid found in strain 22 and that this phenotype was not induced by bile salts. At the transcriptional level, the ldaG-lacZ promoter fusion displayed maximum beta-galactosidase activity when the parent strain was grown in LB supplemented with bile salts. Fluorescence Activated Cell Sorting analysis, immunogold labelling electron microscopy and immunofluorescence using anti-LdaG sera confirmed that LDA is a bile salts-inducible surface-expressed afimbrial adhesin. Finally, LdaG expression was induced in presence of individual bile salts but not by other detergents. We concluded that bile salts increase expression of LDA, conferring a diffuse adherence pattern and having an impact on the adhesion properties of this aEPEC strain.