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Molecular cloning and characterization of the afa-7 and afa-8 gene clusters encoding afimbrial adhesins in Escherichia coli strains associated with diarrhea or septicemia in calves.

ABSTRACT: The afa gene clusters, which encode proteins involved in adhesion to epithelial cells, from Escherichia coli strains associated with urinary and intestinal infections in humans have been characterized. Pathogenic isolates of bovine and porcine origin that possess afa-related sequences have recently been described. We report in this work the cloning and characterization of the afa-7 and afa-8 gene clusters from bovine isolates. Hybridization and sequencing experiments revealed that despite similarity in genetic organization, the afa-7 and afa-8 genes, and the well-characterized afa-3 operon expressed by human-pathogenic isolates, correspond to three different members of the afa family of gene clusters. However, like the afa-3 gene cluster, both the afa-7 and afa-8 gene clusters were found to encode an afimbrial adhesin (AfaE) and an invasin (AfaD). The AfaD peptides encoded by the three gene clusters were only 45% identical, but functional complementation experiments indicated that they belong to the same family of invasins. Hemagglutination and adhesion assays demonstrated that the AfaE-VII and AfaE-VIII adhesins bind to different receptors and that these receptors are not the human decay-accelerating factor recognized to be the receptor of all previously described AfaE adhesins. The AfaE-VIII adhesin is very similar to the M agglutinin of human-uropathogenic strains. We used PCR assays to screen 25 bovine strains for afaD and afaE genes of either the afa-7 or afa-8 gene cluster. The afa-8 gene cluster was highly prevalent in bovine isolates previously reported to carry afa-related sequences (23 of 24 strains), particularly in strains producing cytotoxic necrotizing factors (16 of 16 strains). The location of the afa-8 gene cluster on the plasmids or chromosome of these isolates suggests that it could be carried by a mobile element, facilitating its dissemination among bovine-pathogenic E. coli strains.

SUBMITTER: Lalioui L

PROVIDER: S-EPMC96852 | biostudies-literature | 1999 Oct

REPOSITORIES: biostudies-literature

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Molecular cloning and characterization of the afa-7 and afa-8 gene clusters encoding afimbrial adhesins in Escherichia coli strains associated with diarrhea or septicemia in calves.

Lalioui L L Jouve M M Gounon P P Le Bouguenec C C

Infection and immunity 19991001 10

The afa gene clusters, which encode proteins involved in adhesion to epithelial cells, from Escherichia coli strains associated with urinary and intestinal infections in humans have been characterized. Pathogenic isolates of bovine and porcine origin that possess afa-related sequences have recently been described. We report in this work the cloning and characterization of the afa-7 and afa-8 gene clusters from bovine isolates. Hybridization and sequencing experiments revealed that despite simila ...[more]

PMID: 10496877

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Project description:Operons of the afa family are expressed by pathogenic Escherichia coli strains associated with intestinal and extraintestinal infections in humans and animals. The recently demonstrated heterogeneity of these operons (L. Lalioui, M. Jouve, P. Gounon, and C. Le Bouguénec, Infect. Immun. 67:5048-5059, 1999) was used to develop a new PCR assay for detecting all the operons of the afa family with a single genetic tool. This PCR approach was validated by investigating three collections of human E. coli isolates originating from the stools of infants with diarrhea (88 strains), the urine of patients with pyelonephritis (97 strains), and the blood of cancer patients (115 strains). The results obtained with this single test and those previously obtained with several PCR assays were closely correlated. The AfaE adhesins encoded by the afa operons are variable, particularly with respect to the primary sequence encoded by the afaE gene. The receptor binding specificities have not been determined for all of these adhesins; some recognize the Dr blood group antigen (Afa/Dr(+) adhesins) on the human decay-accelerating factor (DAF) as a receptor, and others (Afa/Dr(-) adhesins) do not. Thus, the afa operons detected in this study were characterized by subtyping the afaE gene using specific PCRs. In addition, the DAF-binding capacities of as-yet-uncharacterized AfaE adhesins were tested by various cellular approaches. The afaE8 subtype (Afa/Dr(-) adhesin) was found to predominate in afa-positive isolates from sepsis patients (75%); it was frequent in afa-positive pyelonephritis E. coli (55.5%) and absent from diarrhea-associated strains. In contrast, Afa/Dr(+) strains (regardless of the afaE subtype) were associated with both diarrhea (100%) and extraintestinal infections (44 and 25% in afa-positive pyelonephritis and sepsis strains, respectively). These data suggest that there is an association between the subtype of AfaE adhesin and the physiological site of the infection caused by afa-positive strains.

Project description:Over the last few years, dramatic increases in our knowledge about diffusely adhering Escherichia coli (DAEC) pathogenesis have taken place. The typical class of DAEC includes E. coli strains harboring AfaE-I, AfaE-II, AfaE-III, AfaE-V, Dr, Dr-II, F1845, and NFA-I adhesins (Afa/Dr DAEC); these strains (i) have an identical genetic organization and (ii) allow binding to human decay-accelerating factor (DAF) (Afa/Dr(DAF) subclass) or carcinoembryonic antigen (CEA) (Afa/Dr(CEA) subclass). The atypical class of DAEC includes two subclasses of strains; the atypical subclass 1 includes E. coli strains that express AfaE-VII, AfaE-VIII, AAF-I, AAF-II, and AAF-III adhesins, which (i) have an identical genetic organization and (ii) do not bind to human DAF, and the atypical subclass 2 includes E. coli strains that harbor Afa/Dr adhesins or others adhesins promoting diffuse adhesion, together with pathogenicity islands such as the LEE pathogenicity island (DA-EPEC). In this review, the focus is on Afa/Dr DAEC strains that have been found to be associated with urinary tract infections and with enteric infection. The review aims to provide a broad overview and update of the virulence aspects of these intriguing pathogens. Epidemiological studies, diagnostic techniques, characteristic molecular features of Afa/Dr operons, and the respective role of Afa/Dr adhesins and invasins in pathogenesis are described. Following the recognition of membrane-bound receptors, including type IV collagen, DAF, CEACAM1, CEA, and CEACAM6, by Afa/Dr adhesins, activation of signal transduction pathways leads to structural and functional injuries at brush border and junctional domains and to proinflammatory responses in polarized intestinal cells. In addition, uropathogenic Afa/Dr DAEC strains, following recognition of beta(1) integrin as a receptor, enter epithelial cells by a zipper-like, raft- and microtubule-dependent mechanism. Finally, the presence of other, unknown virulence factors and the way that an Afa/Dr DAEC strain emerges from the human intestinal microbiota as a "silent pathogen" are discussed.

Dataset Information

Molecular cloning and characterization of the afa-7 and afa-8 gene clusters encoding afimbrial adhesins in Escherichia coli strains associated with diarrhea or septicemia in calves.

Publications

Molecular cloning and characterization of the afa-7 and afa-8 gene clusters encoding afimbrial adhesins in Escherichia coli strains associated with diarrhea or septicemia in calves.

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