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TleA, a Tsh-like autotransporter identified in a human enterotoxigenic Escherichia coli strain.

ABSTRACT: Enterotoxigenic Escherichia coli (ETEC), a leading cause of acute diarrhea, colonizes the intestine by means of adhesins. However, 15 to 50% of clinical isolates are negative for known adhesins, making it difficult to identify antigens for broad-coverage vaccines. The ETEC strain 1766a, obtained from a child with watery diarrhea in Chile, harbors the colonization factor CS23 but is negative for other known adhesins. One clone, derived from an ETEC 1766a genomic library (clone G10), did not produce CS23 yet was capable of adhering to Caco-2 cells. The goal of this study was to identify the gene responsible for this capacity. Random transposon-based mutagenesis allowed the identification of a 4,110-bp gene that codes for a homologue of the temperature-sensitive hemagglutinin (Tsh) autotransporter described in avian E. coli strains (97% identity, 90% coverage) and that is called TleA (Tsh-like ETEC autotransporter) herein. An isogenic ETEC 1766a strain with a tleA mutation showed an adhesion level similar to that of the wild-type strain, suggesting that the gene does not direct attachment to Caco-2 cells. However, expression of tleA conferred the capacity for adherence to nonadherent E. coli HB101. This effect coincided with the detection of TleA on the surface of nonpermeabilized bacteria, while, conversely, ETEC 1766a seems to secrete most of the produced autotransporter to the medium. On the other hand, TleA was capable of degrading bovine submaxillary mucin and leukocyte surface glycoproteins CD45 and P-selectin glycoprotein ligand 1 (PSGL-1). These results suggest that TleA promotes colonization of the intestinal epithelium and that it may modulate the host immune response.

SUBMITTER: Gutierrez D

PROVIDER: S-EPMC4399053 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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TleA, a Tsh-like autotransporter identified in a human enterotoxigenic Escherichia coli strain.

Gutiérrez Daniela D Pardo Mirka M Montero David D Oñate Angel A Farfán Mauricio J MJ Ruiz-Pérez Fernando F Del Canto Felipe F Vidal Roberto R

Infection and immunity 20150223 5

Enterotoxigenic Escherichia coli (ETEC), a leading cause of acute diarrhea, colonizes the intestine by means of adhesins. However, 15 to 50% of clinical isolates are negative for known adhesins, making it difficult to identify antigens for broad-coverage vaccines. The ETEC strain 1766a, obtained from a child with watery diarrhea in Chile, harbors the colonization factor CS23 but is negative for other known adhesins. One clone, derived from an ETEC 1766a genomic library (clone G10), did not produ ...[more]

PMID: 25712927

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Project description:Enterotoxigenic Escherichia coli (ETEC) is a major diarrheal pathogen in developing countries, where it accounts for millions of infections and hundreds of thousands of deaths annually. While vaccine development to prevent diarrheal illness due to ETEC is feasible, extensive effort is needed to identify conserved antigenic targets. Pathogenic Escherichia coli, including ETEC, use the autotransporter (AT) secretion mechanism to export virulence factors. AT proteins are comprised of a highly conserved carboxy terminal outer membrane beta barrel and a surface-exposed amino terminal passenger domain. Recent immunoproteomic studies suggesting that multiple autotransporter passenger domains are recognized during ETEC infection prompted the present studies.Available ETEC genomes were examined to identify AT coding sequences present in pathogenic isolates, but not in the commensal E. coli HS strain. Passenger domains of the corresponding autotransporters were cloned and expressed as recombinant antigens, and the immune response to these proteins was then examined using convalescent sera from patients and experimentally infected mice.Potential AT genes shared by ETEC strains, but absent in the E. coli commensal HS strain were identified. Recombinant passenger domains derived from autotransporters, including Ag43 and an AT designated pAT, were recognized by antibodies from mice following intestinal challenge with H10407, and both Ag43 and pAT were identified on the surface of ETEC by flow cytometry. Likewise, convalescent sera from patients with ETEC diarrhea recognized Ag43 and pAT, suggesting that these proteins are expressed during both experimental and naturally occurring ETEC infections and that they are immunogenic. Vaccination of mice with recombinant passenger domains from either pAT or Ag43 afforded protection against intestinal colonization with ETEC.Passenger domains of conserved autotransporter proteins could contribute to protective immune responses that develop following infection with ETEC, and these antigens consequently represent potential targets to explore in vaccine development.

Project description:Background:As part of the effort to develop an enterotoxigenic Escherichia coli (ETEC) human challenge model for testing new heat-stable toxin (ST)-based vaccine candidates, a controlled human infection model study based on the ST-producing ETEC strain TW11681 was undertaken. Here, we estimate stool TW11681 DNA concentration and evaluate its association with dose, clinical symptoms, and with levels of antibodies targeting the CfaB subunit of the ETEC Colonization Factor Antigen I and the E. coli mucinase YghJ. Nine volunteers ingested different doses of the strain and were subsequently followed for 9 days with daily stool specimen collection and clinical examination. Stool DNA was purified by using a newly developed microplate-based method, and DNA originating from TW11681 was quantified by using a probe-based quantitative PCR assay. Antibody levels against CfaB and YghJ were measured in serum collected before and 10 and 28 days after TW11681 was ingested by using a bead-based flow cytometry immunoassay. Results:For 6 of the 9 volunteers, the stool TW11681 DNA concentration increased sharply a median 3.5 (range 2-5) days after dose ingestion, peaking at a median of 5.4% (range 3.3-8.2%) of the total DNA in the specimen. The concentration then fell sharply during the subsequent days, sometimes even before the onset of antibiotic treatment. The size or timing of these proliferation peaks did not seem to be associated with the number of TW11681 bacteria ingested, but the 2 volunteers who developed diarrhea and all five who experienced abdominal pains or cramps had these peaks. The 3 volunteers who did not have the proliferation peaks experienced fewer symptoms and they generally had relatively low CfaB- and YghJ-specific antibody levels before ingesting the strain and subsequently weaker responses than the other volunteers afterwards. Conclusions:Since the lack of proliferation peaks appears to be associated with fewer clinical symptoms and lower serum antibody responses to virulence factors of the infecting strain, it may be important to account for proliferation peaks when explaining results from controlled human infection model studies and for improving the accuracy of protective efficacy estimates when testing new ETEC diarrhea vaccine candidates.

Dataset Information

TleA, a Tsh-like autotransporter identified in a human enterotoxigenic Escherichia coli strain.

Publications

TleA, a Tsh-like autotransporter identified in a human enterotoxigenic Escherichia coli strain.

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