Dataset Information

Proteomic View of Interactions of Shiga Toxin-Producing Escherichia coli with the Intestinal Environment in Gnotobiotic Piglets.

ABSTRACT:

Background

Shiga toxin (Stx)-producing Escherichia coli cause severe intestinal infections involving colonization of epithelial Peyer's patches and formation of attachment/effacement (A/E) lesions. These lesions trigger leukocyte infiltration followed by inflammation and intestinal hemorrhage. Systems biology, which explores the crosstalk of Stx-producing Escherichia coli with the in vivo host environment, may elucidate novel molecular pathogenesis aspects.

Methodology/principal findings

Enterohemorrhagic E. coli strain 86-24 produces Shiga toxin-2 and belongs to the serotype O157:H7. Bacterial cells were scrapped from stationary phase cultures (the in vitro condition) and used to infect gnotobiotic piglets via intestinal lavage. Bacterial cells isolated from the piglets' guts constituted the in vivo condition. Cell lysates were subjected to quantitative 2D gel and shotgun proteomic analyses, revealing metabolic shifts towards anaerobic energy generation, changes in carbon utilization, phosphate and ammonia starvation, and high activity of a glutamate decarboxylase acid resistance system in vivo. Increased abundance of pyridine nucleotide transhydrogenase (PntA and PntB) suggested in vivo shortage of intracellular NADPH. Abundance changes of proteins implicated in lipopolysaccharide biosynthesis (LpxC, ArnA, the predicted acyltransferase L7029) and outer membrane (OM) assembly (LptD, MlaA, MlaC) suggested bacterial cell surface modulation in response to activated host defenses. Indeed, there was evidence for interactions of innate immunity-associated proteins secreted into the intestines (GP340, REG3-?, resistin, lithostathine, and trefoil factor 3) with the bacterial cell envelope.

Significance

Proteomic analysis afforded insights into system-wide adaptations of strain 86-24 to a hostile intestinal milieu, including responses to limited nutrients and cofactor supplies, intracellular acidification, and reactive nitrogen and oxygen species-mediated stress. Protein and lipopolysaccharide compositions of the OM were altered. Enhanced expression of type III secretion system effectors correlated with a metabolic shift back to a more aerobic milieu in vivo. Apparent pathogen pattern recognition molecules from piglet intestinal secretions adhered strongly to the bacterial cell surface.

SUBMITTER: Pieper R

PROVIDER: S-EPMC3686733 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Publications

Proteomic View of Interactions of Shiga Toxin-Producing Escherichia coli with the Intestinal Environment in Gnotobiotic Piglets.

Pieper Rembert R Zhang Quanshun Q Clark David J DJ Parmar Prashanth P PP Alami Hamid H Suh Moo-Jin MJ Kuntumalla Srilatha S Braisted John C JC Huang Shih-Ting ST Tzipori Saul S

PloS one 20130619 6

<h4>Background</h4>Shiga toxin (Stx)-producing Escherichia coli cause severe intestinal infections involving colonization of epithelial Peyer's patches and formation of attachment/effacement (A/E) lesions. These lesions trigger leukocyte infiltration followed by inflammation and intestinal hemorrhage. Systems biology, which explores the crosstalk of Stx-producing Escherichia coli with the in vivo host environment, may elucidate novel molecular pathogenesis aspects.<h4>Methodology/principal findi ...[more]

PMID: 23840478

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Project description:Many pathogens produce the ?-(1-6)-linked poly-N-acetylglucosamine (PNAG) surface polysaccharide that is being developed as a broadly protective antimicrobial vaccine. However, it is unknown whether systemically injected PNAG vaccines or antibodies would provide protective immunity against pathogens confined to the gastrointestinal tract such as Shiga toxin (Stx)-producing Escherichia coli (STEC), an important group of gastrointestinal (GI) pathogens for which effective immunotherapeutics are lacking. To ascertain whether systemic IgG antibody to PNAG impacts this infectious situation, a vaccine consisting of a synthetic nonamer of nonacetylated PNAG, 9GlcNH2, conjugated to the Shiga toxin 1b subunit (9GlcNH2-Stx1b) was produced. Rabbit antibodies raised to the conjugate vaccine were tested for bacterial killing and toxin neutralization in vitro and protection against infection in infant mice. Cell surface PNAG was detected on all 9 STEC isolates tested, representing 6 STEC serogroups, including E. coli O157:H7. Antibody to the 9GlcNH2-Stx1b conjugate neutralized Stx1 potently and Stx2 modestly. For O157:H7 and O104:H4 STEC strains, antibodies elicited by the 9GlcNH2-Stx1b conjugate possessed opsonic killing and bactericidal activity. Following intraperitoneal injection, antibodies to both PNAG and Stx were needed for infant mouse protection against O157 STEC. These antibodies also mediated protection against the Stx2-producing O104:H4 strain that was the cause of a recent outbreak in Germany, although sufficient doses of antibody to PNAG alone were protective against this strain in infant mice. Our observations suggest that vaccination against both PNAG and Stx, using a construct such as the 9GlcNH2-Stx1b conjugate vaccine, would be protective against a broad range of STEC serogroups. IMPORTANCE The presence of poly-N-acetylglucosamine (PNAG) on many pathogens presents an opportunity to target this one structure with a multispecies vaccine. Whether antibodies to PNAG can protect against pathogens confined to the gastrointestinal tract is not known. As Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are serious causes of infection whose virulence is dependent on elaboration of Stx, we prepared a vaccine containing a synthetic nonamer of PNAG (9GlcNH2) conjugated to Shiga toxin 1b subunit (9GlcNH2-Stx1b) to evaluate bacterial killing, toxin neutralization, and protective efficacy in infant mice. All nine (100%) clinical strains of STEC from different serogroups expressed PNAG. Vaccine-induced antibody mediated in vitro killing of STEC and neutralization of both Stx1 and Stx2. Passive administration of antibody to the conjugate showed protection requiring immunity to both PNAG and Stx for O157 strains, although for an O104 strain, antibody to PNAG alone was protective. Immunity to PNAG may contribute to protection against STEC infections.

Dataset Information

Proteomic View of Interactions of Shiga Toxin-Producing Escherichia coli with the Intestinal Environment in Gnotobiotic Piglets.

Background

Methodology/principal findings

Significance

Publications

Proteomic View of Interactions of Shiga Toxin-Producing Escherichia coli with the Intestinal Environment in Gnotobiotic Piglets.

Similar Datasets

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