Dataset Information

Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

ABSTRACT:

Background

Diarrhea is a prevalent pathological condition frequently associated to the colonization of the small intestine by enterotoxigenic Escherichia coli (ETEC) strains, known to be endemic in developing countries. These strains can produce two enterotoxins associated with the manifestation of clinical symptoms that can be used to detect these pathogens. Although several detection tests have been developed, minimally equipped laboratories are still in need of simple and cost-effective methods. With the aim to contribute to the development of such diagnostic approaches, we describe here two mouse hybridoma-derived single chain fragment variable (scFv) that were produced in E. coli against enterotoxins of ETEC strains.

Methods and findings

Recombinant scFv were developed against ETEC heat-labile toxin (LT) and heat-stable toxin (ST), from previously isolated hybridoma clones. This work reports their design, construction, molecular and functional characterization against LT and ST toxins. Both antibody fragments were able to recognize the cell-interacting toxins by immunofluorescence, the purified toxins by ELISA and also LT-, ST- and LT/ST-producing ETEC strains.

Conclusion

The developed recombinant scFvs against LT and ST constitute promising starting point for simple and cost-effective ETEC diagnosis.

SUBMITTER: Ozaki CY

PROVIDER: S-EPMC4496030 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Publications

Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

Ozaki Christiane Y CY Silveira Caio R F CR Andrade Fernanda B FB Nepomuceno Roberto R Silva Anderson A Munhoz Danielle D DD Yamamoto Bruno B BB Luz Daniela D Abreu Patrícia A E PA Horton Denise S P Q DS Elias Waldir P WP Ramos Oscar H P OH Piazza Roxane M F RM

PloS one 20150708 7

<h4>Background</h4>Diarrhea is a prevalent pathological condition frequently associated to the colonization of the small intestine by enterotoxigenic Escherichia coli (ETEC) strains, known to be endemic in developing countries. These strains can produce two enterotoxins associated with the manifestation of clinical symptoms that can be used to detect these pathogens. Although several detection tests have been developed, minimally equipped laboratories are still in need of simple and cost-effecti ...[more]

PMID: 26154103

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Project description:Enterotoxigenic E. coli (ETEC) are endemic in low-resource settings and cause robust secretory diarrheal disease in children less than five years of age. ETEC cause secretory diarrhea by producing the heat-stable (ST) and/or heat-labile (LT) enterotoxins. Recent studies have shown that ETEC can be carried asymptomatically in children and adults, but how ETEC subvert mucosal immunity to establish intestinal residency remains unclear. Macrophages are innate immune cells that can be exploited by enteric pathogens to evade mucosal immunity, so we interrogated the ability of ETEC and other E. coli pathovars to survive within macrophages. Using gentamicin protection assays, we show that ETEC H10407 is phagocytosed more readily than other ETEC and non-ETEC isolates. Furthermore, we demonstrate that ETEC H10407, at high bacterial burdens, causes nitrite accumulation in macrophages, which is indicative of a proinflammatory macrophage nitric oxide killing response. However, at low bacterial burdens, ETEC H10407 remains viable within macrophages for an extended period without nitrite accumulation. We demonstrate that LT, but not ST, intoxication decreases the number of ETEC phagocytosed by macrophages. Furthermore, we now show that macrophages exposed simultaneously to LPS and LT produce IL-33, which is a cytokine implicated in promoting macrophage alternative activation, iron recycling, and intestinal repair. Lastly, iron restriction using deferoxamine induces IL-33 receptor (IL-33R) expression and allows ETEC to escape macrophages. Altogether, these data demonstrate that LT provides ETEC with the ability to decrease the perceived ETEC burden and suppresses the initiation of inflammation. Furthermore, these data suggest that host IL-33/IL-33R signaling may augment pathways that promote iron restriction to facilitate ETEC escape from macrophages. These data could help explain novel mechanisms of immune subversion that may contribute to asymptomatic ETEC carriage.

Dataset Information

Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

Background

Methods and findings

Conclusion

Publications

Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

Similar Datasets

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