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Genetic rearrangements of the regions adjacent to genes encoding heat-labile enterotoxins (eltAB) of enterotoxigenic Escherichia coli strains.

ABSTRACT: One of the most common bacterially mediated diarrheal infections is caused by enterotoxigenic Escherichia coli (ETEC) strains. ETEC-derived plasmids are responsible for the distribution of the genes encoding the main toxins, namely, the heat-labile and heat-stable enterotoxins. The origins and transfer modes (intra- or interplasmid) of the toxin-encoding genes have not been characterized in detail. In this study, we investigated the DNA regions located near the heat-labile enterotoxin-encoding genes (eltAB) of several clinical isolates. It was found that the eltAB region is flanked by conserved 236- and 280-bp regions, followed by highly variable DNA sequences which consist mainly of partial insertion sequence (IS) elements. Furthermore, we demonstrated that rearrangements of the eltAB region of one particular isolate, which harbors an IS91R sequence next to eltAB, could be produced by a recA-independent but IS91 sequence-dependent mechanism. Possible mechanisms of dissemination of IS element-associated enterotoxin-encoding genes are discussed.

SUBMITTER: Schlor S

PROVIDER: S-EPMC91829 | biostudies-literature | 2000 Jan

REPOSITORIES: biostudies-literature

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Genetic rearrangements of the regions adjacent to genes encoding heat-labile enterotoxins (eltAB) of enterotoxigenic Escherichia coli strains.

Schlör S S Riedl S S Blass J J Reidl J J

Applied and environmental microbiology 20000101 1

One of the most common bacterially mediated diarrheal infections is caused by enterotoxigenic Escherichia coli (ETEC) strains. ETEC-derived plasmids are responsible for the distribution of the genes encoding the main toxins, namely, the heat-labile and heat-stable enterotoxins. The origins and transfer modes (intra- or interplasmid) of the toxin-encoding genes have not been characterized in detail. In this study, we investigated the DNA regions located near the heat-labile enterotoxin-encoding g ...[more]

PMID: 10618247

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Project description:Heat-labile enterotoxin (LT) is an important virulence factor secreted by some strains of enterotoxigenic Escherichia coli (ETEC). The prototypic human-origin strain H10407 secretes LT via a type II secretion system (T2SS). We sought to determine the relationship between the capacity to secrete LT and virulence in porcine-origin wild type (WT) ETEC strains. Sixteen WT ETEC strains isolated from cases of severe diarrheal disease were analyzed by GM1ganglioside enzyme-linked immunosorbent assay to measure LT concentrations in culture supernatants. All strains had detectable LT in supernatants by 2 h of culture and 1 strain, which was particularly virulent in gnotobiotic piglets (3030-2), had the highest LT secretion level all porcine-origin WT strains tested (P<0.05). The level of LT secretion (concentration in supernatants at 6-h culture) explained 92% of the variation in time-to-a-moribund-condition (R2 = 0.92, P<0.0001) in gnotobiotic piglets inoculated with either strain 3030-2, or an ETEC strain of lesser virulence (2534-86), or a non-enterotoxigenic WT strain (G58-1). All 16 porcine ETEC strains were positive by PCR analysis for the T2SS genes, gspD and gspK, and bioinformatic analysis of 4 porcine-origin strains for which complete genomic sequences were available revealed a T2SS with a high degree of homology to that of H10407. Maximum Likelihood phylogenetic trees constructed using T2SS genes gspC, gspD, gspE and homologs showed that strains 2534-86 and 3030-2 clustered together in the same clade with other porcine-origin ETEC strains in the database, UMNK88 and UMN18. Protein modeling of the ATPase gene (gspE) further revealed a direct relationship between the predicted ATP-binding capacities and LT secretion levels as follows: H10407, -8.8 kcal/mol and 199 ng/ml; 3030-2, -8.6 kcal/mol and 133 ng/ml; and 2534-86, -8.5 kcal/mol and 80 ng/ml. This study demonstrated a direct relationship between predicted ATP-binding capacity of GspE and LT secretion, and between the latter and virulence.

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

Genetic rearrangements of the regions adjacent to genes encoding heat-labile enterotoxins (eltAB) of enterotoxigenic Escherichia coli strains.

Publications

Genetic rearrangements of the regions adjacent to genes encoding heat-labile enterotoxins (eltAB) of enterotoxigenic Escherichia coli strains.

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