Development of DNA probes for cytotoxin and enterotoxin genes in enteric bacteria

Brazil, G.; Clayton, C. L.; Sekizaki, Tsutomu; Timmis, Kenneth N.

doi:10.1007/bf01941182

Cited by 5 publications

(3 citation statements)

References 18 publications

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“…In conclusion, the 800 bp ClaI-EcoRI fragment, encoding the stn gene can be utilized as a probe in the identification of bacterial strains that encode similar enterotoxins, and study their 91 phylogenetic relationships [18,19]. Overexpression of STN in the T7 RNA polymerase/promoter system should help in understanding the native structure of the protein and study of its mode of action.…”

Section: Properties Of the Stn Gene Productmentioning

confidence: 99%

Location of the enterotoxin gene from Salmonella typhimurium and characterization of the gene products

Chary

1993

FEMS Microbiology Letters

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The enterotoxin gene (stn) in Salmonella typhimurium (Q1 strain) was confined to an 800 bp ClaI-EcoRI genomic DNA fragment (pCE3) that coded for two polypeptides (25 and 12 kDa) under the control of the T7 RNA polymerase/promoter system. The appearance of the 25 kDa protein corresponded to the enterotoxic activity, as determined by elongation of Chinese hamster ovary (CHO) cells, fluid accumulation in rabbit intestinal loops, and altered vascular permeability in rabbit skin. The stn gene products (STN) caused an elevation of intracellular cAMP in CHO cells. These values were at control levels in stn mutants devoid of enterotoxicity, and the 25-kDa protein concurrently disappeared. The biological activity of the heat-labile enterotoxin was blocked by GM1 ganglioside and neutralized by affinity-purified antibodies made against cholera toxin. The 12 kDa protein however was not correlated with an enterotoxic response.

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Section: Properties Of the Stn Gene Productmentioning

confidence: 99%

Location of the enterotoxin gene from Salmonella typhimurium and characterization of the gene products

Chary

1993

FEMS Microbiology Letters

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“…One SLT-II isolate also produced LT-II, as did 7 of 30 isolates subsequently found to have lost SLT activity. Only three of these eight isolates were LT-IIa probe positive, while all were positive with a probe derived from the LT-II toxin locus from isolate 357900 [53], which we have here determined to encode LT-IIc1. These two probes have a 749 bp region in common that encodes parts of the A and B genes, 343 bp of which are 96% identical (A1 gene) and 406 bp of the A2 and B genes that shows only 59% identity.…”

Section: Discussionmentioning

confidence: 79%

Type II Heat-Labile Enterotoxins from 50 Diverse Escherichia coli Isolates Belong Almost Exclusively to the LT-IIc Family and May Be Prophage Encoded

Jobling

Holmes²

2012

PLoS ONE

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Some enterotoxigenic Escherichia coli (ETEC) produce a type II heat-labile enterotoxin (LT-II) that activates adenylate cyclase in susceptible cells but is not neutralized by antisera against cholera toxin or type I heat-labile enterotoxin (LT-I). LT-I variants encoded by plasmids in ETEC from humans and pigs have amino acid sequences that are ≥95% identical. In contrast, LT-II toxins are chromosomally encoded and are much more diverse. Early studies characterized LT-IIa and LT-IIb variants, but a novel LT-IIc was reported recently. Here we characterized the LT-II encoding loci from 48 additional ETEC isolates. Two encoded LT-IIa, none encoded LT-IIb, and 46 encoded highly related variants of LT-IIc. Phylogenetic analysis indicated that the predicted LT-IIc toxins encoded by these loci could be assigned to 6 subgroups. The loci corresponding to individual toxins within each subgroup had DNA sequences that were more than 99% identical. The LT-IIc subgroups appear to have arisen by multiple recombinational events between progenitor loci encoding LT-IIc1- and LT-IIc3-like variants. All loci from representative isolates encoding the LT-IIa, LT-IIb, and each subgroup of LT-IIc enterotoxins are preceded by highly-related genes that are between 80 and 93% identical to predicted phage lysozyme genes. DNA sequences immediately following the B genes differ considerably between toxin subgroups, but all are most closely related to genomic sequences found in predicted prophages. Together these data suggest that the LT-II loci are inserted into lambdoid type prophages that may or may not be infectious. These findings raise the possibility that production of LT-II enterotoxins by ETEC may be determined by phage conversion and may be activated by induction of prophage, in a manner similar to control of production of Shiga-like toxins by converting phages in isolates of enterohemmorhagic E. coli.

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“…The molecular mass markers are indicated on the left, and from top to bottom they are 97. 4 tac promoter with the synthetic modified 3-lactamase or the aerobactin not clear but it seems likely that the S. typhimunium aroA strains have additional mutations which cause the defect in the export process. These strains are not, however, defective in the transport of other outer membrane proteins normally found in wild-type S. typhimurium, and hence, it seems unlikely that any of the "chaperone" proteins of the Sec-mediated protein translocation pathway (for reviews on the Sec pathway, see reference 1) are mutated.…”

Section: Discussionmentioning

confidence: 99%

Construction of stable LamB-Shiga toxin B subunit hybrids: analysis of expression in Salmonella typhimurium aroA strains and stimulation of B subunit-specific mucosal and serum antibody responses

Brahmbhatt

Wehland

et al. 1992

Infect Immun

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The complete Shiga toxin B subunit and two N-terminal segments of the B subunit have been inserted into a cell surface exposed loop of the LamB protein, and expression of the hybrid proteins from three different promoter systems, i.e., (i) an in vitro-inducible tac promoter that provides high-level expression, (ii) the iron-regulated aerobactin promoter presumably induced in vivo under the iron-limiting conditions of the intestinal mucosal environment, and (iii) a synthetic, modified 13-lactamase promoter providing moderate level constitutive expression, has been analyzed in Escherichia coli, SalmoneUla typhimurium, and attenuated antigen carrier strains of S. typhimurium (aroA mutants). The hybrid vaccine strains were used to immunize mice by the oral and intraperitoneal routes. S. typhimurium aroA mutants apparently have a membrane export defect which prevents the transport of LamB and its derivatives across the cytoplasmic membrane. High-level expression of hybrid proteins through use of the tac promoter proved deleterious to the vaccine strains and prevented the production of viable cells at reasonable cell densities. The lower levels of gene expression observed with the tl-lactamase and aerobactin promoters did not have this effect. Immunization of mice with S. typhimurium aroA strains carrying the hybrid genes expressed from these two promoters resulted in significant B subunit-specific mucosal and serum antibody responses. This suggests that such expression systems may be useful when incorporated into candidate antidysentery live oral vaccines for inducing protection against the effect of Shiga toxin in infections caused by Shigella dysenteriae 1 and other Shiga toxinor Shiga-like toxin-producing pathogens. Bacillary dysentery is an invasive disease of the colon in humans and higher primates and is transmitted via the fecal-oral route. The disease is associated with poor hygiene, overcrowding, and stress, and hence, the majority of cases occur in regions of developing countries that have suboptimal sanitation. However, outbreaks have also been associated with war zones and mental institutions in developed countries. The causative agents are various species of Shigella and enteroinvasive strains of Escherichia coli. The current pandemic of bacillary dysentery caused by Shigella dysenteriae serotype 1 is characterized by high fatality rates * Corresponding author. t This paper is dedicated to the memory of J.-P. Lecocq, acclaimed scientist and friend, who is sorely missed.

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Development of DNA probes for cytotoxin and enterotoxin genes in enteric bacteria

Cited by 5 publications

References 18 publications

Location of the enterotoxin gene from Salmonella typhimurium and characterization of the gene products

Location of the enterotoxin gene from Salmonella typhimurium and characterization of the gene products

Type II Heat-Labile Enterotoxins from 50 Diverse Escherichia coli Isolates Belong Almost Exclusively to the LT-IIc Family and May Be Prophage Encoded

Construction of stable LamB-Shiga toxin B subunit hybrids: analysis of expression in Salmonella typhimurium aroA strains and stimulation of B subunit-specific mucosal and serum antibody responses

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