Sublethal Staphylococcal Enterotoxin B Challenge Model in Pigs To Evaluate Protection following Immunization with a Soybean-Derived Vaccine
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ABSTRACT In an effort to develop a sustainable platform for manufacturing protein-based vaccine candidates, we expressed a triple mutant of staphylococcal enterotoxin B carrying the L45R, Y89A, and Y94A modifications in transgenic soybean seeds (soy-mSEB). Soy-mSEB possessed no detectable superantigen activity in vitro . We found that this soybean-derived, nontoxic mutant of SEB could be stably expressed, stored in seeds for extended periods at room temperature without degradation, and easily purified from contaminating soy proteins. Vaccination of pigs with purified soy-mSEB, or the identical triple mutant expressed in Escherichia coli ( E. coli -mSEB), resulted in high antibody titers against the native toxin in immunized animals. In fact, titers were indistinguishable regardless of the immunogen used, demonstrating the equivalence of soy-mSEB and E. coli -mSEB vaccinations. Antisera from either immunized group were able to block native SEB superantigen activity in an in vitro neutralization assay. Similar results were obtained when immunized animals were challenged with a sublethal dose of native toxin. Significant reductions in toxin-induced serum cytokine levels were observed in soy-mSEB- and E. coli -mSEB-immunized pigs compared to control animals. The reductions in SEB-induced cytokine responses were similar regardless of the immunogen used for vaccination. Surprisingly, however, some clinical symptoms, such as prostration, lethargy, emesis, and/or diarrhea, were still observed in all immunized animals. These studies demonstrate the potential for soybean-derived proteins as a platform technology for sustainable vaccine manufacturing and the usefulness of a sublethal challenge model in pigs for evaluating the efficacy of potential SEB vaccine candidates.Keywords:
Immunogen
Enterotoxigenic Escherichia coli
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
Heat-labile enterotoxin
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The DNA hybridization assay for genes encoding for Escherichia coli enterotoxins was used to examine water specimens in Thailand. In a reconstruction experiment, the DNA hybridization assay was 10(4) times more sensitive than testing random E. coli in the Y-1 adrenal and suckling mouse assays in identifying enterotoxigenic E. coli (ETEC) in water. Drinking and bathing water collected from 2 of 10 different homes of individuals with ETEC-associated diarrhea and 6% (5 of 78) and 11% (11 of 78) of drinking and bathing water samples collected from homes of individuals with diarrhea without ETEC infections, as well as 6% (5 of 77) and 8% (6 of 77) of drinking and bathing water collected from homes in which no inhabitants had diarrhea, were homologous with the DNA probes. Ten E. coli from each of the 31 water specimens which contained bacteria which were homologous with the DNA probes were tested in the Y-1 adrenal and suckling mouse assay. In only 2 of these 31 specimens could ETEC be identified with the standard assays. The DNA hybridization assay is a much more sensitive means of detecting organisms carrying genes coding for enterotoxin production than testing 10 individual colonies in the Y-1 adrenal and suckling mouse assays. This novel application of recombinant DNA technology provides a sensitive method of detecting organisms carrying genes coding for enterotoxin, and this method will be useful in defining the epidemiology of ETEC.
Enterotoxigenic Escherichia coli
DNA–DNA hybridization
Hybridization probe
Verocytotoxin
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The text analysed biological characteristics,molecular mechanisms and application of heat labile(LT)and heat stable(ST)enterotoxin.First of all,different mutants of LT that have been demonstrated as mucosal adjuvants were constructed by site directed mutagenesis and proved to be the excellent candidates of CT.Next,genetically engineered vaccine of fused LT and ST,which was beneficial to preventing and treating diarrhea, was constructed .These results give a base for further research on enterotoxin in ETEC.
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
Heat-labile enterotoxin
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Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
Strain (injury)
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Ten strains of enterotoxigenic Escherichia coli producing heat-labile enterotoxin (LT) were preserved under 12 different conditions. After 1 month, 9 months, and 3 years of preservation, the cultures were recovered and examined for LT production. Preservation of the cultures on Dorset Egg Medium at 4 degrees C and preservation by freezing the cell suspensions in tryptic soy broth with 20% glycerol were found to be suitable preservation methods; all strains were alive for 3 years and had a minimum loss of LT production.
Enterotoxigenic Escherichia coli
Tryptic soy broth
Heat-stable enterotoxin
Heat-labile enterotoxin
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Enterotoxigenic Escherichia coli (ETEC) of infant origin from the Popular Democratic Republic Lao were characterized with respect to there 0 serogroups, biotypes, anti-bioresistances, fimbrial antigens and types of enterotoxins produced. Enterotoxin production was determined by the suckling mice assay, competitive GM1-erythroassay, and cell cultures (CHOK1 and Y1). The presence of genes encoding for the enterotoxins was determined by colony hybridization by using radioactive DNA probes. Profile plasmids from ETEC strains were studied. The plasmids encoding for heat-labile enterotoxin were studied with an acetyl-aminofluorene modified probe.
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
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Enterotoxigenic Escherichia coli
Heat-labile enterotoxin
Heat-stable enterotoxin
Mitomycin C
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The DNA colony hybridization assay was used to identify enterotoxigenic Escherichia coli among E. coli isolated from 803 swine with diarrhea at 10 farms in Thailand. Between 5 September and 8 December 1981, enterotoxigenic E. coli were identified in 40% of 58 litters of piglets under 10 days old and 17% of 29 litters between 10 and 21 days old with diarrhea at farms at four different locations in Thailand. All E. coli that hybridized with one or more of the three enterotoxin gene probes produced heat-labile or heat-stable toxin or both, as determined by testing culture supernatants in the Y1 adrenal and suckling mouse assays. The DNA colony hybridization technique is a specific method of identifying enterotoxigenic E. coli from swine and can be used to further characterize these enteric pathogens.
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
DNA–DNA hybridization
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Enterotoxigenic Escherichia coli (ETEC) strains are important pathogens for humans and farm animals such as pigs. Porcine ETEC strains induce diarrhea through the production of heat-labile enterotoxin (LT) and/or heat-stable enterotoxins (pSTa/STb). Although LT secretion levels differ between porcine ETEC strains, and this has been linked to virulence, it is unclear whether ST secretion levels also differ between porcine ETEC strains. In addition, the molecular mechanism underlying different LT secretion levels has not been elucidated. In this work, multiple porcine ETEC strains were assessed for their capacity to produce and secrete the enterotoxins LT, pSTa, and STb. The strains differed greatly in their capacity to secrete LT, pSTa, and STb. Remarkably, in some strains, periplasmic production did not correlate with their ability to secrete LT, resulting in high periplasmic production and low LT secretion levels. Furthermore, the results indicated that the type II secretion system (T2SS) protein YghG plays a regulatory role in controlling LT secretion levels. These findings highlight YghG as an important mediator of the secretion of the heat-labile enterotoxin LT by porcine ETEC strains and provide better insights into ETEC enterotoxin secretion.IMPORTANCE Enterotoxigenic E. coli strains are a major health concern. Enterotoxins secreted by enterotoxigenic E. coli are crucial for diarrhea induction. Enterotoxin secretion levels differ between strains; however, it is currently unclear what drives these differences. The discrepancy in the production and secretion capacities of enterotoxins in ETEC is important to clarify their function involved in diarrhea induction. Our results further deepen our understanding of how type II secretion system (T2SS) components of ETEC control enterotoxin secretion levels and may lay the foundation for a better understanding of ETEC molecular pathogenesis.
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
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SYNTHESIS OF A HEAT-STABLE ENTEROTOXIN PRODUCED BY A HUMAN STRAIN OF ENTEROTOXIGENICEscherichia coli
Abstract A heat-stable enterotoxin, produced by a human strain of enterotoxigenic Escherichia coli, was synthesized by a solution method. The synthesized peptide has the same biological and physicochemical properties as those of native toxin.
Enterotoxigenic Escherichia coli
Heat-stable enterotoxin
Strain (injury)
Heat-labile enterotoxin
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