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Enterotoxin production and resistance to antimicrobial agents in porcine and bovine Escherichia coli strains.

A select group of porcine and bovine Escherichia coli strains capable of causing diarrheal disease in neonatal pigs and calves, respectively, were tested for enterotoxin production and resistance to 23 different antimicrobial agents. Thirty-four of the 39 porcine strains tested were enterotoxigenic; that is, they synthesized heat-stable (ST), together with heat-labile (LT), enterotoxin; ST toxin alone, or LT toxin alone. Fourteen of the 15 bovine strains tested produced ST toxin only, whereas 1 strain elaborated LT toxin only. All of the strains were multiple drug resistant. Among the porcine strains, 2 were resistant to 6 of the antimicrobial agents, and 1 was resistant to 18 of the drugs. All of these strains were resistant to cloxacillin, lincomycin, and penicillin G. None of them was resistant to chloramphenicol, colistin, gentamicin, polymyxin B, or nalidixic acid. One of the bovine strains was resistant to 7 of the drugs, and 1 strain was resistant to 17 of these antimicrobial agents. All of the bovine strains exhibited resistance to cloxacillin, lincomycin, novobiocin, pencillin G, sulfathiazole, sulfamethizole, and triple sulfa (sulfadiazine, sulfamerazine, and sulfamethazine). None of these strains showed resistance to gentamicin, nalidixic acid, nitrofurazone, or nitrofurantoin.

Animals↗

Improved radioimmunoassay of Staphylococcal enterotoxin a.

A simple solid-phase radioimmunoassay was developed for detecting staphylococcal enterotoxin A (SEA) in food. The method detected 85-100% SEA added to extracts of 6 kinds of foods, and 52-100% SEA added directly to foods at a concentration of 1 ng/g. Assay sensitivity is about 0.3 ng toxin/mL extract. The method is specific for SEA; results are only marginally affected by staphylococcal enterotoxins B and C2 present at concentrations as high as 500 ng/mL extract.

Enterotoxins↗

Incidence of ketamine-induced emesis in cynomologus monkeys (Macaca fascicularis) used for staphylococcal enterotoxin bioassay.

Ten (24%) of 41 cynomologus monkeys (Macaca fascicularis) showed emetic response to 2.5-20 mg/Kg of ketamine injected i.m. Reduction of the levels of ketamine to one half or less of the emetic level resulted in faster recovery from sedation yet provided adequate time for intubation and subsequent intragastric feeding of staphylococcal enterotoxin (SE) in only 6 of the 10 monkeys without emesis. The onset of the first emetic episode with ketamine was similar to that induced by staphylococcal enterotoxin A (SEA). Cynomologus monkeys showing emetic response to ketamine could still be used for SE bioassay if an experimentally determined non-emetic dose for individual monkeys is employed for sedation.

Animals↗

[Comparative data on the ability of Staphylococcus aureus to produce enterotoxins and other biologically active substances].

Of 201 staphylococcal strains tested, the majority had coagulase activity and oxidized mannitol under anaerobic conditions. Among the staphylococci, enterotoxigenous strains that partially or completely lost their enzymatic activity occurred in an insignificant number of cases. Enzymatic activity (plasma-coagulation, the presence of DNase and mannitol fermentation under anaerobic conditions) makes it possible to classify the strains isolated with Staphylococcus aureus. However, these strains did not always exhibit enterotoxigenous properties. Pigment formation is not the leading sign that allows for classifying staphylococci with Staphylococcus aureus or enterotoxigenous strains. The ability to enterotoxin production was found to be inherent in staphylococci whatever the object from which they were isolated. Enterotoxigeneity was mostly observed in the strains isolated during food poisoning. Such strains produced primarily serotype A or B enterotoxins.

Deoxyribonucleases↗

[Study of possibilities of accumulation of S. aureus enterotoxins during the production and storage of processed cheeses].

The conditions of survive of enterogenic staphylococci in models of production and storage of processed cheeses were studied. The possibilities of S.aureus propagation and accumulation of great doses of thermo-stable enterotoxin were observed in cheese mass before processing. It was established probability of detection of staphylococcus enterotoxins in processed cheeses, produced from the poor quality raw materials.

Cheese↗

[Identification of enterotoxins and cytotoxins of Escherichia coli by Vero cell culture and solid-phase hybridization (colony blot)].

We studied 40 E. coli strains from meat and hamburger and 64 strains from stools of people: 14/64 from traveler's diarrhea and 50/64 from infants with and without diarrhea. We want to know if they could be producing of cytotoxin and enterotoxin in Vero culture cells as well as phenotypic and genotypic relationships. The serotypes that we isolated were different than O:157H:7. We found 3 cytotoxic strains, 77 heat labile enterotoxic strains on Vero culture cells, and 19 E. coli strains with cytotoxin and LT enterotoxin. One strain isolated from infant with diarrhea was negative sorbitol but cytotoxic effect and it was O:55 serotype. The colony blot and cytotonic were found in 90 (86.5%) E. coli strains. The sensitivity of colony blot was 93.75%.

Adult↗

Alteration of the microenvironment in plasma membranes of rat enterocytes after Escherichia coli heat stable enterotoxin treatment: effect on protein kinase C activity.

Plasma membranes isolated from Escherichia coli heat stable enterotoxin (STa) treated rat enterocytes were studied in respect to protein kinase C activity and fluidity change. Pretreatment of enterocytes with STa increased the membrane bound protein kinase C activity about 5 fold as compared to control. STa treatment made the membrane more fluid as evident from a higher phospholipid/cholesterol ratio and greater unsaturated fatty acid levels. Moreover, the phase transition temperature of the STa treated membrane appeared to be significantly lower than that of the corresponding control membrane, thereby further indicating a rise in fluidity of the membrane in the former case. Our results, therefore, suggested that following STa enterotoxin treatment an appropriate fluid environment in the rat intestinal cell membrane was essential for the activation of protein kinase C.

Animals↗

Inhibition of antigen-specific T cell activation by staphylococcal enterotoxins.

The staphylococcal enterotoxins SEA, SEB, SEC2, and TSST-1 bind to MHC class II molecules and stimulate polyclonal T cell populations on the basis of the expression of responsive TCR V beta domains. CL-1 is a human T cell clone that is specific for a peptide derived from influenza hemagglutinin (HA 307-319) presented in the context of HLA-DR1. CL-1 expresses the TCR V beta 13.1 domain, and does not respond to SEA, SEB, or TSST-1. This T cell was used to test the effect of nonstimulatory staphylococcal enterotoxins on a response to antigenic peptide. These toxins inhibit peptide-specific activation of CL-1 in a concentration-dependent manner. These toxins also inhibit the response of an HLA-DR1-specific alloreactive T cell clone. This inhibition seems to be a result of impaired access of TCR to the MHC/peptide complex rather than negative signaling by toxin via class II interaction or induction of T cell anergy. SEA, but neither SEB nor TSST-1 impedes avidin access to a biotin group attached to the amino terminus of HA 307-319. SEA partially impairs access of avidin to HA peptide biotinylated at residue 313, but is unable to inhibit avidin access to biotin at residue 318. This demonstrates that SEA binds to HLA-DR molecules that have also bound the antigenic peptide and suggests a topology for the interaction of SEA with class II, whereby the toxin interferes with peptide/MHC-TCR contact.

Amino Acid Sequence↗

Correlation of phospholipase A & enterotoxin production by Salmonella typhimurium with reference to virulence parameters.

Seventy strains of S. typhimurium along with 10 strains each of S. typhi and S. paratyphi B, 3 strains of S. enteritidis and 2 strains of S. senftenberg were examined for phospholipase A (PLA) activity. S. typhimurium positive for PLA activity (17.14%) were subjected to study of other virulence parameters like enterotoxin production, haemolytic activity, surface hydrophobicity and antibiotic sensitivity. A high degree of correlation was observed between PLA activity and enterotoxin production, antibiotic sensitivity and cell surface hydrophobicity respectively.

Child, Preschool↗

Biological activity and complementation of the two peptides of staphylococcal enterotoxin B formed by limited tryptic hydrolysis.

Tryptic hydrolysis after reduction and carboxamidomethylation of staphylococcal enterotoxin B cleaved the single susceptible bond located between the 2 half-cystines of the molecule, Lys-Thr at positions 97 and 98 (Spero, L., Warren, J. R., and Metzger, J. F. (1973) J. Biol. Chem. 248, 7289-7294). The product remained as a single particle but was separated into the two constituent peptides by denaturants, and purification of the two fragments was accomplished by chromatography on CM-cellulose in 8 M urea. Antigenic activity was exhibited by the separated peptides after dialysis of urea solutions against dilute buffer, but was very labile. No emetic activity in rhesus monkeys was found for either separated peptide. The derivative behaved as two random coil peptides in 6 M guanidine hydrochloride but upon removal of guanidine refolded to a single molecular entity. Viscosity and unfolding kinetics in 1.5 M guanidine indicated physical identity of the recombined peptides with the derivative prior to treatment with guanidine. Three biological measures (serological, mitogenic, and emetic activity) were also essentially unaltered for the recombined material. Since these biological activities are dependent upon different aspects of enterotoxin structure, it is concluded that the recombined derivative was restored to its original conformation.

Animals↗

Effect of ketoprofen on Escherichia coli heat-stable enterotoxin-induced diarrhea of calves.

To evaluate the effect of ketoprofen on fecal output during secretory diarrhea, 16 calves were given approximately 200 micrograms of heat-stable Escherichia coli enterotoxin by suckling on 2 occasions. On one day, treatment was not administered. On the other day, either 3 mg of ketoprofen/kg of body weight (n = 8) or 6 mg of ketoprofen/kg (n = 8) was administered 1 hour before and 3 hours after administration of enterotoxin. Fecal output was no different after 8 or 24 hours from calves given 3 mg/kg, but fecal output was less at 8 hours and 24 hours for calves given 6 mg/kg (P = 0.0588), compared with the control day.

Animals↗

Purification & antigenicity of Salmonella newport enterotoxin.

The enterotoxic moiety present in cell free culture supernatant (CFCS) of S. newport strains (MP/120 and BM/704) was purified and antigenically characterised. Purification was achieved to homogeneity by salt precipitation, dialysis and successive gel filtration through Sephadex G-100 and G-200 columns with the help of FPLC set. It was non-dialysable and purified enterotoxin yielded a single protein band in PAGE. It appeared to be of high molecular weight (100 KDa) and highly immunogenic in the rabbit. Antigenically, it was not related to cholera toxin, Shiga toxin or heat-labile enterotoxin of Escherichia coli.

Antibody Formation↗

[Problems in obtaining antiserum to staphylococcal enterotoxin D].

Experimentally were demonstrated the difficulties overcome and the possibilities of producing a Staphylococcus-enterotoxin D antiserum of a 1:80 titer. The product was shown to be suitable for work as well as for further studies. Given are the schemes of immunizing a rabbit serving as a serum producer and those for purifying the antiserum (eliminating heterologous antibodies), its titration with a microimmunodiffusion test, etc. In this connection some problems are referred to concerning the production of the amounts necessary of enterotoxin D, its purification and toxicity for test animals, etc., that are successfully solved by the authors.

Animals↗

Enterotoxin production and plasmid profiles in Salmonella typhimurium isolated from man.

One hundred twenty-six isolates of Salmonella typhimurium from various clinical sources were tested for enterotoxin production and characterization of plasmid profile. Cell-free culture supernates and polymyxin B extracts of all the strains were assayed by rabbit ileal loop and skin permeability tests. Enterotoxic activity was detected in culture supernates of 32 strains. Twenty-one strains by both rabbit ileal loop and skin permeability tests, nine strains by skin permeability test, and two strains by rabbit ileal loop test were positive. Live culture of three enterotoxic strains, positive in culture supernates produced ileal secretion. Polymyxin B extracts from 6 hours and 18 hours broth cultures of all the strains were devoid of enterotoxicity. Ileal mucosa exposed to culture supernate of enterotoxigenic strains showed swollen and blunted villi with submucosal oedema while those exposed to polymyxin B extracts showed shortening of villi and sloughing of epithelial lining. Plasmid profiles of enterotoxigenic strains were heterogenous and grouped into 20 different profiles. No correlation could be established between plasmid profile, R-pattern, and enterotoxin production.

Animals↗

Effect of growth hormone, prolactin and insulin on the release of IL-1 alpha, IFN-gamma and IL-4 by staphylococcal enterotoxin A-stimulated splenocytes.

The regulation by peptide hormones (Growth Hormone, Prolactin, Insulin) of cytokine secretion by splenocytes stimulated with Staphylococcal Enterotoxin A was studied. Growth hormone increases the release of IFN-gamma from splenocytes stimulated with Enterotoxin A by 50% but considerably decreases IL-1 alpha release by 93%. Prolactin decreases the release of IL-1 alpha by 80%, but has no significant effects on IFN-gamma release. Insulin causes a 50% decrease in IFN-gamma and 95% decrease in IL-1 alpha. IL-4 release was not changed. The results are discussed in terms of the possibility of an interesting function for these endocrine peptides which expands their range of biologic activities within the immune system.

Animals↗

Staphylococcal enterotoxin D functions as a human B cell superantigen by rescuing VH4-expressing B cells from apoptosis.

Staphylococcal enterotoxins are potent superantigens, in that they activate T cells bearing specific V beta-chain gene segments. In this study, we analyzed the capacity of staphylococcal enterotoxin D (SED) to function as a B cell superantigen. SED induced T cell-dependent polyclonal proliferation and differentiation of B cells. In the absence of T cells, SED induced survival of B cells uniquely expressing VH4 containing IgM. The mechanism of survival of VH4-expressing B cells appeared to relate to the countering of apoptosis initiated by the engagement of HLA-DR by SED. Analysis of the VH4 gene products expressed by SED-stimulated B cells revealed the usage of six of the known functional VH4 genes with a variety of different CDR3 regions, employing different DH and JH gene segments. Moreover, the sequence analysis identified a possible site for SED binding of VH4 that includes the solvent-exposed surfaces of 3' CDR2/FR3 and/or FR1. Thus, SED appears to function as a unique B cell superantigen by inducing survival of VH4-expressing B cells.

Adult↗

Enterotoxin septic shock protection and deficient T helper 2 cytokine production in growth hormone transgenic mice.

Neuroendocrine hormones have long been thought to play a role in lymphoid development and function. In particular, growth hormone has been shown to mediate thymic development as well as to promote T cell engraftment in severe combined immunodeficiency mice. Murine T helper cells are classified into two subsets based on their cytokine production pattern. Here, we report that transgenic mice for bovine growth hormone show significant alterations in T cell function and decreased capability for cytokine production, an effect that is more acute in T helper cells as measured by their inability to produce IL-4 upon in vivo injection with Staphylococcus aureus enterotoxin B. Furthermore, upon immunization with conventional Ags, growth hormone transgenic mice produce an altered Ig isotype pattern characterized by a response shift from IgG1 in nontransgenic mice to IgG2 in transgenic mice. The impaired T cell responses correlated with survival from septic shock mediated by bacterial enterotoxins. We conclude that growth hormone may have the potential of regulating immune responses in pathologic processes associated with hyperactivation of T cells or with massive cytokine production.

Animals↗

[Comparative study of the synthesis and specificity of enterotoxin from Vibrio cholerae O139 serotype using monoclonal antibodies].

Strains of V. cholerae serovar 0139 showed a higher intensity of multiplication and lesser nutritive requirements and produced 2-5 times higher amounts of enterotoxin during in-depth culturing than cholera vibrios of groups O1 and non-O1. R-forms of V. cholerae were characterized by the highest production of toxin. Dot-immunoanalysis and immunoblotting with monoclonal antibodies demonstrated the identity of cholera toxin and enterotoxin of V. cholerae serovar O139. The authors come to a conclusion that selective efficacy of the developed vaccines against diarrhea caused by V. cholerae serovar O139 depends on the presence of homologous O-antigen in their composition.

Antibodies, Monoclonal↗