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PubMed1974

Enterotoxin production by strains of Staphylococcus aureus isolated from foods and human beings.

Enterotoxin production by strains of Staphylococcus aureus isolated from routine samples of foods and from human beings was investigated. Twenty-one to 26% of 112 strains isolated from raw meat, sausages and poultry and 32-36% of 183 strains isolated from cooked foods, e.g. meat, chicken and frozen seafoods, produced enterotoxins A, B, C, D or E. Staph. aureus isolated from raw meat and chicken less frequently produced enterotoxins A, B, C or E and more frequently enterotoxin D, than those from cooked meat and seafoods. Of the 113 strains isolated from cheese and raw milk 6-11% produced enterotoxin and most of these produced enterotoxin D. Only a few strains isolated from foods produced enterotoxin E. Results of enterotoxin tests on Staph. aureus from human beings resembled those on strains from cooked foods.

Bacteriological Techniques↗

PubMed1983

Enterotoxin production, phage typing and serotyping of Staphylococcus aureus strains isolated from clinical materials and food.

The production of enterotoxins A, B, C and F by strains of Staphylococcus aureus isolated from various clinical sources and from isolates implicated in food poisoning was investigated. One hundred and ninety one of the 374 clinical strains (51.1%) were found to be enterotoxigenic; of these, 81 (27.7%) strains produced enterotoxin A, 57 (15.3%) strains produced enterotoxin B, 23 (6.2%) strains produced enterotoxin C, and 64 (17.1%) strains produced enterotoxin F. These enterotoxigenic strains were most frequently lysed by phages of group III (21.5%) or were not typable (22%). Eighteen of the 29 strains implicated in food poisoning were enterotoxigenic. The correlation of antigens and bacteriophage patterns with enterotoxigenicity was determined: enterotoxin A being related to a4 antigen, enterotoxin B to phages of 94/96 complex with c1, o antigens, and enterotoxin F to phages of group I with 2632, k1k2, m antigens.

Bacterial Toxins↗

PubMed1996

Nasal carriage of enterotoxin-producing Staphylococcus aureus among restaurant workers in Kuwait City.

Enterotoxin-producing Staphylococcus aureus is a common cause of staphylococcal food poisoning. To determine the incidence of carriage of enterotoxin-producing S. aureus in a sample of the healthy population in Kuwait city, restaurant workers in the city were screened for nasal carriage of S. aureus. 26.6% of 500 workers studied carried S. aureus and 86.6% of the S. aureus produced staphylococcal enterotoxins. 28% produced enterotoxin A, 28.5% produced enterotoxin B, 16.4% produced enterotoxin C and 3.5% produced enterotoxin D. Ten isolates produced both enterotoxins A and B or A and C. 73% of the isolates were untypeable with standard phages. However, 17.1%, 3% and 6% belonged to phage groups I, II and III respectively. The results demonstrated a high level of enterotoxigenic S. aureus carriage among restaurant workers which although lower than that reported for the general population and hospital workers may be important in the restaurant industry.

Bacteriophage Typing↗

PubMed2001

Antagonistic effects of the staphylococcal enterotoxin a mutant, SEA(F47A/D227A), on psoriasis in the SCID-hu xenogeneic transplantation model.

Psoriasis is a T-cell-mediated immune dermatosis probably triggered by bacterial superantigens. This pathomechanism has been experimentally reproduced in a SCID-hu xenogeneic transplantation model. We analyzed the effects of different bacterial superantigens on the induction of psoriasis in this model. Staphylococcal enterotoxin B and exfoliative toxin triggered the onset of psoriasis when administered repetitively intracutaneously over a period of 2 wk, whereas staphylococcal enterotoxin A representing a distinct subfamily of staphylococcal enterotoxins only mimicked certain aspects of psoriasis. The biologic effects of staphylococcal enterotoxin A were more pronounced when a mutated form, SEA(H187A), of this superantigen with reduced affinity to major histocompatibility complex class II was coinjected. Another mutated variant, SEA(F47A/D227A), exhibiting no measurable major histocompatibility complex class II affinity blocked the effects triggered by wild-type staphylococcal enterotoxin A when injected in a 10-fold higher dose. Inhibition was specific as induction of psoriasiform epidermal changes by staphylococcal enterotoxin B could not be blocked. As staphylococcal enterotoxin A, in contrast to the other superantigens tested, is capable of inducing epidermal thickening but not the typical appearance of psoriasis, we conclude that bacterial superantigens may differ with regard to their effects on human nonlesional psoriatic skin. Staphylococcal-enterotoxin-A-mediated effects were blocked by a genetically engineered superantigen highlighting the potential therapeutic use of mutated superantigens.

Animals↗

PubMed2005

Nosocomial antibiotic-associated diarrhea associated with enterotoxin-producing strains of methicillin-resistant Staphylococcus aureus.

OBJECTIVES: The aim of this study is to present new evidence that enterotoxin-producing strains of methicillin-resistant Staphylococcus aureus may cause nosocomial antibiotic-associated diarrhea. METHODS: We conducted a prospective study that utilized standard methods to exclude other bacterial, parasitic, and viral pathogens as causes of nosocomial diarrhea in patients with heavy growth of methicillin-resistant S. aureus in their stool. Staphylococcal enterotoxin assays were performed on S. aureus strains recovered from patients' stools and on stool specimens from affected patients. Retrospective cohort studies compared the severity of diarrhea in patients with methicillin-resistant S. aureus-associated diarrhea with that of patients whose stool did not contain the organism and with patients colonized or infected with enterotoxin-negative methicillin-resistant S. aureus strains. RESULTS: During an 18-month period, 11 patients had nosocomial antibiotic-associated diarrhea associated with enterotoxin-producing strains of methicillin-resistant S. aureus. Other common bacterial, parasitic, and viral pathogens were excluded. S. aureus strains from the 11 patients produced staphylococcal enterotoxin A, A and B, or D. Eighty-nine percent of patients had the same enterotoxin(s) in stool specimens as produced by the strain recovered from their stool. Case patients had a greater number of days of diarrhea than patients without methicillin-resistant S. aureus in their stool (p < 0.001), or randomly selected patients colonized or infected with enterotoxin-negative methicillin-resistant S. aureus (p < 0.001). CONCLUSIONS: Our findings provide evidence that enterotoxin-producing strains of methicillin-resistant S. aureus may cause nosocomial antibiotic-associated diarrhea. Greater recognition of this disease should result in more rapid and appropriate treatment of affected patients.

Anti-Bacterial Agents↗

PubMed2007

Staphylococcus aureus-derived enterotoxins enhance house dust mite-induced patch test reactions in atopic dermatitis.

BACKGROUND: Up to 65% of Staphylococcus aureus strains isolated from the skin of patients with atopic dermatitis (AD) produce exotoxins with superantigenic properties that may also act as allergens leading to an induction of exotoxin-specific IgE antibodies. Staphylococcal enterotoxin B (SEB) applied epicutaneously in a concentration of 10 micro g/cm(2), i.e. 200 micro g/ml, under occlusion induces cutaneous inflammation in patients with AD and healthy individuals. METHODS: We performed patch tests in 32 adult patients with AD using different concentrations (i.e. 2-200 micro g/ml) of SEA, SEB and house dust mite (HDM) extract (500 micro g/ml). Furthermore, the respective enterotoxins and HDM extract were applied simultaneously to the same patch test site. Specific IgE levels to SEA, SEB and HDM were measured with the CAP FEIA. RESULTS: The rates of patch test reactions to SEA and SEB increased with rising enterotoxin concentrations. There were no differences in the rates of patch test reactions to SEA and SEB between patients sensitized to the corresponding enterotoxin and non-IgE-sensitized patients. The number of patch test reactions to the mixture of enterotoxin and HDM extract was higher than the number of patch test reactions to either the enterotoxins or HDM extract. We identified 11 patients with AD who reacted neither to the enterotoxins nor to HDM extract, but who showed patch test reactions to the mixture. These reactions were not predicted by the presence of enterotoxin- or HDM-specific IgE. CONCLUSIONS: Colonization with exotoxin-producing S. aureus may influence the outcome of patch tests in patients with AD.

Adolescent↗

PubMed1976

Effect of starter culture on staphylococcal enterotoxin and thermonuclease production in dry sausage.

Different amounts of enterotoxin A-, B-, and C1-producing staphylococci were added to dry sausage prepared by normal processes, either alone or in conjunction with a starter culture (micrococci and lactobacilli). The sausage was examined after 0, 3, 7, 14, and 30 days for staphylococci, micrococci, and lactobacilli, and measurements were made of water activity, pH, enterotoxin, and thermostable nuclease. The results showed that in the absence of starter culture measurable amounts of enterotoxin A were formed in a 200-g sample of dry sausage in 3 days, the level of Staphylococcus aureus infection being over 10(6) cells/g. Enterotoxin B was not found, although the total number of staphylococci was over 10(8) cells/g. Enterotoxin C1 was observed when the Staphylococcus count was about 8 X 10(7) cells/g, but was no longer detectable after 7 days. The starter culture prevented the production of enterotoxin A in all cases investigated. By contrast, a very high-level inoculation of an enterotoxin C1-producing strain gave a positive result after 3 days of incubation even in the presence of a starter culture. Heat-stable nuclease was observed in all sausages to which enterotoxin-producing staphylococci were added. The cell count determined in a sample of sausage had no definite correlation with the thermonuclease activity of the sample.

Antibiosis↗

PubMed1976

Comparison of different purification procedure for extraction of staphylococcal enterotoxin A from foods.

Different procedures commonly used for extraction, purification, and concentration of staphylococcal enterotoxins from foods were investigated with 131I- and 125I-labeled staphylococcal enterotoxin A. Loss of labeled enterotoxin A was compared with loss of total nitrogen. The results showed that in most of the common procedures, such as gel filtration, ion exchange, and heat treatment, the percentage of loss of labeled enterotoxin A was greater than the loss of total nitrogen. Chloroform extraction and acid precipitation with hydrochloric acid had nearly the same effect on the purification of both labeled enterotoxin A and total nitrogen. Ammonium sulfate precipitation proved to be practical and was successfully used for purification of enterotoxin A from sausage extract. Simultaneous use of trypsin and Pseudomonas peptidase for treatment of food extracts considerably reduced food proteins capable of interfering with serological detection of enterotoxins but did not essentailly influence the loss of enterotoxin A.

Enterotoxins↗

PubMed1981

Endogenous radiolabeling of enterotoxin from Clostridium perfringens type A on a defined medium.

Four enterotoxin-positive strains of Clostridium perfringens were tested for sporulation and enterotoxin production on defined media. The medium described by Sacks and Thompson (Appl. Environ. Microbiol. 35:405-409, 1978) gave the highest enterotoxin production and was selected for the production of endogenously labeled enterotoxin. The specific radioactivity of the enterotoxin was 16,000 dpm/microgram when the tritiated amino acids were added to the growth medium just before the inoculum. Addition of the radioactive amino acids during the growth period gave consistently lower specific radioactivity. When the enterotoxin was produced on the medium described by Duncan and Strong (Appl. Microbiol. 16:82-89, 1968), the highest specific radioactivity of the enterotoxin was found when the radioactive amino acids were added to the growth medium 4 h after inoculation. In this case, the specific activity of the enterotoxin was 10,000 dpm/microgram.

Amino Acids↗

PubMed1987

Production of staphylococcal enterotoxin in mixed cultures.

Two Staphylococcus aureus strains were grown in brain-heart infusion (BHI) broth and a meat medium with Bacillus cereus, Streptococcus faecalis, Escherichia coli, and Pseudomonas aeruginosa. Both S. aureus strains grew well and produced enterotoxin in the presence of S. faecalis in BHI broth; however, enterotoxin production was observable in the meat medium only when the S. aureus inoculum was greater than the S. faecalis inoculum. S. aureus FRI-100 grown with B. cereus produced enterotoxin in both media only when the S. aureus inoculum was much higher than the B. cereus inoculum (10 versus 10(4) CFU), whereas S. aureus FRI-196E produced enterotoxin in both media at all inoculum combinations except in the meat medium, when the inocula of the two organisms were the same. S. aureus grown with E. coli in BHI broth produced enterotoxin at all inoculum combinations except when the E. coli inoculum was greater than the S. aureus inoculum; however, in the meat medium, enterotoxin was produced only when the S. aureus inoculum was much greater than the E. coli inoculum (10 versus 10(4) CFU), S. aureus FRI-100 grown with P. aeruginosa in either medium produced enterotoxin only when the S. aureus inoculum was much greater than the P. aeruginosa inoculum (10 versus 10(3) or 10(4) CFU). It can be concluded from these results that enterotoxin production is unlikely in mixed cultures unless the staphylococci outnumber the other contaminating organisms.

Bacillus cereus↗

PubMed1997

Cloning of novel enterotoxin genes from Bacillus cereus and Bacillus thuringiensis.

A novel enterotoxin gene was cloned from Bacillus cereus FM1, and its nucleotide sequence was determined. Previously, a 45-kDa protein causing characteristic enterotoxin symptoms in higher animals had been isolated (K. Shinagawa, p. 181-193, in A. E. Pohland et al., ed., Microbial Toxins in Foods and Feeds, 1990) from the same B. cereus strain, but no report of cloning of the enterotoxin gene has been published. In the present study, a specific antibody to the purified enterotoxin was produced and used to screen the genomic library of B. cereus FM1 made with the lambda gt11 vector. An immunologically positive clone was found to contain the full protein-coding region and some 5' and 3' flanking regions. The deduced amino acid sequence of the cloned gene indicated that the protein is rich in beta structures and contains some unusual sequences, such as consecutive Asn residues. In order to clone enterotoxin genes from Bacillus thuringiensis, two PCR primers were synthesized based on the nucleotide sequence of the B. cereus gene. These primers were designed to amplify the full protein-coding region. PCR conducted with DNA preparations from the B. thuringiensis subsp. sotto and B. thuringiensis subsp. israelensis strains successfully amplified a segment of DNA with a size almost identical to that of the protein-coding region of the B. cereus enterotoxin. Nucleotide sequences of the amplified DNA segments showed that these B. thuringiensis strains contain an enterotoxin gene very similar to that of B. cereus. Further PCR screening of additional B. thuringiensis strains with four primer pairs in one reaction revealed that some additional B. thuringiensis strains contain enterotoxin-like genes.

Amino Acid Sequence↗

PubMed1971

Staphylococcal enterotoxin B: solid-phase radioimmunoassay.

An immunoassay employing (125)I-labeled enterotoxin B and polystyrene tubes coated with specific antibody was used for assaying purified and crude enterotoxin. Antibody was adsorbed to untreated polystyrene tubes. Unlabeled enterotoxin competed with (125)I-labeled enterotoxin for antibody-combining sites. The uptake of (125)I-labeled toxin reflected the concentration of unlabeled toxin present. The test is sensitive to 1 to 5 ng of purified and crude enterotoxin B per ml, and cross-reactions with heterologous enterotoxins did not interfere with the specificity. This test possesses the combination of sensitivity and objectivity absent in current methods for assaying enterotoxin and provides a model for investigating other enterotoxin serotypes.

Animals↗

PubMed1979

Effect of purified Escherichia coli heat-stable enterotoxin on intestinal cyclic nucleotide metabolism and fluid secretion.

Enterotoxigenic Escherichia coli cause diarrhea by elaborating two enterotoxins. The large-molecular-weight, heat-labile toxin causes intestinal secretion by stimulating cyclic adenosine 5'-monophosphate production. The mechanism by which the small-molecular-weight, heat-stable enterotoxin induces secretion is unclear. The present study tested the hypothesis that heat-stable enterotoxin induces secretion by altering intestinal cyclic nucleotide concentrations. This was studied in suckling mice by using highly purified E. coli heat-stable enterotoxin obtained from a strain pathogenic for humans. At 3 min after administration of this toxin, intestinal cyclic guanosine 5'-monophosphate (GMP) levels were increased 10-fold. Cyclic GMP levels decreased thereafter, but still were greater than control levels at 120 min. Cyclic adenosine 5'-monophosphate levels fell to one-half of control levels at 3 min and remained below control levels for 120 min. When the time course of enterotoxin-induced secretion was compared with changes in cyclic GMP levels, fluid secretion was not evident until 15 to 30 min after enterotoxin administration. Thus, the increase in intestinal cyclic GMP concentration preceded measurable fluid secretion. And finally, administration of the 8-bromo analog of cyclic GMP evoked fluid secretion, the time course of which was similar to that induced by enterotoxin. These, and other data, strongly suggest that E. coli heat-stable enterotoxin induces intestinal secretion by increasing intestinal cyclic GMP levels.

Animals↗

PubMed1980

Isolation of specific and common antibodies to staphylococcal enterotoxins B, C1, and C2.

Staphylococcal enterotoxins C1 and C2 elicit the production of antibodies in rabbits that give precipitin reactions with both toxins and gel diffusion plates. These enterotoxins also elicit the production of antibodies that are specific for each of the enterotoxins. Enterotoxin B elicits the production of antibodies in some rabbits that react with enterotoxins B, C1, and C2 in gel diffusion plates and in radioimmunoassay. Antibodies specific for enterotoxin B are produced also. Enterotoxin C1 elicits the production of antibodies that cross-react weakly with enterotoxin B, indicating that the antigenic sites involved in the cross-reactions are not identical in the two toxins. The antibodies have been isolated by affinity chromatography.

Animals↗

PubMed1984

Properties of crude Campylobacter jejuni heat-labile enterotoxin.

The amount of crude Campylobacter jejuni enterotoxin present in culture products was quantitated by comparing the response of these preparations with that of pure Escherichia coli heat-labile toxin (LT) in the Chinese hamster ovary assay and in enzyme-linked immunosorbent assays that used GM ganglioside or antisera to LT or both. Maximum C. jejuni enterotoxin production was achieved by growth at 42 degrees C for 24 h under agitation in supplemented GC medium. Adding polymyxin separately to either the broth supernatant or the cells enhanced the recovery of toxin; the yield from cell lysates was much lower. The quantity of C. jejuni enterotoxin produced by clinical isolates obtained locally or provided from Mexico varied widely, over a spectrum from none to large amounts; quantitative values for the amount of C. jejuni enterotoxin determined by the Chinese hamster ovary and enzyme-linked immunosorbent assays correlated with the degree of secretory potency of this material in ligated rat ileal loops. The cytotonic activity of C. jejuni enterotoxin in Chinese hamster ovary cells was abolished by heating at 96 degrees C for 10 min and by preincubation either with GM ganglioside or with LT or cholera toxin antisera. The secretory activity of C. jejuni enterotoxin in ligated rat ileal loops was passively neutralized by antiserum to LT, and immunizing rats with either LT or its B subunit significantly (P less than 0.001) reduced fluid response to active challenge with C. jejuni enterotoxin in ligated ileal loops. These observations indicate that strains of C. jejuni vary in their capacity to elaborate a heat-labile enterotoxin that has close immunological homology with LT and cholera toxin.

Animals↗

PubMed1986

Production and characterization of monoclonal antibodies to Clostridium perfringens enterotoxin.

Four hybridoma cell lines producing monoclonal antibodies to Clostridium perfringens enterotoxin were established by fusion of mouse myeloma and spleen cells obtained from mice immunized with the enterotoxin and its toxoid. An enzyme-linked immunosorbent assay indicated that the two antibodies, 2-B-4 and 3-G-10, bound to those regions that were located close each other; the others, 3-B-2 and 2-H-2, bound to other independent regions on the enterotoxin. Release of 51Cr from Vero cells with the enterotoxin was inhibited by either 2-B-4 or 3-G-10, both of which inhibited the binding of 125I-labeled enterotoxin to the cells. Neither binding nor cytotoxicity of the enterotoxin was affected by 2-H-2; 3-B-2 only barely inhibited the binding but neutralized the enterotoxin shown by 51Cr release. It seems justified to conclude that 3-B-2 blocks the toxic action after the enterotoxin has bound to Vero cells.

Animals↗

PubMed1972

Sporulation and enterotoxin production by mutants of Clostridium perfringens.

The ability of Clostridium perfringens type A to produce an enterotoxin active in human food poisoning has been shown to be directly related to the ability of the organism to sporulate. Enterotoxin was produced only in a sporulation medium and not in a growth medium in which sporulation was repressed. Mutants with an altered ability to sporulate were isolated from an sp(+) ent(+) strain either as spontaneous mutants or after mutagenesis with acridine orange or nitrosoguanidine. All sp(0) (-) mutants were ent(-). Except for one isolate, these mutants were not disturbed in other toxic functions characteristic of the wild type and unrelated to sporulation. A total of four of seven osp(0) mutants retained the ability to produce detectable levels of enterotoxin. None of the ent(-) mutants produced gene products serologically homologous to enterotoxin. A total of three sp(-) mutants, blocked at intermediate stages of sporulation, produced enterotoxin. Of these mutants, one was blocked at stage III, one probably at late stage IV, and one probably at stage V. A total of three sp(+) revertants isolated from an sp(-) ent(-) mutant regained not only the ability to sporulate but also the ability to produce enterotoxin. The enterotoxin appears to be a sporulation-specific gene product; however, the function of the enterotoxin in sporulation is unknown.

Animals↗

PubMed1987

Cloning and expression of the Salmonella enterotoxin gene.

This report examines the genetic basis for Salmonella typhimurium Q1 enterotoxin production. A 918-base-pair XbaI-HincII fragment of plasmid pJM17, composed of cholera toxin (CT) coding sequences (ctxAB), was used as a gene probe. With this probe, the S. typhimurium enterotoxin was identified on a 6.3-kilobase EcoRI-PstI fragment of chromosomal DNA from plasmidless strain Q1. We cloned this 6.3-kilobase fragment into Escherichia coli RR1. The genetic map of the cloned Salmonella enterotoxin (stx) gene was similar but not identical to the CT and E. coli heat-labile enterotoxin genes. By using synthetic oligonucleotides derived from the sequences of CT subunits A (ctxA) and B (ctxB), it was revealed that there were some conserved regions of DNA encoding the enterotoxins of strain Q1 and Vibrio cholerae. Expression of the cloned stx gene in minicells and subsequent Western blot (immunoblot) analysis with CT antitoxin demonstrated that the Salmonella enterotoxin had two or more subunits with molecular sizes of 45, 26, and 12 kilodaltons. Crude cell lysates of E. coli RR1(pCHP4), containing the cloned Salmonella enterotoxin gene, elicited fluid secretion in ligated rabbit intestinal loops and firm induration in rabbit skin. Both of these enterotoxic responses were neutralized by antisera specific for CT. Mucosal tissue from positive intestinal loops contained elevated levels of cyclic AMP. These data suggest some evolutionary relatedness between the enterotoxin genes of S. typhimurium and V. cholerae.

Animals↗