Phenotypic and genotypic characteristics of Staphylococcus aureus isolates from raw bulk-tank milk samples of goats and sheep

Two hundred and ninety-three isolates of Staphylococcus aureus obtained from 127 bulk-tank milk samples of goats and sheep from Switzerland were characterised by pheno- and genotypic traits. Of the 293 S. aureus isolates, 193 (65.9%) were egg yolk-negative and 15 (5.1%) were negative for clumping factor and/or protein A determined by a latex agglutinating test system. For 285 isolates, PCR amplification of the 3' end of the coagulase gene showed a single amplicon. Five differently sized PCR products of 500, 580, 660, 740 and 820 bp were distinguished. In 191 isolates (n = 293) staphylococcal enterotoxin (SE) genes were detected: 123 isolates tested positive for SEC gene, 31 for SEG gene, 28 for SEA gene, 26 for SEJ gene, 24 for SEI gene, 4 for SEB gene and 4 for SED gene. Furthermore, 126 isolates were positive for the gene encoding the toxic shock syndrome toxin 1. Coagulase gene restriction profile analysis of the 145 isolates harbouring SEA or SEC genes revealed six different patterns using AluI and five different patterns using HaeIII. In summary, within these two groups, high genotypic uniformity within the different sized coagulase gene amplicons was demonstrated. This is the first study providing comprehensive characterisation data of S. aureus strains originating from bulk-tank milk samples of goats and sheep. Remarkable differences in phenotypic traits between S. aureus originating from goats and sheep and bovine milk were found. Moreover, the high prevalence of toxin-producing S. aureus may be important as it is relevant to food hygiene.     

Molecular typing of enterotoxigenic Staphylococcus aureus isolated from bovine mastitis in Korea

One hundred and sixty-six Staphylococcus aureus isolates from mastitic milk samples from different cows on 26 farms were investigated for staphylococcal enterotoxins(SEs) and toxic shock syndrome toxin-1(TSST-1) by polymerase chain reaction(PCR) and reverse passive latex agglutination assay(RPLA). SEs and the TSST-1 gene were detected in thirty-seven isolates based on a multiplex PCR; SEA was detected in 32 isolates, SEB in 3 isolates, SEC in 1 isolate, and SEA and the TSST-1 gene in 1 isolate. Of the 37 enterotoxigenic isolates, thirty-three isolates were enterotoxigenic according to RPLA, where 29 isolates produced SEA, 3 isolates produced SEB, and 1 isolate produced SEC. The enterotoxin-producing S. aureus isolates were further characterized by pulsed-field gel electrophoresis(PFGE). A macrorestriction analysis revealed 11 PFGE patterns. Among the 33 enterotoxigenic S. aureus isolates, 45.4% exhibited the same PFGE pattern I. Accordingly, although the enterotoxin-producing S. aureus isolates from bovine mastitis were genetically diverse, 1 common genotype prevailed on the farms, indicating that PFGE pattern I isolates may be the most disseminated in Korea.     

Identification of Staphylococcus aureus strains negative for enterotoxins A, B and C isolated from bovine mastitis in México

A total of 41 isolates of Staphylococcus aureus obtained from bovine mastitis in 7 different states in Mexico were analyzed by the polymerase chain reaction (PCR) to determine the presence of encoding genes for enterotoxins A, B and C. The oligonucleotides were designed by specific regions of the sea, seb, sec genes. Surprisingly, none of the isolates presented the prospective amplification bands when they were run on agarose gels. On the contrary, reference strains CECT 976 SEA; CECT 5191 SEB; and CECT 4465 SEC showed the prospective amplification products. In order to confirm these results, enterotoxin production A, B, C, D, and E was determined by enzyme linked fluorescent assay (ELFA) using a MiniVIDAS system, on 15 Staphylococcus aureus selected at random from among the 41 isolates. None of the analyzed strains was positive to the test, whereas reference strains enterotoxins producing: CECT 976 SEA; CECT 5191 SEB; CECT 4465 SEC, CECT 4466 SED; CECT 5192 SEE produced concentrations of the toxins detected for this technique. The role of enterotoxins in the pathogenicity of S. aureus in bovine mastitis in Mexico is discussed.     

Pattern of enterotoxin genes seg, seh, sei and sej positive Staphylococcus aureus isolated from bovine mastitis

PCR detection of the genes encoding the newly described staphylococcal enterotoxins (SE) SEG, SEH, SEI and SEJ was carried out for 104 randomly selected Staphylococcus aureus field strains isolated from cases of bovine mastitis. Sixty-one (58.7%) isolates were positive for one or more of these novel enterotoxin genes. Thirty-six field strains were classified as carrier of seg, 22 of sei gene and 23 were positive for sej gene. None of the 104 investigated ruminant S. aureus strains carried the seh gene. Thirty-seven of these S. aureus strains showed a combination of genes encoding enterotoxin types SEA to SEE or toxic shock syndrome toxin 1 (TSST 1). Thirteen cultures harboured only one, 28 two, 12 three and 8 four enterotoxin genes. Among the 61 S. aureus field strains 14 (23.0%) were positive for the genes encoding SEJ and SED and 10 (16.4%) isolates for those encoding SEG and SEI. Isolates harbouring the sed/sej genes were further characterized by macrorestriction analysis and pulsed-field-gelelectrophoresis (Pfge). Macrorestriction analysis revealed six patterns. Nine of these14 S. aureus isolates (64.3%) exhibited two patterns with a high degree of relationship (>80%).     

Prevalence and characterization of Staphylococcus aureus and enterotoxigenic Staphylococcus aureus in retail raw chicken meat throughout Japan

A total of 444 samples of raw chicken meat (thighs, breasts, wings, livers, gizzards, hearts and ovaries) that retailed at 145 different supermarkets in 47 prefectures in Japan were examined for contamination with Staphylococcus aureus in association with its enterotoxigenicity. S. aureus was isolated from 292 (65.8%) of the samples, and from 131 of the 145 supermarkets. There was no significant difference in the detection rate of S. aureus according to the type of meat examined. About 80% of 714 isolates belonged to the poultry (57.1%) and human biotypes (22.1%). Seventy-eight (21.7%) of 360 isolates were enterotoxigenic and isolated from 78 samples in 53 supermarkets in 31 prefectures. Staphylococcal enterotoxins (SEs) produced were SEB (50 isolates), SEA (14), SEC (8), SED (2), SEA+SEB (2), and SEA+SEC (2). Most of the enterotoxigenic isolates belonged to the human and poultry biotypes, coagulase type VII, VIII or IV, and were lysed by phages of group III. Identical SE types, biotypes, coagulase types and pulsed-field gel electrophoresis (PFGE) patterns were shown in isolates from different types of meat at the same supermarket and from samples taken from different supermarkets in the same prefectures or in isolates from samples obtained from several different prefectures. Among the 50 SEB-producing isolates, 27 yielded three similar PFGE patterns that differed by only a few fragments, suggesting that they were closely related genetically. The three patterns were found in isolates of samples that retailed at 17 supermarkets in 11 prefectures, indicating that they may be disseminated among raw chicken meat in Japan.     

Detection of classical enterotoxins and identification of enterotoxin genes in Staphylococcus aureus from milk and dairy products

Milk and dairy products are frequently contaminated with enterotoxigenic Staphylococcus aureus, which is often involved in staphylococcal food poisoning. The distribution of genes encoding staphylococcal enterotoxins (SE) in S. aureus isolated from bovine, goat, sheep and buffalo milk and dairy products was verified by the presence of the corresponding SE production. A total of 112 strains of S. aureus were tested for SE production by immuno-enzymatic (SEA-SEE) and reversed passive latex agglutination (SEA-SED) methods, while multiplex-PCR was applied for SE genes (sea, sec, sed, seg, seh, sei, sej and sel). Of the total strains studied, 67% were detected to have some SE genes (se), but only 52% produced a detectable amount of the classic antigenic SE types. The bovine isolates frequently had enterotoxin SEA, SED and sej, while SEC and sel predominated in the goat and sheep strains. The results demonstrated (i) marked enterotoxigenic S. aureus strain variations, in accordance with strain origin and (ii) the two methods resulted in different information but concurred on the risk of foodstuff infection by S. aureus.     

PCR detection of staphylococcal enterotoxin genes in Staphylococcus aureus strains isolated from raw and pasteurized milk

Milk is considered a nutritious food because it contains several important nutrients including proteins and vitamins. Conversely, it can be a vehicle for several pathogenic bacteria such as Staphylococcus aureus. This study aimed to analyze the frequency of genes encoding the staphylococcal enterotoxins (SEs) SEA, SEB, SEC, SED, SEE, SEG, SEH, SEI and SEJ in S. aureus strains isolated from raw or pasteurized bovine milk. S. aureus was found in 38 (70.4%) out of 54 raw milk samples at concentrations of up to 8.9 x 10(5) CFU/ml. This microorganism was present in eight samples of pasteurized milk before the expiration date and in 11 samples analyzed on the expiration date. Of the 57 strains studied, 68.4% were positive for one or more genes encoding the enterotoxins, and 12 different genotypes were identified. The gene coding for enterotoxin A, sea, was the most frequent (16 strains, 41%), followed by sec (8 strains, 20.5%), sed (5 strains, 12.8%), seb (3 strains, 7.7%) and see (2 strains, 5.1%). Among the genes encoding the other enterotoxins, seg was the most frequently observed (11 strains, 28.2%), followed by sei (10 strains) and seh and sej (3 strains each). With the recent identification of new SEs, the perceived frequency of enterotoxigenic strains has increased, suggesting that the pathogenic potential of staphylococci may be higher than previously thought; however, further studies are required to assess the expression of these new SEs by S. aureus, and their impact in foodborne disease. The quality of Brazilian milk is still low, and efforts from the government and the entire productive chain are required to attain consumer safety.     

Occurrence of enterotoxigenic strains of Staphylococcus aureus in raw poultry meat

The aim of this work was to determine the contamination of raw poultry meat with enterotoxigenic strains of S. aureus, using the PCR method. PCR is a rapid and sensitive method, which can show the presence in food of enterotoxigenic strains of S. aureus on the basis of specific gene sequences and detect the potential source of contamination before enterotoxins are produced. No coagulase-positive staphylococci strains were found in 65 samples of chicken parts, but these bacteria were present in 11 of 23 examined samples of minced turkey meat (48%). Using the primers for enterotoxin genes A to C, 4 of the 11 isolated S. aureus strains showed a positive result in the PCR. Three of the isolates represented the SEB gene and remaining one the SEC gene. The results obtained showed that PCR is sensitive and rapid method which may be used for detection and identification of enterotoxigenic S. aureus.     

Frequency of superantigen-producing Staphylococcus intermedius isolates from canine pyoderma and proliferation-inducing potential of superantigens in dogs

This preliminary study investigated the potential role of staphylococcal superantigens in the pathogenesis of canine pyoderma. The staphylococcal enterotoxins A (SEA), SEB, SEC and SED, and the toxic shock syndrome toxin-1 (TSST-1) were assayed in isolates from skins of dogs with pyoderma. Culture supernatants from 25 of 96 isolates were positive for multiple superantigens, with SEA and SEC being the most frequently detected. In in vitro stimulation of canine peripheral blood mononuclear cells and quantitative flow cytometry revealed that low concentrations of SEA and SEB were potent stimulators of blastogenesis of T cells.     

Multiplex PCR and RPLA Identification of Staphylococcus aureus enterotoxigenic strains from bulk tank milk

Enterotoxigenic Staphylococcus aureus in raw milk poses a potential health hazard to consumers, and the identification of such strains should be used as part of a risk analysis of milk and milk products. The primary purpose of this study was to investigate the occurrence of enterotoxigenic S. aureus strains in raw milk supplied for dairy processing in the Czech Republic. A further aim was to compare the production of staphylococcal enterotoxins (SEs) with the presence of the corresponding genes. This was undertaken using multiplex polymerase chain reaction (PCR) and reversed passive latex agglutination (RPLA). Out of 440 bulk tank milk samples from 298 dairy herds, 70 proved positive for S. aureus (15.9%). Staphylococcal enterotoxin genes (ses) were detected in 39 (55.7%) isolates. The genes most commonly detected were sei (38.6%), seg (31.4%) and sea (27.1%). Genes seb, seh, sed, sej and sec were observed in 10%, 4.3%, 2.9%, 2.9% and 1.4% of strains respectively. Genes see and sel did not occur. The most frequently detected genotypes were seg, sei at 11.4%; sea at 10.0%; and sea, seg, sei at 8.6%. Toxin production was observed in nine (12.9%) S. aureus isolates. Seven strains were detected as SEB- (10%) and two as SED- (2.9%) producing. A relatively high number (32%) of discrepancies between the results with multiplex PCR and RPLA assays was obtained, particularly on account of SEA. Nineteen strains were sea positive by PCR but SEA negative by RPLA, and one strain was sec positive and SEC negative. The results of both methods were identical concerning SEB and SED. It was concluded that detection of ses by PCR was a useful additional tool to support identification of enterotoxigenic strains.     

Quantitative analysis of Staphylococcus aureus in skimmed milk powder by real-time PCR

A large-scale outbreak of food poisoning caused by consumption of skimmed milk powder contaminated with staphylococcal enterotoxin A (SEA) occurred in Japan. No viable Staphylococcus aureus was detected in the skimmed milk powder, however, sea and nuc genes of S. aureus were detected in it by PCR. The number of S. aureus in skimmed milk powder was estimated by quantitative real-time PCR.     

Isolation of enterotoxigenic strains of staphylococci from dogs

The ability of 309 staphylococcal isolates from household dogs to produce enterotoxin, coagulase, thermonuclease and hemolysin was investigated. A total of 52 (16.8%) isolates from 45 out of 150 dogs examined were enterotoxigenic when tested for enterotoxin types A, B and C. Based on sites sampled, 33 (20.5%) out of 161 isolates from the anterior nares were enterotoxigenic while from dorsal skins 19 (12.8%) out of 148 isolates were enterotoxigenic. Staphylococcal enterotoxin C(SEC) was predominantly produced as 21 (6.8%) isolates elaborated it and also accounted for 40.4% of all enterotoxins produced by isolates. Staphylococcal enterotoxins A(SEA) and B(SEB) were produced by 10 (3.2%) and 16 (5.2%) strains, respectively. Mixed enterotoxin types AB, AC and BC were produced by 1,3 and 1 strains, respectively. With human plasma, 17.1% of coagulase-positive and 15.0% of coagulase-negative strains were enterotoxigenic. However, using canine plasma, 19.1% and 6.9% of the coagulase-positive and negative isolates, respectively, were enterotoxigenic. The incidence of enterotoxigenicity was 16.9% amongst thermonuclease-positive isolates and 16.3% for thermonuclease-negative strains.Alpha hemolysin was predominantly produced by 180 (60.2%) isolates and 19.9% of these were enterotoxigenic. Beta hemolysin was produced by 36 (11.7%) isolates with 13.9% enterotoxigenic, while 87 (28.2%) exhibited gamma hemolytic pattern amongst which 11.5% were enterotoxigenic.Based on data provided on coagulation of human and canine plasmas and hemolytic patterns, it is concluded that a large proportion of canine isolates from this environment are not of canine biotypes, but are most probably human biotypes.     

Characterization of leukocytotoxic and superantigen-like factors produced by Staphylococcus aureus isolates from milk of cows with mastitis.

Staphylococcus aureus is a major pathogen for cattle, causing various forms of subclinical and clinical mastitis. Two groups of virulence factors (leukotoxins and superantigens) are supposed to play an important role in the initiation and/or the exacerbation of this disease. In order to detect all known and putative members of leukotoxins and SAgs (superantigens), we tested secreted factors of different S. aureus isolates in flow cytometry-based assays.Isolates were sampled from 68 cows of different farms and cultured for 24h in vitro. Supernatants were then coincubated with purified polymorphonuclear granulocytes (PMN) or combinations of blood mononuclear cells (MNC) and PMN. Viable PMN and MNC were determined by quantitative flow cytometry. In addition, we recorded the proliferation-inducing potential of isolate supernatants for bovine MNC. Based on these criteria, the supernatants of S. aureus isolates fell in three groups. The first group (n=32), termed LT-SNs (leukotoxin-containing supernatants), killed purified granulocytes (neutrophils and eosinophils) in vitro. The second group of supernatants (n=20), termed SAg-SN (superantigen-containing supernatants), induced activation and proliferation of mononuclear cells (MNC) and, only in the presence of MNC, resulted in a selective depletion of neutrophils after 24h in vitro. The third group of supernatants (n=16) contained neither LTs or SAgs. Functionally, SAg-SNs behaved like purified staphylococcal enterotoxin A (SEA) or SEB tested in parallel. The absence of SAg-like activity in LT-SNs was confirmed by heat treatment of LT-SNs, which destroyed the leukocytotoxic activity, but did not reveal any MNC-activating potential. This study, therefore, suggests, that pathogenic S. aureus isolates either produce leukotoxins or superantigens and that both groups of virulence factors can easily be differentiated by the functional assays described.The prevalence of leukotoxin- or superantigen-producing isolates was comparable among cattle with subclinical (LT=41%; SAg=30.8%) mastitis. The higher frequency of LT-producing isolates in cases of clinical mastitis (LT=55.2%; SAg=27.6%) was not significant. At least, these findings argue against the dominant role of superantigens or leukotoxins in S. aureus-induced bovine mastitis.     

荧光定量PCR检测牛乳中金黄色葡萄球菌肠毒素A基因方法的建立

为建立检测牛乳中金黄色葡萄球菌(S.aureus)肠毒素A基因(SEA)定性定量的检测方法,本研究针对S.aureus SEA基因片段设计1对引物,将构建的重组质粒作为阳性对照,建立了S.aureus SEA DNA的SYBR Green I real-time PCR检测方法.结果显示,特异性产物Tm值为78.2℃～78.5℃,最低可检测到49.5 fg/μL(16.5拷贝)的阳性质粒.标准曲线的相关系数为0.99.与其他常见的产SEB的S.aureus、产SEC的S.aureus、无乳链球菌、大肠杆菌、嗜热链球菌、伤寒沙门氏菌、大肠杆菌DH5 α及JM109均无交叉反应.该检测方法具有较好的特异性和敏感性,为牛乳中S.aureus的快速检测提供了新的技术手段.     

Antimicrobial susceptibility and presence of resistance genes in staphylococci from poultry

The species distribution, susceptibility to 19 antimicrobial agents and presence of selected genes encoding resistance to macrolides, streptogramins and tetracyclines were examined among 118 staphylococcal isolates from infections of poultry in Denmark. Isolates were identified using a combination of conventional biochemical testing and 16S rDNA sequencing. The most common species were Staphylococcus aureus (83), Staphylococcus hyicus (11), Staphylococcus xylosus (9) and Staphylococcus cohnii (6). The isolates were susceptible to most antimicrobials tested. A high frequency of S. aureus (30%) was resistant to ciprofloxacin. Only six (7%) S. aureus isolates and one Staphylococcus saprophyticus were penicillin resistant. Resistance to sulphamethoxazole was observed among 16 (19%) of S. aureus isolates and two coagulase negative staphylococci (CNS). Twenty (24%) of the S. aureus isolates were resistant to erythromycin and 19 of these isolates contained the ermA gene, whereas the remaining isolate contained the ermC gene. Eleven (48%) of the novobiocin resistant CNS were resistant to erythromycin and all these isolates contained the ermA gene. Two isolates identified as S. xylosus, were found to be resistant to streptogramins and both contained the vatB- and the vgaB-genes. Thirty-nine (47%) of the S. aureus isolates, three of nine S. hyicus and eight of the 23 novobiocin resistant CNS were tetracycline resistant and all contained the tet(K) gene. A single S. aureus isolate also contained the tet(M) gene. The present study showed a frequent occurrence of resistance to fluoroquinolones, tetracycline and macrolides among staphylococci isolated from broilers in Denmark, whereas the occurrence of resistance to other antimicrobial agents remains low. Similar genes, encoding resistance to erythromycin, tetracycline and streptogramins to those previously observed, were detected.     

牛乳腺炎金黄色葡萄球菌肠毒素A基因的克隆及序列分析

根据GenBank上公布的金黄色葡萄球菌肠毒素A（staphylococcus enterotoxin A,SEA）基因的全序列,设计一对特异性引物扩增内蒙古分离株的SEA基因序列。经基因克隆和序列测定,表明扩增的基因片段长度为582 bp,与标准菌株ATCC13565的SEA基因片段序列相似性为100%,与GenBank上公布的金黄色葡萄球菌菌株（EF520720.1）SEA基因相似性达到99.14%。本研究结果为进一步研究建立牛乳中SEA分子检测技术奠定了实验基础。     

Characterization of enterotoxigenic Staphylococcus aureus strains isolated from bovine mastitis in north-east Switzerland

Thirty-four strains of enterotoxin-producing Staphylococcus aureus obtained from milk samples of 34 dairy cows suffering from mastitis from 34 different farms in north-east Switzerland were identified and further characterized by pheno- and genotypic methods. This included the identification of staphylococcal enterotoxin (SE) types, an antibiotic resistance testing, the appraisal of hemolysis, the egg yolk reaction, the detection of the clumping factor and protein A by means of a latex agglutination, the PCR amplification of a S. aureus specific part of the gene encoding the 16S-23S rRNA "intergenic spacer" region and a species specific part of the 23S rRNA-gene, the PCR amplification of the clumping factor (clfA) gene, the X region and the IgG-binding region of the protein A (spa) gene, the coagulase (coa) gene and additionally a macrorestriction analysis of the chromosomal DNA. Within the 26 cultures which formed a single SE, there were 23 SEC- and three SED-formers. Eight cultures were SEAD formers. It was remarkable that 22 SEC formers were also positive for TSST-1. Eighteen of the 23 SEC-formers could be classified as being of the same phenotype. Most of the cultures of one enterotoxin type also showed a great uniformity in the size and number of repeats of the X region as well as in the size of the IgG-binding region of protein A gene and in the size of the coagulase gene. Macrorestriction analysis revealed 11 PFGE patterns. These were in part only different from each other in a few fragments and thus displayed close clonal relations. The results of the present investigation show that a broad distribution of identical or closely related enterotoxin-producing S. aureus clones seem to contribute to the bovine mastitis problem in north-east Switzerland.     

Determination of staphylococcal exotoxins, SCCmec types, and genetic relatedness of Staphylococcus intermedius group isolates from veterinary staff, companion animals, and hospital environments in Korea

The Staphylococcus (S.) intermedius group (SIG) has been a main research subject in recent years. S. pseudintermedius causes pyoderma and otitis in companion animals as well as foodborne diseases. To prevent SIG-associated infection and disease outbreaks, identification of both staphylococcal exotoxins and staphylococcal cassette chromosome mec (SCCmec) types among SIG isolates may be helpful. In this study, it was found that a single isolate (one out of 178 SIG isolates examined) harbored the canine enterotoxin SEC gene. However, the S. intermedius exfoliative toxin gene was found in 166 SIG isolates although the S. aureus-derived exfoliative toxin genes, such as eta, etb and etd, were not detected. SCCmec typing resulted in classifying one isolate as SCCmec type IV, 41 isolates as type V (including three S. intermedius isolates), and 10 isolates as non-classifiable. Genetic relatedness of all S. pseudintermedius isolates recovered from veterinary staff, companion animals, and hospital environments was determined by pulsed-field gel electrophoresis. Strains having the same band patterns were detected in S. pseudintermedius isolates collected at 13 and 18 months, suggesting possible colonization and/or expansion of a specific S. pseudintermedius strain in a veterinary hospital.     

Differences between Danish bovine and human Staphylococcus aureus isolates in possession of superantigens

PCR-assays for the detection of staphylococcal enterotoxins A-E, and H, toxic shock toxin 1, and exfoliative toxins A and B were evaluated against phenotypic methods, and performed well. Four hundred and fourteen isolates of Staphylococcus aureus from Danish cases of bovine mastitis were screened for genes encoding these superantigens. One hundred isolates from Danish human carriers were also included in the study. In contrast to the frequent presence of genes encoding and in vitro expression of superantigens among the human carrier isolates, only one of 414 isolates from bovine mastitis carried the genes encoding enterotoxin C and toxic shock toxin-1. These results further support the hypothesis that the bovine and human S. aureus reservoirs constitute two separate sub-populations of the species S. aureus. The results also show that these superantigens are generally not present in Danish S. aureus isolates from bovine mastitis, and thus play no essential role in the pathogenesis of bovine S. aureus mastitis.     

Detection of staphylococcal enterotoxins in milk and milk products

For food evaluation the determination of the number of Staphylococcus aureus (hereinafter S. aureus) colonies is insufficient in view of present scientific knowledge. The results, advantages and shortcomings of diagnostic methods are demonstrated on an example of three methods of detection of staphylococcal enterotoxins in milk and milk products. 133 strains were investigated by the method of biotyping of S. aureus strains. Four strains of S. aureus were included in biotype A, seven xin-producing strains were isolated seventeen times by detection of 96 S. aureus strains were not included in any biotype, the other strains belonged to biotypes C and E. This method can be used as an auxiliary method of evaluation of foods containing S. aureus bacteria. The agar-gel precipitation method of enterotoxin detection in isolated strains of S. aureus has just restricted validity. The enteroto-strains. The main shortcoming of this method is a fact that the result concerning the isolated strains need not be identical with the result of enterotoxin detection in food. Direct assays of staphylococcal enterotoxins in milk and milk products using an enzymoimmunological method seem to be the most promising, mainly due to their high sensitivity (0.0001-0.001 micrograms.ml1-) and other advantages. Positive and negative results are presented on an example of two model trials with winter sheep milk cheese.