Enterotoxin production by Staphylococcus aureus isolated from mastitic cows

Staphylococcus aureus is an important cause of mastitis in cows. The ability of S. aureus strains to produce one or more enterotoxins in milk and dairy products is linked to staphylococcal food poisoning. To determine whether staphylococci causing bovine mastitis could cause human foodborne intoxication, the production of staphylococcal enterotoxins A through D (SEA, SEB, SEC, and SED) by 160 S. aureus isolates was evaluated with the use of a reverse passive latex agglutination enterotoxin kit. All S. aureus strains were isolated over a 9-month period from 2,343 routine submissions of a composite quarter collection of individual mastitic cows at 18 dairy farms in the San Joaquin Valley in California. Prior to enterotoxin detection, isolates were grown by a method that enhances the in vitro synthesis of enterotoxin. Twenty-two of 160 S. aureus isolates produced enterotoxin. Seven produced SEC, 12 produced SED, and 3 produced both SEC and SED. None of the isolates produced SEA or SEB.     

Use of novel PCR primers specific to the genes of staphylococcal enterotoxin G, H, I for the survey of Staphylococcus aureus strains isolated from food-poisoning cases and food samples in Taiwan

Data regarding the incidence of the newly found enterotoxigenic Staphylococcus aureus strains in food poisoning cases and in food samples were to date not available in Taiwan. In this study, PCR primers specific for the detection of SEG, H and I genes, i.e., seg, seh and sei, were used for the assay of 55 human isolates of S. aureus negative to the classical enterotoxins (SEA-->SEE) detection. These isolates were from the fecal specimens of the patients suffering from food poisoning outbreaks. Only eight strains were found to have the seg, seh and sei. The presence of other bacterial pathogens, such as Vibrio parahaemolyticus, Bacillus cereus and Salmonella spp. and perhaps, strains producing other new staphylococcal enterotoxins, in the fecal specimens of these patients, may account for these food poisoning cases. For 139 strains from food samples, such as frozen Chinese foods, Chinese sausages and lunch meals, sea strains accounted for the major portion and it seemed to be the most common SE type to coexist with seg, seh and sei. Only two strains had sec and none of them had seg, seh or sei. For strains without the classical SE genes, only 13 strains had seg, seh and/or sei. The above results imply that seg, seh and sei S. aureus strains play only a minor role in food-borne outbreaks in Taiwan.     

Development of two multiplex polymerase chain reactions for the detection of enterotoxigenic strains of Staphylococcus aureus isolated from foods

Two multiplex polymerase chain reactions were developed for the detection of enterotoxigenic strains of Staphylococcus aureus: one multiplex reaction for the simultaneous detection of enterotoxigenic strains type A (entA), type B (entB), and type E (entE) and another for the simultaneous detection of enterotoxigenic strains type C (entC) and type D (entD). Both reactions were standardized with the use of the reference enterotoxigenic strains of S. aureus: FRI 722, producer of staphylococcal enterotoxin (SE) type A (SEA); FRI 1007, producer of SEB; FRI 137, producer of SEC1; FRI 472, producer of SED; and FRI 326, producer of SEE. Optimized methods were used to determine the presence of enterotoxigenic types for 51 S. aureus strains isolated from meat (sausage, ham, and chorizo) and dairy (powdered milk and cheese) products by the Baird-Parker technique. The enterotoxigenic capacities of the strains were determined by the indirect enzyme-linked immunosorbent assay (ELISA) with the use of reference staphylococcal toxins and antitoxins. Fifty of the 51 strains isolated were enterotoxigenic and produced one to four enterotoxin types, with the most frequently produced types being SEA and SED. Levels of correlation between the presence of genes that code for the production of SE (as determined by polymerase chain reaction) and the expression of these genes (as determined by the indirect ELISA) were 100% for SEA and SEE, 86% for SEC, 89% for SED, and 47% for SEB.     

Real-time PCR detection of Staphylococcus aureus in milk and meat using new primers designed from the heat shock protein gene htrA sequence

Staphylococcus aureus may cause foodborne disease outbreaks and staphylococcal infections and is one of the major causes of mastitis. Rapid and reliable methods for detection of this microorganism in milk and other foods are needed. In this study, we designed a primer set from the sequence of the heat shock protein gene htrA, a gene coding for high-temperature-requirement A (HtrA) protein, and used it for real-time PCR detection of S. aureus isolates: 16 reference strains and 40 strains isolated from food-poisoning cases. All strains tested generated positive results. Bacterial strains other than S. aureus, including strains of other Staphylococcus species, did not produce positive results. When this primer set was used for the real-time PCR detection of S. aureus in milk and meat samples without the preenrichment step, samples with target cell numbers greater than 10(3) CFU/ml or CFU/g could be detected, indicating the potential quantitative ability of this real-time PCR assay. With a 10-h preenrichment step, however, a detection limit of 1 CFU/ml or CFU/g could be obtained.     

Coagulase-positive Staphylococci and Staphylococcus aureus in food products marketed in Italy

Staphylococcus aureus is a very common organism capable of producing several enterotoxins (SEs) that cause intoxication symptoms of varying intensity in humans when ingested through contaminated food. This paper reports the results of an investigation on the presence of Coagulase-Positive Staphylococci (CPS) and S. aureus in several food products marketed in Italy and on food contact surface swabs sampled from the food industry. A total of 11,384 samples were examined and 1971 of them (17.3%) were found to contain CPS. The assays performed on 541 CPS strains led to the identification of 537 S. aureus strains on which characterization of type A, B, C and D staphylococcal enterotoxins (SEA, SEB, SEC and SED) was performed. A total of 298 S. aureus strains (55.5%) produced one or more SEs: 33.9% of the strains produced SEC, 26.5% SEA, 20.5% SEA+SED, 13.4% SED, 2.7% SEB, 1.7% SEA+SEB, 0.7% SEC+SED and 0.3% produced SEA+SEC and SEB+SEC. The investigation highlighted that these organisms are very common and constitute a potential risk for consumers' health.     

Occurrence of toxigenic Staphylococcus aureus in ready-to-eat food in Korea

Toxigenic Staphylococcus aureus contamination in ready-to-eat (RTE) food is a leading cause of foodborne illness in Korea. To monitor food contamination by S. aureus, a total of 3332 RTE food samples were selected from nationwide wholesale marts between 2003 and 2004 and examined. A total of 285 (8.6%) of the overall samples were contaminated by S. aureus. According to the analysis, 31.6% of the tested cream-cakes, 19.8% of the raw fish, and 19.3% of the rice cakes with filling were contaminated with S. aureus. Forty-seven percent of the strains isolated from the contaminated food were enterotoxigenic S. aureus. The phenotypic result of the strain isolated from food showed that 48% of the strains produced one or more toxins, such as staphylococcal enterotoxins A, B, and C (SEA, SEB, and SEC). At least one SEA was produced by over 90% of the toxigenic strains. Other toxins, such as SEB, SEC, SED, SEA+SEC, and SEC+SED, were each detected. Toxic shock syndrome toxin 1 (TSST-1), a causative agent of toxic shock syndrome, was detected in 13 strains of the toxigenic isolates from the food. As the result of genotyping, 22 strains with a toxin gene that was not detected in the phenotypic analysis were also detected. Sixty-nine percent of the toxigenic strains had at least one sea gene, and the most prevalent genotype was sea+seh (34.4%), followed by sea (18.8%) and sea+seg+sei (15.6%). The tst gene encoding TSST-1 was found in 13 strains (13.5%). The genes (eta and etb) encoding exfoliative toxins A and B were not detected in any of the samples.     

PCR detection of Staphylococcal enterotoxins (SEs) N, O, P, Q, R, U, and survey of SE types in Staphylococcus aureus isolates from food-poisoning cases in Taiwan

Staphylococcal enterotoxins (SEs) are superantigenic toxins. They are five major classical types, i.e., SEA, SEB, SEC, SED, SEE, and new SEs or SE-like superantigens, such as SEG to SEU. Only the staphylococcal superantigens (SAgs) that induce emesis following oral administration in a monkey model are designated as SEs while other related toxins are called SE-like (SEl) superantigens. To survey the enterotoxin genotypes for S. aureus strains isolated from food-poisoning cases in Taiwan, we developed PCR primers specific for SEN, SEO, SEP, SEQ, SER, and SEU genes. The complete SE sequences and their expression potential for strains positive to sen, seo, sep, seq, ser, and seu specific primers were also determined. These strains were used as reference strains. With the PCR primers specific for all SEs or SAgs, including toxic shock syndrome toxin I (TSST-1), we assayed the genotypes of 147 S. aureus strains isolated from patients associated with staphylococcal food-poisoning outbreaks occurred during 2001-2003. For these 147 strains, 135 (91.8%) were found positive for one or more SE or SAg genes. For classical enterotoxin and TSST-1 types, the major one was tsst-1 (59.1%) following by sea (29.2%), seb (19.7%), sec (6.8%), and sed (2.0%). For new SE and SAg types, the major one was sei (29.9%) and sep (27.9%) followed by, sek (16.3%), seo (14.3%), seu (14.2%), sem (11.6%), sen (10.9%), seq (10.9%), seh (8.2%), sel (6.8%), and ser (5.4%) etc. This report reveals the whole SE and SAg genotypes for S. aureus strains isolated from staphylococcal food-poisoning cases in Taiwan.     

Application of extended single-reaction multiplex polymerase chain reaction for toxin typing of Staphylococcus aureus isolates in South Korea

The extended single-reaction multiplex PCR (esr-mPCR) developed in this study to detect staphylococcal enterotoxins (SEs), including SEA, SEB, SEC, SED, SEE, SEH, SEI, and SEJ, requires fewer sets of primers than other conventional multiplex PCRs and can be used to detect newly identified staphylococcal enterotoxins SEs more readily. Esr-mPCR analysis of 141 isolates of Staphylococcus aureus obtained from abattoir and livestock product samples revealed that 27 of the S. aureus isolates were toxigenic, and two were 2 multitoxigenic isolates. The most prevalent SE type was SEI followed by SEA and SEH. In addition, we investigated the clonal relatedness of toxigenic S. aureus isolates by arbitrarily primed PCR (AP-PCR). AP-PCR analysis of toxigenic S. aureus isolates revealed that the discriminatory power of AP-PCR was 9 (D=0.81), 8 (D=0.77), and 10 types (D=0.83) with primers AP1, ERIC2, and AP7, respectively. The combination of three each AP-PCR result could rearrange toxigenic S. aureus isolates into 10 types and five subtypes, with the D-value of 0.92. Interestingly, our data showed that toxigenic S. aureus isolates from different sources had different fingerprinting patterns although some of them carried the same types of SE genes. These data suggest that combinations of esr-mPCR and AP-PCR can provide a powerful approach for epidemiological investigation of toxigenic S. aureus isolates.     

Occurrence, characterization and antimicrobial resistance of enterotoxigenic Staphylococcus aureus isolated from meat and dairy products

Staphylococcus aureus is considered to be one of the leading causes of food-borne illnesses. Milk, dairy products and meats are often contaminated with enterotoxigenic strains of this bacterium. Foodstuff contamination may occur directly from infected food-producing animals or may result from poor hygiene during production processes, or the retail and storage of foods, since humans may carry the microorganism. The number of S. aureus strains that exhibits antimicrobial-resistance properties has increased, together with the potential risk of transmitting the same properties to the human microflora via foods or inducing infections hard to be treated. This paper reports the results of a 3-year survey (2003-2005) on the occurrence of S. aureus in meat and dairy products. Of 1634 samples examined, 209 (12.8%) were contaminated with S. aureus. A total of 125 enterotoxigenic S. aureus strains were biotyped and their antimicrobial resistance pattern tested. Most of the isolated strains produced SED (33.6%), followed by SEA (18.4%), SEC (15.2%), SEB (6.4%) and belonged mainly to the Human ecovar (50.4%), followed by Ovine (23.2%), Non-Host-Specific (17.6%), Bovine (7.2%) and Poultry-like (1.6%) ecovars. Finally, the 68.8% analysed strains showed antimicrobial resistance properties at least at one of antibiotics tested. Human biotype showed antimicrobial resistance at more than one antibiotic than the other biotypes (p<0.05). The results provided evidence that the presence of enterotoxigenic and antimicrobial resistant strains of S. aureus has become remarkably widespread in foods. This calls for better control of sources of food contamination and of the spread of antimicrobial-resistance organisms.     

Enterotoxin production and DNA fingerprinting in Staphylococcus aureus isolated from human and food samples. Relations between genetic types and enterotoxins

A total of 224 Staphylococcus aureus strains from human carriers (110 strains) and manually handled foods (114 strains) collected in the Principality of Asturias, Spain over 1995-1999 were analysed for the production of enterotoxins (SEs) A, B, C, and D by a reversed passive latex agglutination test and by amplification of ent genes (A, B, C, D, E, and J) using PCR. Sixty-two strains were enterotoxigenic and a good relation between detection of SEs and their ent genes was found. No strain carried entE and all strains producing SED carried entD and entJ genes. Among the enterotoxigenic strains the percentages registered were 29, 8, 35, 18, 2, 2, and 6 for SEA, SEB, SEC, SEDJ, SEAC, SEADJ and SECDJ, respectively. DNA fingerprinting of 77 strains (the SE prototypes, 62 enterotoxigenic and 10 non-enterotoxigenic [NE]) was carried out by randomly amplified polymorphic DNA using two selected primers independently. Combining results from both primers, 10 genetic types were defined, which showed a different degree of relationship (similarity coefficient: 0.9-0.36) and were clustered into three lineages. One lineage clustered five genetic types and a wide diversity of strains, mainly SEA, SEB, SEDJ, and NE. Another lineage clustered only SEC, SECDJ and NE strains. These two lineages showed a low genetic relationship and appeared as endemic in healthy humans living in the Principality of Asturias. The third lineage included only the prototype strains for SEA and SEE.     

The detection of enterotoxins and toxic shock syndrome toxin genes in Staphylococcus aureus by polymerase chain reaction

A simple polymerase chain reaction (PCR)-based procedure was developed for the detection of fragments of staphylococcal enterotoxins (SEs) SEA, SEB, SEC, SED, SEE, SEG, SEH, and SEI together with the toxic shock syndrome toxin (TSST-1) genes of Staphylococcus aureus. One hundred and twenty-nine cultures of S. aureus were selected, 39 of which were recovered from 38 suspected staphylococcal food-poisoning incidents. The method was reproducible, and 32 different toxin genotypes were recognized. The presence of SE genes was associated with S. aureus strains reacting with phages in group III, and the TSST-1 gene with phages in group I. There was a 96% agreement between the PCR results for detection of SEA-D and TSST-1 as compared with a commercial reverse passive latex agglutination assay for the detection of SEs from cultures grown in vitro. Enterotoxin gene fragments were detected in S. aureus cultures recovered from 32 of the 38 suspected staphylococcal food poisoning incidents, and of these, 17 were associated with SEE, SEG, SEH, and SEI in the absence of SEA-D. Simple PCR procedures were also developed for the detection of SE directly in spiked food samples, and this was most successfully achieved in mushroom soup and ham. Detection was less successful in three types of cheese and in cream. SEA or SEB were detected by enzyme-linked immunosorbent assay in three food samples (two of which were associated with food poisoning incidents) naturally heavily contaminated with S. aureus: the appropriate SEA or SEB gene fragments were detected directly in these three foods by PCR.     

The use of DNA probes for confirming enterotoxin production by Staphylococcus aureus and micrococci

DNA-DNA colony hybridization was employed to evaluate the results obtained by different immunological methods for detection of staphylococcal enterotoxin. Staphylococcus aureus strains tested for staphylococcal enterotoxin production by immuno-assays and micrococci not previously tested for staphylococcal enterotoxin production were examined for presence of the genes encoding for staphylococcal enterotoxin A, B, C and E by using three corresponding DNA probes. The staphylococcal enterotoxin A probe also detected staphylococcal enterotoxin E gene because of 100% homology. The optimal sensitivity plate method showed the best accordance between the immuno-assay and the hybridization reactions. The enzyme-linked immunosorbent assay detected 12.5 to 17% staphylococcal enterotoxin producers without hybridization reactions. The microslide gel double diffusion test and the reversed passive latex agglutination test showed rather poor accordance with the hybridization reactions. All 17 strains of different micrococci investigated were negative in hybridization with all three DNA probes.     

Prevalence of Staphylococcus aureus and staphylococcal enterotoxins in raw pork and uncooked smoked ham--a comparison of classical culturing detection and RFLP-PCR.

In many countries Staphylococcus aureus is considered to be the second or third most common pathogen causing outbreaks of food poisoning, only outnumbered by Salmonella spp. and in competition with Clostridium perfringens. Often the consumption of ham or meat containing staphylococcal enterotoxins (SE) is identified as cause of the illness. Thus, to gain an insight into the prevalence of S. aureus and its emetic enterotoxins in raw pork and uncooked smoked ham and to investigate how the prevalence of the pathogen is influenced during the fabrication process, a total of 135 samples of raw pork, salted meat and ready-for-sale uncooked smoked ham were examined for the prevalence of S. aureus and staphylococcal enterotoxins A to D (SEA-SED). To this means classical cultural methods were employed as well as molecular biological techniques (PCR) and the results were compared. In 25.9% of all samples S. aureus was detected by culture whereas 51.1% of the samples showed a positive result when PCR was used for the detection of the pathogen. Fresh meat was contaminated most often. By PCR, 62.2% were identified as being S. aureus positive compared to 57.7% positive samples using the cultural technique. The detection rate during the fabrication process declined significantly. The pathogen was cultivated from 8.9% of the salted meat samples. Here, 55.6% of the samples reacted positively in the PCR, and finally, in approximately a third of the ready-for-sale smoked hams, S. aureus genes were found. From 11.1% of these samples, the pathogen could be isolated by culture. From these results, we conclude that the PCR used in this study is more sensitive than the classical cultural method. By PCR, one or more staphylococcal enterotoxin genes were found in 24 of the 135 examined samples. This means that 34.8% of the staphylococcal strains identified using the PCR technique were enterotoxigenic. Using the SET-RPLA, a percentage of 28.6% enterotoxigenic isolates was ascertained. No staphylococcal enterotoxin formation was detected by the SET-RPLA in ready-for-sale ham, although SE-genes were found by PCR. The detection of SE-genes by PCR is faster and easier to perform than the SET-RPLA.     

Discrimination of Staphylococcus aureus strains from different species of Staphylococcus using Fourier transform infrared (FTIR) spectroscopy

Staphylococcus aureus is a widespread opportunistic pathogen that can cause food-borne illness and is sometimes associated with raw milk and raw milk cheese products. The traditional taxonomic procedures for classification of staphylococcal species are time consuming and often several tests are required. FTIR spectroscopy offers a rapid method for the discrimination and identification of S. aureus strains isolated from raw milk and raw milk cheeses. FTIR spectroscopy was used to discriminate S. aureus from other species of Staphylococcus. This was achieved by using a model composed of 39 species and subspecies of Staphylococcus. The model was validated using a set of spectra of strains isolated from raw milk and different varieties of French raw milk cheese. S. aureus was successfully discriminated from the other species of Staphylococcus and all the strains of S. aureus isolated from raw milk and different varieties of French raw milk cheese were also successfully identified as such. These results demonstrated that FTIR spectroscopy is a rapid (results obtained within 24 h starting from a pure strain or a single colony) and robust method for the identification of S. aureus isolates of dairy origin and food-borne origin in general.     

Characterization of Staphylococcus aureus strains associated with food poisoning outbreaks in France

Enterotoxins produced by Staphylococcus aureus are responsible for staphylococcal food-poisoning outbreaks (SFPO). In France, SFPO are the second cause of food-borne diseases after Salmonella. However, very little is known about the strains involved. The objective of this study was to characterize the staphylococcal strains related to these SFPO through phenotypic and genotypic analyses. A total of 178 coagulase-positive staphylococcal isolates recovered from 31 SFPO (1981-2002) were screened through biotyping. Thirty-three strains representative of the different biotypes in each SFPO were further examined for SmaI macrorestriction-type, phage-type, resistance to various antimicrobial drugs, presence of staphylococcal enterotoxin (se) genes sea to sei, and production of enterotoxins SEA to SED. All these 33 strains were identified as S. aureus species: 27 were of human biotypes and six ovine or non-host-specific biotypes. Most (74.1%) strains reacted with group III phages. Eleven strains were resistant to at least two classes of antibiotics and among them, two were resistant to methicillin. Twenty-nine strains carried one or several of the eight se genes tested; the gene sea was most common (n=23), and often linked to sed (n=12) or seh (n=5). The novel se genes seg-i were in all cases associated with se genes sea to sed except for one strain which carried only seg and sei. Pulsed-Field Gel Electrophoresis (PFGE) of SmaI macrorestriction digests of the 33 strains discriminated 32 PFGE patterns grouped into nine biotype-specific clusters. All five strains carrying sea and seh were grouped together into the same sub-cluster. Three of the four se-gene-negative strains were in one PFGE cluster: all four should be tested for se genes not included in this study and, if negative, be further investigated for the presence of unidentified SEs.     

Phage inactivation of Staphylococcus aureus in fresh and hard-type cheeses

Bacteriophages are regarded as natural antibacterial agents in food since they are able to specifically infect and lyse food-borne pathogenic bacteria without disturbing the indigenous microbiota. Two Staphylococcus aureus obligately lytic bacteriophages (vB_SauS-phi-IPLA35 and vB_SauS-phi-SauS-IPLA88), previously isolated from the dairy environment, were evaluated for their potential as biocontrol agents against this pathogenic microorganism in both fresh and hard-type cheeses. Pasteurized milk was contaminated with S. aureus Sa9 (about 10(6)CFU/mL) and a cocktail of the two lytic phages (about 10(6)PFU/mL) was also added. For control purposes, cheeses were manufactured without addition of phages. In both types of cheeses, the presence of phages resulted in a notorious decrease of S. aureus viable counts during curdling. In test fresh cheeses, a reduction of 3.83log CFU/g of S. aureus occurred in 3h compared with control cheese, and viable counts were under the detection limits after 6h. The staphylococcal strain was undetected in both test and control cheeses at the end of the curdling process (24h) and, of note, no re-growth occurred during cold storage. In hard cheeses, the presence of phages resulted in a continuous reduction of staphylococcal counts. In curd, viable counts of S. aureus were reduced by 4.64log CFU/g compared with the control cheeses. At the end of ripening, 1.24log CFU/g of the staphylococcal strain was still detected in test cheeses whereas 6.73log CFU/g was present in control cheeses. Starter strains were not affected by the presence of phages in the cheese making processes and cheeses maintained their expected physico-chemical properties.     

酸奶制作中金黄色葡萄球菌污染的模拟实验研究

金黄色葡萄球菌是乳制品中检出率较高的食源性致病菌.该论文采用模拟实验的方式,探讨了在金黄色葡萄球菌污染的情况下,酸奶制作过程中乳酸菌和金黄色葡萄球菌消长的动态规律,并初步估计了被污染酸奶中金黄色葡萄球菌耐热肠毒素的危害水平.研究结果显示,当发酵温度偏低、奶粉添加量偏高和金葡菌污染程度较高时,金黄色葡萄球菌生长速率和最大菌数明显提高.参考有关文献进行估算,对于一个体重为61kg的成人来说,如果酸奶食用量少于1300mL,一般不会有酸奶金黄色葡萄球菌耐热肠毒素中毒的危险;但对于儿童来说则有一定风险.本研究对于酸奶制作工艺控制及酸奶的安全消费具有参考意义.     

金黄色葡萄球菌基因组岛对多重耐药表型与生物被膜能力的影响机制

本文选择常见的典型食源性微生物金黄色葡萄球菌,从耐药性微生物感染防控角度出发,对广州地区临床分离的127株葡萄球菌的耐药表型、基因组岛分型与生物被膜生长能力进行研究。通过微量肉汤稀释法检测确定菌株对26种抗菌药物的药敏结果;PCR扩增葡萄球菌属特异性基因16S r RNA、金黄色葡萄球菌菌株特异性基因fem A、耐药基因mec A以检测确定菌株耐药特性;多重PCR检测金葡菌基因组岛SCCmec基因元件中ccr复合物,mec复合物以对其进行分型。107株耐药型金葡菌均为多重耐药,且呈耐9种或以上抗生素占76.1%(86/113)。113株葡萄球菌SCCmec分型结果为:I型0株,II型12株,III型73株,IV型10株,V型11株,5株为无法分型;本文对基因组岛和金黄色葡萄球菌耐药表型与生物被膜能力的相关性进行分析与探讨,为进一步对各种食源性微生物引起的食品污染进行安全控制,提供了研究基础。     

Characterization of Staphylococcus aureus isolates from white-brined Urfa cheese

The aim of this study was to investigate the presence of Staphylococcus aureus and staphylococcal enterotoxin (SE) genes in Urfa cheese samples and to characterize the enterotoxigenic potential of these isolates. From a total of 127 Urfa cheese samples, 53 isolates (from 41.7% of the samples) were identified by a species-specific PCR assay as S. aureus. Of these isolates, 40 (75.5%) gave positive PCR results for the 3' end of the coa gene. The coa-positive S. aureus strains were characterized for their population levels and enterotoxigenic properties, including slime factor, β-lactamase, antibiotic susceptibilities, production of the classical SEs (SEA through SEE), in both cheese and liquid cultures by enzyme-linked immunosorbent assay (ELISA) and for the presence of specific genes, including classical SE genes (sea through see), mecA, femA, and spa, by PCR. The genetic relatedness among the coa-positive S. aureus isolates was investigated by PCR-based restriction fragment length polymorphism (RFLP) analysis and the 23S rRNA gene spacer. The 23S rRNA gene spacer and coa RFLP analysis using AluI and Hin6I revealed 14 different patterns. SEB, SEC, and SEA and SEE were detected by ELISA in three cheese samples. Fourteen S. aureus strains harbored enterotoxin genes sea through see, and three strains carried multiple toxin genes. The most commonly detected toxin gene was sec (25% of tested strains). Of the 40 analyzed S. aureus strains, 3 (7.5%) were mecA positive. Based on tandem repeats, four coa and spa types were identified. The results of this study indicate that S. aureus and SEs are present at significant levels in Urfa cheese. These toxins can cause staphylococcal food poisoning, creating a serious hazard for public health.