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ST7 Staphylococcus aureus is highly prevalent in humans, pigs, as well as food in China; however, staphylococcal food poisoning (SFP) caused by this ST type has rarely been reported. On May 13, 2017, an SFP outbreak caused by ST7 S. aureus strains occurred in two campuses of a kindergarten in Hainan Province, China. We investigated the genomic characteristics and phylogenetic analysis of ST7 SFP strains combined with the 91 ST7 food-borne strains from 12 provinces in China by performing whole-genome sequencing (WGS). There was clear phylogenetic clustering of seven SFP isolates. Six antibiotic genes including blaZ, ANT (4′)-Ib, tetK, lnuA, norA, and lmrS were present in all SFP strains and also showed a higher prevalence rate in 91 food-borne strains. A multiple resistance plasmid pDC53285 was present in SFP strain DC53285. Among 27 enterotoxin genes, only sea and selx were found in all SFP strains. A ФSa3int prophage containing type A immune evasion cluster (sea, scn, sak, and chp) was identified in SFP strain. In conclusion, we concluded that this SFP event was caused by the contamination of cakes with ST7 S. aureus. This study indicated the potential risk of new emergencing ST7 clone for SFP. [ABSTRACT FROM AUTHOR]

Gastric cancer is one of the most common malignancies worldwide and it is the fourth leading cause of cancer-related death. Staphylococcus aureus enterotoxin A (SEA) is one of the toxins affecting the expression of apoptotic-related genes. The aim of this research is to investigate the effects of S. aureus enterotoxin A on the expression of BAK, FAS, BAX, TNF-α, BCL-2 and Survivin in AGS cell lines. In this study, AGS cells were transfected with the pcDNA3.1(+)-enterotoxin A or empty pcDNA3.1(+) plasmids using Lipofectamine 2000. The cells were grown in Dulbecco's modified Eagle's medium (DMEM) with 10% FBS. Then, the mRNA expression level of BAK, FAS, BAX, TNF-α, BCL-2 and Survivin was measured by qRT-PCR method. Our findings indicated that Staphylococcus aureus enterotoxin A markedly altered the expression level of the apoptotic-related gene of the AGS cell line. The mRNA expression of BAK, FAS, BAX and TNF-α genes increased statistically significant in enterotoxin A treated AGS cells compared with the control group. On the contrary, BCL2 and Survivin mRNA expression showed a significant decrease compared with AGS cells were transformed by empty pcDNA3.1(+) plasmid. We suggested that SEA could represent an agent that can significantly changes apoptosis-related genes expression in AGS cell line, leading to reduction of tumor cell proliferation.

Staphylococcus aureus (S. aureus) is a Gram-positive bacteria considered one of the leading causes of community and hospital-acquired illnesses or public health concerns. Antibiotic resistance in this microorganism is one of the greatest issues in global health care. The use of metal nanoparticles and their oxides is one of the potential approaches to combating bacteria resistance to antibiotics. The antibacterial properties of ZnO NPs against enterotoxigenic S. aureus were studied. ZnO NPs were tested in vitro by agar diffusion test. They resulted in 26 and 22 mm zones of inhibition for a size of 20 nm and a concentration of 20 mM against 105 and 107 CFU/mL S. aureus, respectively. The MIC of ZnO NPs of various sizes, 20 and 50 nm, with 105 CFU/mL was 2.5 and 5 mM, respectively. MIC with 107 CFU/mL was five mM for 20 and 50 nm ZnO NPs. Further, the highest growth reduction percentage, 98.99% in the counts of S. aureus was achieved by ZnO NPs of size 20 nm and concentration of 10 mM. Moreover, the obtained ELISA results indicated a significantly decreased concentration of enterotoxin A with all concentrations and sizes of ZnO NPs. PCR analysis showed a significant effect on sea gene in response to ZnO NPs treatments leading to loss of the gene, unlike the unaffected nuc gene. Moreover, morphological changes and cell shape distortion were detected by scanning electron microscope for bacterial cells treated with ZnO NPs. [ABSTRACT FROM AUTHOR]

Milk, a nutritious global important food commodity, serves as an excellent medium for microbial growth as well. The foodborne pathogen Staphylococcus aureus is a commensal member of human microflora that enters the food chain through poor hygienic practices and cross contamination and causes various clinical manifestations in humans. During this study, raw milk and swab samples (milker's hand, udder, towel, milking bucket, and farm floor) were collected from four middle-scale buffalo dairy farms. The results revealed S. aureus presence in 11.6% ( n = 56/448) bucket raw milk samples and 2.6% ( n = 12/448) udder raw milk samples. Contrarily, S. aureus prevalence was significantly higher in farm floors (100%, n = 84/84), towel (35.7%, n = 10/28), milking bucket (11.6%, n = 56/448), milker's hand (10.7%, n = 3/28), and udder swab samples (4.0%, n = 18/448). The chi-square test yielded p values of 0.000, 0.005, and 0.0011 for udder raw milk, udder swab, and milker's hand swab, respectively. The p values of the milking bucket ( p = 0.048) and farm floors ( p = 0.0183) confirmed their possible role in S. aureus cross contamination. Gene amplifications of nuclease and enterotoxin A indicate potential virulence of S. aureus isolates in collected samples. Antibiotic susceptibility testing revealed multidrug resistance in 44% ( n = 239) of S. aureus isolates with the highest resistance of 61% against penicillin. Resistance against ampicillin, streptomycin, and lincomycin was observed. Fewer S. aureus isolates were resistant to kanamycin and erythromycin, whereas the lowest number of resistant isolates was observed against chloramphenicol. A high prevalence of S. aureus in the farm environment and milking equipment suggested the cross contamination of potentially enterotoxin-producing and multidrug-resistant S. aureus to raw milk. Therefore, good hygiene practices should be enforced to avoid foodborne and zoonotic infections.

Staphylococcus aureus (S. aureus) is one of the most important microorganisms causing foodborne outbreaks worldwide. the present work was planned to study the occurrence, count, and identification of S. aureus and investigation of the obtained isolates for the presence of enterotoxin A encoding gene (sea) in milk and some milk products in New Valley governorate, Egypt. A total of 150 random samples of marketable milk and some milk products including soft cheese (Kareish, Domiati) & ice cream (30 for each) were collected from different localities in the New Valley governorate. S. aureus occurred in 11.6, 10, 3.3 and10% for marketable milk, Kariesh cheese, Domiati cheese, and ice cream, respectively. The presence of the enterotoxin sea gene was identified in 4 (57%) and 2 (50%) in milk and soft cheese, respectively, while none of the isolates obtained from ice cream samples harbored this gene. The investigation revealed that the examined milk samples and its products were contaminated with S. aureus with average counts of 2.8×108, 7.5×105, 7.6×106, and 1.7×107 for milk, Kariesh cheese, Domiati cheese, and ice cream samples, respectively. In conclusion, the presence of S. aureus in milk and some milk products indicated that the examined samples were of low quality which reflects unhygienic measures during milking, processing, storage, and distribution chain. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a serious pathogen unleashing its virulence through several classes of exotoxins such as hemolysins and enterotoxins. In this study, we designed a novel multi-antigen subunit vaccine which can induce innate, humoral and cellular immune responses. Alpha hemolysin, enterotoxins A and B were selected as protective antigens for combining into a triple antigen chimeric protein (HAB). Immunoinformatics analysis predicted HAB protein as a suitable vaccine candidate for inducing both humoral and cellular immune responses. Tertiary structure of the HAB protein was predicted and validated through computational approaches. Docking studies were performed between the HAB protein and mice TLR2 receptor. Furthermore, we constructed and generated recombinant HAB (r-HAB) protein in E. coli and studied its toxicity, immunogenicity and protective efficacy in a mouse model. Triple antigen chimeric protein (r-HAB) was found to be highly immunogenic in mouse as the anti-r-HAB hyperimmune serum was strongly reactive to all three native exotoxins on Western blot. In vitro toxin neutralization assay using anti-r-HAB antibodies demonstrated > 75% neutralization of toxins on RAW 264.7 cell line. Active immunization with r-HAB toxoid gave ~ 83% protection against 2 × lethal dosage of secreted exotoxins. The protection was mediated by induction of strong antibody responses that neutralized the toxins. Passive immunization with anti-r-HAB antibodies gave ~ 50% protection from lethal challenge. In conclusion, in vitro and in vivo testing of r-HAB found the molecule to be nontoxic, highly immunogenic and induced excellent protection towards native toxins in actively immunized and partial protection to passively immunized mice groups. Key points: • HAB protein was computationally designed to induce humoral and cellular responses. • r-HAB protein was found to be nontoxic, immunogenic and protective in mouse model. • r-HAB conferred protection against lethal challenge in active and passive immunization. [ABSTRACT FROM AUTHOR]

Foods with high protein content such as milk, meat and salads have been frequently incriminated in staphylococcal enterotoxins (SЕ) outbreaks. Staphylococcal enterotoxin A (SEA) is most toxic from all 20 SЕ. Potency of SEA requires that the methods of analysis in food must be sensitive enough to measure very low concentrations. A fluorescent immunoassay, sandwich format, for SEA detection in milk was developed, using immobilized F(ab′)2 fragment and anti-SEA monoclonal antibody (mAb) bonded onto the surface of magnetic nanoparticles (MNPs). F(ab′)2 fragment of anti-SEA mAb was obtained by pepsin hydrolysis of mAb. The fragment and antibody were conjugated with fluorescein 5(6)-isothiocyanate (FITC) and formed F(ab')2 fragment-FITC and mAb-FITC conjugates. The obtained conjugates were purified by Sephadex 25 size exclusion chromatography. A fluorescent immunoassay was developed by using immobilized F(ab')2 fragment and anti-SEA mAb and were obtained conjugates F(ab')2 fragment-FITC and mAb-FITC. The analytical parameters of the immunoassay with F(ab′)2 fragment were compared with those obtained from the immunoassay with mAb. When using a immobilized F(ab')2 and fluorescent conjugate F(ab')2 fragment - FITC, the linear range of SEA in buffer was from 0.7 to 300 pg/mL, and in milk from 1.0 to 300 pg/mL. With an immobilized mAb and mAb-FITC conjugate, the linear range of SEA in buffer and in milk was from 50 to 300 pg/mL. It was found that the immunoassay with F(ab')2 fragment allow the measurement of SEA at very low concentrations. The limit detection of SEA in milk samples with F(ab')2 fragment was 0.9 pg/mL and with whole antibody 30 pg/mL. [ABSTRACT FROM AUTHOR]

Staphylococcal enterotoxin A (SEA) is an exotoxin that releases from gram-positive bacteria, and it is a source of food poisoning. Here, the luminescence method is introduced to quantify and detect SEA using carbon quantum dots (CQDs) as fluorophore; SEA antibody loaded Fe3O4 nanoparticles (Fe3O4@Ab) are prepared for SEA extraction. SEA in standards and milk samples were conjugated with Fe3O4@Ab (Fe3O4@Ab@SEA), separated by a 1.4 T magnet, and dispersed in phosphate buffer saline. The fluorescence of CQDs is quenched when mixed with Fe3O4@Ab@SEA, without elution of SEAs from the Fe3O4@Ab@SEA. It is a fast and reliable method compared with the immunoassay counterpart. At optimum quantification conditions, a linear relationship between SEAs concentration and CQDs fluorescent quenching was observed from 4 to 2000 pg mL−1 with the detection limit 2 pg mL−1. Repeatability and reproducibility were calculated as a percentage of relative standard deviations and were from 1.56 to 11.76%. Finally, the proposed method was applied to measure the SEA contents of four sterilized and homogenized milk samples without fat precipitations and have shown significant reliability. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus (S. aureus) is a Gram-positive bacteria considered one of the leading causes of community and hospital-acquired illnesses or public health concerns. Antibiotic resistance in this microorganism is one of the greatest issues in global health care. The use of metal nanoparticles and their oxides is one of the potential approaches to combating bacteria resistance to antibiotics. The antibacterial properties of ZnO NPs against enterotoxigenic S. aureus were studied. ZnO NPs were tested in vitro by agar diffusion test. They resulted in 26 and 22 mm zones of inhibition for a size of 20 nm and a concentration of 20 mM against 105 and 107 CFU/mL S. aureus, respectively. The MIC of ZnO NPs of various sizes, 20 and 50 nm, with 105 CFU/mL was 2.5 and 5 mM, respectively. MIC with 107 CFU/mL was five mM for 20 and 50 nm ZnO NPs. Further, the highest growth reduction percentage, 98.99% in the counts of S. aureus was achieved by ZnO NPs of size 20 nm and concentration of 10 mM. Moreover, the obtained ELISA results indicated a significantly decreased concentration of enterotoxin A with all concentrations and sizes of ZnO NPs. PCR analysis showed a significant effect on sea gene in response to ZnO NPs treatments leading to loss of the gene, unlike the unaffected nuc gene. Moreover, morphological changes and cell shape distortion were detected by scanning electron microscope for bacterial cells treated with ZnO NPs.

Gastric cancer is one of the most common malignancies worldwide and it is the fourth leading cause of cancer-related death. Staphylococcus aureus enterotoxin A (SEA) is one of the toxins affecting the expression of apoptotic-related genes. The aim of this research is to investigate the effects of S. aureus enterotoxin A on the expression of BAK, FAS, BAX, TNF-a, BCL-2 and Survivin in AGS cell lines. In this study, AGS cells were transfected with the pcDNA3.1(+)-enterotoxin A or empty pcDNA3.1(+) plasmids using Lipofectamine 2000. The cells were grown in Dulbecco's modified Eagle's medium (DMEM) with 10% FBS. Then, the mRNA expression level of BAK, FAS, BAX, TNF-a, BCL-2, and Survivin was measured by the qRT-PCR method. Our findings indicated that Staphylococcus aureus enterotoxin A markedly altered the expression level of the apoptotic-related gene of the AGS cell line. The mRNA expression of BAK, FAS, BAX, and TNF-a genes increased statistically significantly in enterotoxin A treated AGS cells compared with the control group. On the contrary, BCL2 and Survivin mRNA expression showed a significant decrease compared with AGS cells were transformed by empty pcDNA3.1(+) plasmid. We suggested that SEA could represent an agent that can significantly change apoptosis-related genes expression in the AGS cell line, leading to reduction of tumor cell proliferation. [ABSTRACT FROM AUTHOR]

Ochratoxin A (OTA) and staphylococcus enterotoxin A (SEA) are highly toxic contaminants and have induced human health problems. They commonly occur in milk and milk products. A competitive fluorescent immunoassay was developed for rapid and simultaneous determination of these toxins in milk samples. The procedure was based on the competitive immunoreactions between antigens in sample and antigen-fluorescent dye conjugates with immobilised antibodies on magnetic nanoparticles (MNPs). Each monoclonal antibody specifically recognises its corresponding toxin (antigen), and there is no cross-reactivity in the assay. First, monoclonal antibodies against OTA and SEA were produced. The activity of the obtained antibodies was determined by fluorescent-linked immunosorbent assay. Then, the monoclonal antibodies were immobilised on MNPs. The amounts of immobilised anti-OTA antibody and anti-SEA antibody were determined to be 20 and 22 μg mL−1, respectively. The antigen-fluorescent dye conjugates OTA–OVA–ATTO620 and SEA-FITC were prepared. The optimal amount of immobilised antibodies for competitive immunoassay was determined. It was found that the linear range of OTA in buffer was larger (0.001–100 ng mL−1) than the linear range of SEA (0.001–20 ng mL−1). The results for simultaneous determination of OTA and SEA in sixfold diluted milk were almost the same in buffer; the linear range for OTA was from 0.005 to 100 ng mL−1 and for SEA from 0.005 to 20 ng mL−1. The detection limit for both OTA and SEA in milk was 0.004 ng mL−1. The developed method took half the time of the individual assays (20 min). The assay was evaluated using spiked milk samples. The influences of somatic cell count, fat, pH and protein concentration in milk on immunoassay were studied. In summary, this developed immunoassay could provide an effective and rapid approach for detecting multi-toxins in milk samples. [ABSTRACT FROM AUTHOR]

The fish hydrolyzed protein is the potential source of bioactive peptides. These peptides are inactive in the original structure of the protein molecule, but they can be activated after enzymatic hydrolysis. In this research, the effect of the half maximal inhibitory concentration (IC50) rainbow trout hydrolyzed proteins (< 3 kDa) was evaluated on enterotoxin A gene expression of Staphylococcus aureus ATCC 29,213 (the strain of enterotoxin A producer). After affecting IC50 of the fish hydrolyzed protein (< 3 kDa) and S. aureus 1.5 × 108 cfu/ml in BHI broth medium and incubating for the durations 8 and 12 h after exposure (logarithmic phase and stationary phase of S. aureus growth, respectively), the gene expression of enterotoxin A was investigated via Real-Time PCR. gyrA used as a housekeeping endogenous control. Notably only < 3 kDa fish hydrolyzed protein by Flavourzyme enzyme (HF3) showed an inhibitory effect on gene expression of S. aureus enterotoxin A in logarithmic phase. Therefore, fish hydrolyzed proteins can provide a new strategy as natural factors in preventing toxin production, due to the high quality and safety of sensitive foods. [ABSTRACT FROM AUTHOR]

Background and purpose: Staphylococcus aureus (S. aureus) produces various toxins. One of the most important toxins is enterotoxin. Enterotoxin A and B play a major role in food poisoning. This research was conducted to determine the enterotoxin A and B genes in S. aureus strains. Materials and methods: In this study, 223 specimens were collected from the skin and nose of patients, medical personnel and kitchen staff in Imam Khomeini and Bouali Sina hospitals, Sari, Iran. Culture of samples and biochemical tests were used to detect S. aureus. Polymerase Chain Reaction (PCR) was used to detect enterotoxin A and B genes. Results: Out of 223 specimens, 49 (21.97%) were positive for S. aureus, from which 17 (34.7%) isolates were positive for enterotoxin A gene, while none contained enterotoxin B gene. Conclusion: In this study, isolates of S. aureus were positive for presence of enterotoxin A gene. This bacterium has a major role in causing food poisoning, therefore, its prevalence in hospital strains can lead to secondary infections in patients and should be regarded as a serious threat to community health.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogen of nosocomial infection, which is resistant to most antibiotics. Presently, anti-virulence therapy and anti-biofilm therapy are considered to be promising alternatives. In the current work, we investigated the influence of bisdemethoxycurcumin (BDMC) on the virulence-related exoproteins and the biofilm formation using a reference strain and clinic isolated strains. Western blotting, quantitative RT-PCR, and tumor necrosis factor (TNF) release assay were performed to assess the efficacy of BDMC in reducing the expression of Staphylococcus enterotoxin-related exoproteins (enterotoxin A, enterotoxin B) and α-toxin in MRSA. The anti-biofilm activity of BDMC was evaluated through a biofilm inhibition assay. The study suggests that sub-inhibitory concentrations of BDMC significantly inhibited the expression of sea, seb, and hla at the mRNA level in MRSA. Moreover, the expression of virulence-related exoproteins was significantly decreased by down-regulating accessory gene regulator agr, and the inhibition of biofilms formation was demonstrated by BDMC at sub-inhibitory concentrations. Consequently, the study suggests that BDMC may be a potential natural antibacterial agent to release the pressure brought by antibiotic resistance.

Staphylococcus aureus is a potential pathogen contaminating raw milk and dairy products, where it is able to produce thermostable enterotoxins that can cause staphylococcal food poisoning. This study was undertaken to investigate the inhibitory activity of a Lactococcus lactis strain (isolated from milk) on S. aureus growth and staphylococcal enterotoxin A (SEA) production. In the presence of L. lactis, the number of the pathogen decreased significantly (p

The present study investigates the nature of the link between the staphylococcal enterotoxin A (SEA) gene and the lifecycle of Siphoviridae bacteriophages, including the origin of strain variation regarding SEA production after prophage induction. Five strains representing three different genetic lines of the sea region were studied under optimal and prophage-induced growth conditions and the Siphoviridae lifecycle was followed through the phage replicative form copies and transcripts of the lysogenic repressor, cro. The role of SOS response on prophage induction was addressed through recA transcription in a recA-disruption mutant. Prophage induction was found to increase the abundance of the phage replicative form, the sea gene copies and transcripts and enhance SEA production. Sequence analysis of the sea regions revealed that observed strain variances were related to strain capacity for prophage induction, rather than sequence differences in the sea region. The impact of SOS response activation on the phage lifecycle was demonstrated by the absence of phage replicative form copies in the recA-disruption mutant after prophage induction. From this study it emerges that all aspects of SEA-producing strain, the Siphoviridae phage and the food environment must be considered when evaluating SEA-related hazards.

A simple, cheap, and less aggressive immobilization procedure for biomolecules using reduced graphene oxide (rGO) was employed to prepare an impedimetric immunosensor for detection of staphylococcal enterotoxin A (SEA) from Staphylococcus aureus in milk samples. The scanning electron microscopy, cyclic voltammetry, and electrochemical impedance spectroscopy (EIS) were used to monitor the single steps of the electrode assembly process. The glassy carbon (GC)/rGO platform detected the antigen-antibody binding procedures of SEA with concentrations of 0.5 to 3.5 mg L−1 via impedance changes in a low frequency range. The impedimetric immunosensor was successfully applied for the determination of SEA in milk samples.

Background and purpose: Staphylococcus aureus (S. aureus) produces various toxins. One of the most important toxins is enterotoxin. Enterotoxin A and B play a major role in food poisoning. This research was conducted to determine the enterotoxin A and B genes in S. aureus strains. Materials and methods: In this study, 223 specimens were collected from the skin and nose of patients, medical personnel and kitchen staff in Imam Khomeini and Bouali Sina hospitals, Sari, Iran. Culture of samples and biochemical tests were used to detect S. aureus. Polymerase Chain Reaction (PCR) was used to detect enterotoxin A and B genes. Results: Out of 223 specimens, 49 (21.97%) were positive for S. aureus, from which 17 (34.7%) isolates were positive for enterotoxin A gene, while none contained enterotoxin B gene. Conclusion: In this study, isolates of S. aureus were positive for presence of enterotoxin A gene. This bacterium has a major role in causing food poisoning, therefore, its prevalence in hospital strains can lead to secondary infections in patients and should be regarded as a serious threat to community health. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is closely associated with food poisoning incidences, more, because of intoxication due to contamination of food with its enterotoxins. Staphylococcus enterotoxin A (SEA) is highly resistant to heat and other food processing conditions and has been implicated in food borne diseases from dairy products. Here we describe a rapid, gold nanoparticle based lateral flow immunoassay (LFIA) for detecting SEA in milk. Assay is in sandwich format and uses two specific monoclonal antibodies against SEA, one of which is labeled with gold nanoparticles and other is immobilized at control line. We also demonstrate the effect of using a combination of additives in capture antibody in order to enhance assay sensitivity. The developed LFIA is specific and has a lower detection limit of 0.5 μg/ml. LFIA is tested both on spiked buffer and milk samples and display good reproducibility. The assembled strips stable for a month at 4 °C. The method may be suitable for testing SEA in milk samples as it is quick, easy to perform and reproducible. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a potential pathogen contaminating raw milk and dairy products, where it is able to produce thermostable enterotoxins that can cause staphylococcal food poisoning. This study was undertaken to investigate the inhibitory activity of a Lactococcus lactis strain (isolated from milk) on S. aureus growth and staphylococcal enterotoxin A (SEA) production. In the presence of L. lactis, the number of the pathogen decreased significantly (p<0.05) after 6 h of incubation in a laboratory medium and milk (3 log CFU/mL reduction compared to pure cultures). SEA concentration was reduced by 79 % in the co-cultures. S. aureus was unable to reach population levels permitting SEA production in the cheese inoculated with L. lactis during 32 days of storage. In contrast, during the same period, it attained 7 log CFU/g in the cheese manufactured without the lactococcal strain, a level which permitted SEA detection in the cheese extracts. However, this enterotoxin was never detected in the cheese harbouring L. lactis. These results demonstrate the anti-staphylococcal enterotoxinogenesis potential of the L. lactis strain and its usefulness in raw milk cheese biopreservation. [ABSTRACT FROM AUTHOR]

Background: Infections due to Staphylococcus aureus, a nosocomial and community-acquired pathogen, is a major public health problem. Wide range of diseases caused by S. aureus from mild infections of the skin and soft tissue to life threatening diseases which is due to having several virulence factors such as enzymes, toxins and also enterotoxins. Enterotoxin A (SEA) and enterotoxin B (SEB) are superantigens and gasterointestinal toxins causing food poisoning. The sea and seb genes encode SEA and SEB, respectively. The goal of this study was determine the prevalence of sea and seb genes in S. aureus isolated from patients and healthy carriers in Gorgan city, north of Iran. Materials and Methods: 170 isolates of S. aureus (95 from patients and 75 healthy carriers) were collected during 1 year. After identification and purification, DNA extraction was done by phenol - chloroform method. Amplification of sea and seb genes was done by specific primers and polymerase chain reaction method. Results: Among the 170 isolates of S. aureus, 60.6% and 27.1% contained sea and seb genes, respectively. The frequencies of isolates containing sea and seb genes were 58.8% and 61.3%, respectively, in methicillin-resistant Staphylococcus aureus (MRSA) isolates and 23.5% and 28.6%, respectively, in methicillin-sensitive Staphylococcus aureus (MSSA) isolates which were not statistically significant. The frequency of these genes was not related to age, sex and source of isolation in the patients. Conclusion: This study showed that a high proportion of S. aureus isolates carried sea gene, whereas the frequency of seb gene in this region was predictable.

We investigated the cytokine profile of peripheral mononuclear cells from chronic osteomyelitis (OST) patients following in vitro stimulation with staphylococcal enterotoxin A (SEA). We demonstrate that stimulation with SEA induced prominent lymphocyte proliferation and high levels of tumour necrosis factor (TNF)-α, interleukin (IL)-4 and IL-10 secretion in both OST and non-infected individuals (NI). Even though stimulation with SEA had no impact on IL-6 production in either patient group, the baseline level of IL-6 production by cells from OST patients was always significantly less than that produced by cells from NI. After classifying the osteomyelitic episodes based on the time after the last reactivation event as "early" (1-4 months) or "late" osteomyelitis (5-12 months), we found that increased levels of TNF-α and IL-4 in combination with decreased levels of IL-6 were observed in the early episodes. By contrast, increased levels of IL-10, IL-2 and IL-6 were hallmarks of late episodes. Our data demonstrate that early osteomyelitic episodes are accompanied by an increased frequency of "high producers" of TNF-α and IL-4, whereas late events are characterised by increased frequencies of "high producers" of IL-10, IL-6 and IL-2. These findings demonstrate the distinct cytokine profiles in chronic osteomyelitis, with a distinct regulation of IL-6 production during early and late episodes.

"nBackground: Staphylococcus aureus is a major foodborne pathogen throughout the world. Enterotoxins and toxic shock syndrome toxin-1 are important virulence factors and as pyrogenic toxin superantigens have profound effects on the ir host. Thus circulation of TSST1 producing S.aureus among people and food chain is a worrying issue. The present paper was conducted to study Prevalence of tst, entC, entA and entA/C genes in staphylococcus aureus strains isolated from different foods. "n"nMethods: Over 1040 food samples have been analyzed differentially according to Iran national standard (number= 1194) for S.aureus identification. After DNA extraction, PCR reactions were carried out by reference strain as positive control, adequate primers. "n"nResults: At present study, prevalence of foodstuffs contaminated by S.aureus isolates was about 9.5% (100 strains). Of 25% of isolates producing entC, 28% (seven strains) had tst gene at the same time and of 8% of isolates producing entA, 12.5% (one strain) were positive for tst genes simultaneously. Altogether of 9% isolates producing combination of entC and entA, 44.4% (four strains) were also producer of tst gene. "n"nConclusion: Prevalence of TSST1 producing strains in combination with enterotoxin genes is considerable especially with entC and A plus C. On the other hand, circulation of these isolates in humans, animals, foods and environment has hazardous effect for general public health.

Staphylococcus aureus is a major human pathogen producing different types of toxins. Enterotoxin A (SEA) is the most common type among clinical and food-related strains. The aim of the present study was to examine the prevalence of sea in clinical isolates of S. aureus. Moreover, the correlation between sea producing strains and type of infection as well as resistance to antibiotics is also considered. 128 S. aureus isolates randomly collected from different clinical samples in Tehran University Hospitals from February 2008 to June 2008. Patients’ information including sex, infection type and the hospital where samples come from were recorded. The sea gene was observed among 60 (46.9%) of clinical isolates. There was a significant correlation between prevalence of sea gene and type of infection (P = 0.01). Furthermore, significant correlation was detected between the presence of sea gene and resistance to the most of antibiotics used in this study. The significant relationship between the type of infection and S. aureus isolates carrying sea indicates the interaction quality of the S. aureus pathogen and the host as well as the pathogenic role of S. aureus.

"nBackground: Clindamycin is a suitable antibiotic for treatment of skin and soft tissue infections. Moreover, it can suppress toxin production in many pathogenic bacteria such as S. aureus. There are two mechanisms of resistance in this antibiotic. Constitutive resistance can be detected by standard disk diffusion method but in the case of inducible resistance, D-test should be carried out. The main aim of this study is to determine prevalence of clindamycin inducible resistance among methicillin resistant and susceptible isolates of S. aureus isolated from different clinical samples. "nMethods: A total of 87 clinical isolates from clinical samples were collected. Methicillin resistance was determined using standard disk diffusion method. Subsequently, D-test was carried out according to CLSI guideline. Presence of the sea gene (enterotoxin A) was detected by PCR using specific primers. "nResults: Out of 87 isolates, 18(20.7%) were clindamycin inducible resistant while constitutive resistance was detected among 21(24.1%) isolates. The 95% Confidence intervals for the proportion of inducible clindamycin resistance among clinical isolates of S. aureus was 12.2% to 29.2%. The inducible phenotype in MRSA isolates was more common than that of MSSA isolates (33.3% vs 5.1%).Significant differences were found between prevalence of inducible clindamycin resistance and type of infection (p=0.045). Importantly, there was a significant correlation between sea gene and the constitutive/inducible resistance (p

The aim of this study was to investigate the prevalence, molecular characteristics and antimicrobial susceptibility of S. aureus in raw eggs during production and sales in Yangling, Shaanxi Province, China, and assess whether enterotoxin-A-carrying strains could grow and produce enterotoxin A (SEA) in egg contents at 3 different temperatures (10, 25, and 37 °C) and culture times (1, 3, 5, and 7 days). A total of 102 isolates of 627 fresh eggs were identified, including 100 (15.9%) isolates from eggshells and 2 (0.3%) isolates from egg contents. All isolates harbored at least one virulence gene, hemolysin genes and novel enterotoxin genes were commonly detected. 95.1% of isolates were multi-drug resistant, with most commonly resistant to penicillin, trimethoprim, erythromycin, ciprofloxacin, and tetracycline. CC5-ST5-t002 (78.4%) was the most predominant clone. PFGE analysis found that isolates between the eggshell and egg contents of the same egg had the same dendrogram. In the test for artificial contamination of egg contents with S. aureus , three isolates could grow and produce SEA. This study indicates eggshells are often contaminated with high levels of S. aureus , and the discovery of S. aureus in egg contents implies the risk of entering the food chain. • The eggshell is often contaminated with high levels of S. aureus during production and sale. • Most S. aureus isolates harbored at least one virulence gene and exhibited multi-drug resistance. • CC5-ST5-t002 S. aureus was the dominant type among 102 S. aureus isolates. • Close genetic relatedness of S. aureus from eggshell and egg liquid was found based on PFGE analysis. • S. aureus could grow and product SEA at 10 °C, 25 °C and 37 °C in egg contents. [ABSTRACT FROM AUTHOR]

ST7 Staphylococcus aureus is highly prevalent in humans, pigs, as well as food in China; however, staphylococcal food poisoning (SFP) caused by this ST type has rarely been reported. On May 13, 2017, an SFP outbreak caused by ST7 S. aureus strains occurred in two campuses of a kindergarten in Hainan Province, China. We investigated the genomic characteristics and phylogenetic analysis of ST7 SFP strains combined with the 91 ST7 food-borne strains from 12 provinces in China by performing whole-genome sequencing (WGS). There was clear phylogenetic clustering of seven SFP isolates. Six antibiotic genes including blaZ, ANT (4')-Ib, tetK, lnuA, norA, and lmrS were present in all SFP strains and also showed a higher prevalence rate in 91 food-borne strains. A multiple resistance plasmid pDC53285 was present in SFP strain DC53285. Among 27 enterotoxin genes, only sea and selx were found in all SFP strains. A ФSa3int prophage containing type A immune evasion cluster ( sea, scn, sak , and chp ) was identified in SFP strain. In conclusion, we concluded that this SFP event was caused by the contamination of cakes with ST7 S. aureus . This study indicated the potential risk of new emergencing ST7 clone for SFP., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Guo, Yu, Wang, Hua, You, Wu, Ji, Zhang, Li, Hu, Wu, Wei, Jin, Meng, Yang, Hu, Long, Hu, Qi, Ma, Bei, Yan, Wang and He.)

The present study investigates the nature of the link between the staphylococcal enterotoxin A (SEA) gene and the lifecycle of Siphoviridae bacteriophages, including the origin of strain variation regarding SEA production after prophage induction. Five strains representing three different genetic lines of the sea region were studied under optimal and prophage-induced growth conditions and the Siphoviridae lifecycle was followed through the phage replicative form copies and transcripts of the lysogenic repressor, cro. The role of SOS response on prophage induction was addressed through recA transcription in a recA-disruption mutant. Prophage induction was found to increase the abundance of the phage replicative form, the sea gene copies and transcripts and enhance SEA production. Sequence analysis of the sea regions revealed that observed strain variances were related to strain capacity for prophage induction, rather than sequence differences in the sea region. The impact of SOS response activation on the phage lifecycle was demonstrated by the absence of phage replicative form copies in the recA-disruption mutant after prophage induction. From this study it emerges that all aspects of SEA-producing strain, the Siphoviridae phage and the food environment must be considered when evaluating SEA-related hazards. [ABSTRACT FROM AUTHOR]

Treatment of Staphylococcus aureus infections has become complicated owing to growing antibiotic resistance mechanisms and due to the multitude of virulence factors secreted by this organism. Failures with traditional monovalent vaccines or toxoids have brought a shift towards the use of multivalent formulas and neutralizing antibodies to combat and prevent range of staphylococcal infections. In this study, we evaluated the efficacy of a fusion protein (r-ET) comprising truncated regions of staphylococcal enterotoxin A (SEA) and toxic shock syndrome toxin (TSST-1) in generating neutralizing antibodies against superantigen induced toxicity in murine model. Serum antibodies showed specific reactivity to both SEA and TSST-1 native toxins. Hyperimmune serum from immunized animals protected cultured splenocytes from non-specific superantigen induced proliferation completely. Passive antibody administration prevented tissue damage from acute inflammation associated with superantigen challenge from S. aureus cell free culture supernatants. Approximately 80% and 50% of actively and passively immunized mice respectively were protected from lethal dose against S. aureus toxin challenge. This study revealed that r-ET protein is non-toxic and a strong immunogen which generated neutralizing antibodies and memory immune response against superantigen induced toxic effects in mice model. [ABSTRACT FROM AUTHOR]

Background: Staphylococcus aureus enterotoxins are known to induce an inflammatory response of the airways, increase sensitization to inhaled allergens, and decrease T-cell sensitivity to steroids. However, the effects of Staphylococcus enterotoxin A (SEA) on mucin secretion of airway epithelial cells still have not been reported. Therefore, in this study, the effect and brief signaling pathway of SEA on MUC5B expression were investigated in human airway epithelial cells. Methods: In the mucin-producing human NCI-H292 airway epithelial cells and the primary cultures of normal nasal epithelial cells, the effect and signaling pathway of SEA on MUC5B expression were investigated using reverse transcriptase-polymerase chain reaction (PCR), real-time PCR, enzyme immunoassay, and immunoblot analysis with several specific inhibitors and small interfering RNA (siRNA). Results: SEA increased MUC5B mRNA and protein expression. Toll-like receptor 2 (TLR2) mRNA expression was significantly increased after treatment with SEA. Knockdown of TLR2 by siRNA significantly blocked SEA-induced MUC5B mRNA expression. SEA significantly activated phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK). U0126 (ERK1/2 MAPK inhibitor) and SB203580 (p38 MAPK inhibitor) significantly inhibited SEA-induced MUC5B mRNA expression. In addition, knockdown of ERK1/2 and p38 MAPK by siRNA significantly blocked SEA-induced MUC5B mRNA expression. Furthermore, the phosphorylation of ERK1/2 and p38 MAPK was significantly blocked by knockdown of TLR2 by siRNA. Conclusion: These results show for the first time that SEA induces MUC5B expression via TLR2, ERK1/2, and p38 MAPK signaling pathway in human airway epithelial cells. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a major human pathogen producing different types of toxins. Enterotoxin A (SEA) is the most common type among clinical and food-related strains. The aim of the present study was to examine the prevalence of sea in clinical isolates of S. aureus. Moreover, the correlation between sea producing strains and type of infection as well as resistance to antibiotics is also considered. 128 S. aureus isolates randomly collected from different clinical samples in Tehran University Hospitals from February 2008 to June 2008. Patients’ information including sex, infection type and the hospital where samples come from were recorded. The sea gene was observed among 60 (46.9%) of clinical isolates. There was a significant correlation between prevalence of sea gene and type of infection (P = 0.01). Furthermore, significant correlation was detected between the presence of sea gene and resistance to the most of antibiotics used in this study. The significant relationship between the type of infection and S. aureus isolates carrying sea indicates the interaction quality of the S. aureus pathogen and the host as well as the pathogenic role of S. aureus.

The article presents a study of the dependence of staphylococcal enterotoxin production to temperature. 11 strains of Staphylococcus aureus was incubated at 10° to 37°C through a brain heart infusion broth and examined their enterotoxin with western blot analysis to evaluate the effect of incubation temperature to the production of A staphylococcal enteroxin (SEA). The study shows that the maximal concentration of SEA to most strains increased as the temperature increases.

Abstract: The effect of transient exposure of Staphylococcus aureus enterotoxin A (SEA) to high pressure and/or denaturing agents was examined by assessing the toxin superantigenicity and immunoreactivity, and by monitoring pressure-induced changes in fluorescence emission spectra. Pressurization of SEA at 600MPa and 45°C in Tris–HCl buffer (20mM, pH 7.4) resulted in a marked increase in both T-cell proliferation (superantigenicity) and immunoreactivity. In opposite, pressurization at 20°C did not change significantly SEA superantigenicity and immunoreactivity, indicating some toxin baro-resistance. Exposure of SEA to 8M urea at atmospheric pressure or at 600MPa and 20°C, also led to a marked increase of superantigenicity (but not of immunoreactivity). In contrast, exposure of SEA to sodium-dodecylsulfate (30mM) led to an increase of immunoreactivity with some effect on superantigenicity after pressurization at 45°C only. High pressure up to 600MPa induced spectral changes which at 20°C were fully reversible upon decompression. At 45°C, however, a sharp break of the centre of spectral mass mainly due to tryptophan residues was observed at 300MPa, and irreversible spectral changes mainly related to tyrosine residues subsisted after pressure release, indicating a marked protein conformational transition. Urea 8M further increased SEA structural changes at 600MPa and 20°C. These results indicate that SEA, under a combination of high pressure and mild temperature, as well as in the presence of urea, partly unfolds to a structure of strongly increased T-cell proliferative ability. [Copyright &y& Elsevier]

Background: Staphylococcus aureus is a major foodborne pathogen throughout the world. Enterotoxins and toxic shock syndrome toxin-1 are important virulence factors and as pyrogenic toxin superantigens have profound effects on the ir host. Thus circulation of TSST1 producing S.aureus among people and food chain is a worrying issue. The present paper was conducted to study Prevalence of tst, entC, entA and entA/C genes in staphylococcus aureus strains isolated from different foods. Methods: Over 1040 food samples have been analyzed differentially according to Iran national standard (number= 1194) for S.aureus identification. After DNA extraction, PCR reactions were carried out by reference strain as positive control, adequate primers. Results: At present study, prevalence of foodstuffs contaminated by S.aureus isolates was about 9.5% (100 strains). Of 25% of isolates producing entC, 28% (seven strains) had tst gene at the same time and of 8% of isolates producing entA, 12.5% (one strain) were positive for tst genes simultaneously. Altogether of 9% isolates producing combination of entC and entA, 44.4% (four strains) were also producer of tst gene. Conclusion: Prevalence of TSST1 producing strains in combination with enterotoxin genes is considerable especially with entC and A plus C. On the other hand, circulation of these isolates in humans, animals, foods and environment has hazardous effect for general public health. [ABSTRACT FROM AUTHOR]

Methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogen of nosocomial infection, which is resistant to most antibiotics. Presently, anti-virulence therapy and anti-biofilm therapy are considered to be promising alternatives. In the current work, we investigated the influence of bisdemethoxycurcumin (BDMC) on the virulence-related exoproteins and the biofilm formation using a reference strain and clinic isolated strains. Western blotting, quantitative RT-PCR, and tumor necrosis factor (TNF) release assay were performed to assess the efficacy of BDMC in reducing the expression of Staphylococcus enterotoxin-related exoproteins (enterotoxin A, enterotoxin B) and α-toxin in MRSA. The anti-biofilm activity of BDMC was evaluated through a biofilm inhibition assay. The study suggests that sub-inhibitory concentrations of BDMC significantly inhibited the expression of sea, seb, and hla at the mRNA level in MRSA. Moreover, the expression of virulence-related exoproteins was significantly decreased by down-regulating accessory gene regulator agr, and the inhibition of biofilms formation was demonstrated by BDMC at sub-inhibitory concentrations. Consequently, the study suggests that BDMC may be a potential natural antibacterial agent to release the pressure brought by antibiotic resistance. [ABSTRACT FROM AUTHOR]

s. ananou, e. valdivia, m. martínez bueno, a. gálvez and m. maqueda. 2004. Control of the enterotoxigenic Staphylococcus aureus CECT 976 strain by enterocin AS-48 in laboratory cultures, and behaviour of the AS-48 activity in the presence of food preservatives. Enterocin AS-48 shows inhibitory activity on the majority of the Staphylococcus species tested. This enterocin has a bactericidal and bacteriolytic mode of action on S. aureus CECT 976, a strain selected for this study by its enterotoxigenic character (SEA production). The inhibitory effect of AS-48 was pH and temperature dependent, and enterocin activity was higher at pH 5. The minimum bactericidal concentration (MBC) of AS-48, decreased from 15 μg ml−1 at 37°C to 10 μg ml−1 at 15°C. Sublethally injured cells showed an increased sensitivity with a MBC of 5 μg ml−1. In this way, the highest effectiveness of Ent AS-48 against S. aureus CECT 976 was obtained at 4°C in combination with high concentrations of NaCl (6 and 7%). Interestingly, enterotoxin SEA production by strain CECT 976 was markedly inhibited by subinhibitory concentrations of Ent AS-48. These low concentrations also provoked a delay of bacterial growth. The results presented indicated that Ent AS-48 has a potential for application as a protective agent against S. aureus in foods. In this study, we have established the conditions for an efficient inhibition of growth and enterotoxin production by S. aureus CECT 976 in culture media by a combination of environmental factors and Ent AS-48. [ABSTRACT FROM AUTHOR]

Activation of lymphocytes by interleukin-2 (IL-2) induces lymphokine-activated killer (LAK) cells that show promising effects on tumour growth in clinical trials. We examined the effect of the superantigen staphylococcal enterotoxin A (SEA) on anti-tumour activity of freshly prepared human lymphocytes. Picomolar amounts of SEA rapidly induced cytotoxic activity against K562 and Raji cells as well as some natural-killer(NK)-resistant tumour cell lines. Cytotoxic activity was not dependent on target cell expression of either major histocompatibility complex (MHC) class I or II antigens as shown using mutated cell lines. Cell-sorting experiments showed that the activity was expressed by NK (CD5CD56) as well as T (CD5) cells, although the former contained the majority of cytotoxic activity. NK cells could not be directly activated by SEA. In contrast, SEA activated purified T cells to the same extent as in bulk cultures. It is suggested that SEA activation of NK cells is secondary to that brought about by lymphokines produced by T cells. Activation of LAK cells with SEA was comparable in magnitude as well as target cell spectrum to that of IL-2. In addition to the LAK-like cytotoxic activity induced by SEA, a superimposed cytotoxicity towards target cells expressing MHC class II antigens coated with SEA was observed. This staphylococcal-enterotoxin-dependent cell-mediated cytotoxicity (SDCC) was exclusively mediated by T cells. It is well established that MHC class II antigens function as receptors for staphylococcal enterotoxins on mammalian cells and that the complex between MHC class II antigen and - SEA apparently functions as a target structure for activated T cells with target cell lysis as a consequence. Activation of T lymphocytes with IL-2 also resulted in the capability to mediate SDCC. Staphylococcal enterotoxins represent a novel way of inducing anti-tumour activity in human lymphocytes, which could be of value in therapeutic applications. [ABSTRACT FROM AUTHOR]

A simple, cheap, and less aggressive immobilization procedure for biomolecules using reduced graphene oxide (rGO) was employed to prepare an impedimetric immunosensor for detection of staphylococcal enterotoxin A (SEA) from Staphylococcus aureus in milk samples. The scanning electron microscopy, cyclic voltammetry, and electrochemical impedance spectroscopy (EIS) were used to monitor the single steps of the electrode assembly process. The glassy carbon (GC)/rGO platform detected the antigen-antibody binding procedures of SEA with concentrations of 0.5 to 3.5 mg L−1 via impedance changes in a low frequency range. The impedimetric immunosensor was successfully applied for the determination of SEA in milk samples. [ABSTRACT FROM AUTHOR]