Search Results (79)

Foodborne diseases are the most common causes of illness worldwide. Bacterial pathogens, including Staphylococcus aureus, are often involved in foodborne disease and pose a serious threat to human health. S. aureus is commonly found in humans and a variety of animal species. Staphylococcal enteric disease, specifically staphylococcal food poisoning (SFP), accounts for numerous gastrointestinal illnesses, through the contamination of food with its enterotoxins, and its major impact on human health imposes a heavy economic burden in society. Commonly, antibiotics and antimicrobials are used to treat SFP. However, a range of complications may arise with these treatments, impeding the control of S. aureus diseases specifically caused by methicillin-resistant S. aureus (MRSA). Natural alternative options to control S. aureus diseases, such as bacteriophages, plant-based antimicrobials, nanoparticle-based or light-based therapeutics, and probiotics, are promising in terms of overcoming these existing problems as they are environmentally friendly, abundant, unlikely to induce resistance in pathogens, cost-effective, and safe for human health. Recent findings have indicated that these alternatives may reduce the colonization and infection of major foodborne pathogens, including MRSA, which is crucial to overcome the spread of antibiotic resistance in S. aureus. This review focuses on the present scenario of S. aureus in foodborne disease, its economic importance and current interventions and, most importantly, the implications of natural antimicrobials, especially probiotics and synbiotics, as alternative antimicrobial means to combat pathogenic microorganisms particularly, S. aureus and MRSA. Full article

Staphylococcus aureus is a leading cause of foodborne intoxication globally, driven by its heat-stable enterotoxins (SEs), which pose significant public health risks. This review critically evaluates modern and traditional methodologies for detecting S. aureus and its enterotoxins in food matrices, emphasizing their principles, applications, and limitations. The review includes a dedicated section on sample preparation and pretreatment methods for diverse food substrates, addressing a critical gap in practical applications. Immunological techniques, including ELISA and lateral flow assays, offer rapid on-site screening but face matrix interference and variable sensitivity challenges. Molecular methods, such as PCR and isothermal amplification, provide high specificity and speed for bacterial and toxin gene detection but cannot confirm functional toxin production. Sequencing-based approaches (e.g., WGS and MLST) deliver unparalleled genetic resolution for outbreak tracing but require advanced infrastructure. Emerging biosensor technologies leverage nanomaterials and biorecognition elements for ultra-sensitive real-time detection, although scalability and matrix effects remain hurdles. Mass spectrometry (MALDI-TOF MS) ensures rapid species identification but depends on pre-isolated colonies. Traditional microbiological methods, while foundational, lack the precision and speed of molecular alternatives. The review underscores the necessity of context-driven method selection, balancing speed, sensitivity, and resource availability. Innovations in multiplexing, automation, AI-based methods, and integration of complementary techniques are highlighted as pivotal for advancing food safety surveillance. Standardized validation protocols and improved reporting of performance metrics are urgently needed to enhance cross-method comparability and reliability in outbreak settings. Full article

Staphylococcal enterotoxins (SEs) are major contributors to foodborne intoxications. Reliable detection methods for SEs are essential to maintain food safety and protect public health. Since the heat-stable toxins also exert their toxic effect in the absence of the bacterium, reliance on DNA detection alone can be misleading: it does not allow for determining which specific toxins encoded by a given strain are produced and epidemiologically linked with a given outbreak. Commercially available diagnostic assays for SE detection are so far limited in sensitivity and specificity as well as in the range of targeted toxins (SEA–SEE), thus non-targeted SEs linked to foodborne illness remain undetected at the protein level. This study aimed to develop a highly sensitive and specific multiplex suspension immunoassay (SIA) for SEA to SEI. To this end, high-affinity monoclonal antibodies (mAbs) for the specific detection of the individual SEs were generated. When implemented in sandwich ELISAs and multiplex SIA, these mAbs demonstrated exceptional sensitivity with detection limits in the low picogram per millilitre range. When applied for the analysis of SE production in liquid cultures of a panel of 145 whole-genome sequenced strains of Staphylococcus spp. and Enterococcus faecalis, the novel multiplex SIA detected and differentiated the eight SEs with assay accuracies of 86.9–100%. Notably, the multiplex SIA covered one to four sequence variants for each of the individual SEs. Validation confirmed high recovery rates and reliable performance in three representative complex food matrices. The implementation of the novel mAbs in a multiplex SIA enabled, for the first time, simultaneous detection, differentiation, and quantification of multiple SEs from minimal sample volumes using Luminex^® technology. As a result, the multiplex SIA will help strengthen food safety protocols and public health response capabilities. Full article

Antibiotic-resistant Staphylococcus aureus (S. aureus) in retail meat poses a public health threat requiring continuous surveillance. This study investigated the frequency of isolation, toxin genes, and antibiotic resistance profile of S. aureus recovered from retail poultry meat samples and presented results beneficial to public health interventions. Of 200 samples collected, 16% (32/200) tested positive for S. aureus, and these were recovered from thigh 37.5% (12/32), wing 34.4% (11/32), gizzard (15.6% (5/32), and liver 12.5% (4/32) samples. Findings of spa typing analysis revealed that 68.8% (22/32), 18.8% (6/32), 9.4% (3/32), and 3.0% (1/32) of the isolates belonged to the spa types t267, t160, t548, and t008, respectively. For antibiotic susceptibility testing, 12.5% (4/32) of the isolates were resistant to only penicillin, but one isolate (1/32; 3%) showed resistance to the antibiotics penicillin, erythromycin, ampicillin, and oxacillin. PCR analysis revealed that 9.4% (3/32) of the isolates carried the mecA gene associated with methicillin-resistant Staphylococcus aureus (MRSA) isolates. One MRSA isolate was identified as a t008 spa type, and harbored a 26,974 bp-sized plasmid, which was the source of its resistance to penicillin, ampicillin, erythromycin, and oxacillin. The staphylococcal enterotoxin (SE) genes seg, sei, sek, seb, selm, and seln were also identified among the isolates, and mostly the antimicrobial and enterotoxin genes were carried on plasmids of the isolates. This study raises awareness on the continuous circulation of pathogenic microbes like S. aureus in retail poultry meat. Full article

Food poisoning outbreaks frequently involve staphylococcal enterotoxins (SEs). SEs include 33 distinct types and multiple sequence variants per SE type. Various mass spectrometry methods have been reported for the detection of SEs using a conventional bottom-up approach. However, the bottom-up approach cannot differentiate between all sequence variants due to partial sequence coverage, and it requires a long trypsin digestion time. While the alternative top-down approach can theoretically identify any sequence modifications, it generally provides lower sensitivity. In this study, we optimized top-down mass spectrometry conditions and incorporated a fully ¹⁵N-labeled SEA spiked early in the protocol to achieve sensitivity and repeatability comparable to bottom-up approaches. After robust immunoaffinity purification of the SEA, mass spectrometry signals were acquired on a Q-Orbitrap instrument operated in full-scan mode and targeted acquisition by parallel reaction monitoring (PRM), enabling the identification of sequence variants and precise quantification of SEA. The protocol was evaluated in liquid and solid dairy products and demonstrated detection limits of 0.5 ng/mL or ng/g in PRM and 1 ng/mL or ng/g in full-scan mode for milk and Roquefort cheese. The top-down method was successfully applied to various dairy products, allowing discrimination of contaminated versus non-contaminated food, quantification of SEA level and identification of the variant involved. Full article

Staphylococcus aureus stands out as one of the most virulent pathogens in the genus Staphylococcus. This characteristic is due to its ability to produce a wide variety of staphylococcal enterotoxins (SEs) and exotoxins, which in turn can cause staphylococcal food poisoning (SFP), clinical syndromes such as skin infections, inflammation, pneumonia, and sepsis, in addition to being associated with the development of inflammation in the mammary glands of dairy cattle, which results in chronic mastitis and cell necrosis. SEs are small globular proteins that combine superantigenic and emetic activities; they are resistant to heat, low temperatures, and proteolytic enzymes and are tolerant to a wide pH range. More than 24 SE genes have been well described (SEA-SEE, SEG, SEH, SEI, SEJ, SElK, SElL, SElM, SElN, SElO, SElP, SElQ, SElR, SElS, SElT, SElU, SElV, SElW, SElX, SElY, and SElZ), being a part of different SFP outbreaks, clinical cases, and isolated animal strains. In recent years, new genes (sel26, sel27, sel28, sel31, sel32, and sel33) from SEs have been described, as well as two variants (seh-2p and ses-3p) resulting in a total of thirty-three genes from Ses, including the nine variants that are still in the process of genetic and molecular structure evaluation. SEs are encoded by genes that are located in mobile genetic elements, such as plasmids, prophages, pathogenicity islands, and the enterotoxin gene cluster (egc), and housed in the genomic island of S. aureus. Both classical SEs and SE-like toxins (SEls) share phylogenetic relationships, structure, function, and sequence homology, which are characteristics for the production of new SEs through recombination processes. Due to the epidemiological importance of SEs, their rapid assessment and detection have been crucial for food security and public health; for this reason, different methods of identification of SEs have been developed, such as liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS), molecular methods, and whole-genome sequencing; providing the diagnosis of SEs and a better understanding of the occurrence, spread, and eradication of SEs. This review provides scientific information on the enterotoxins produced by S. aureus, such as structural characteristics, genetic organization, regulatory mechanisms, superantigen activity, mechanisms of action used by SEs at the time of interaction with the immune system, methods of detection of SEs, and recent biocontrol techniques used in food. Full article

Food poisoning caused by Staphylococcal enterotoxins (SEs) is prevalent globally, making efficient detection of these toxins very important. Traditionally, liquid chromatography–mass spectrometry required immunosorbent enrichment by magnetic bead-coupled antibodies obtained by animal-specific immunization. However, this method is time-consuming and costly. In this study, two recombinant protein capture molecules were designed based on the principle of toxins binding to Major Histocompatibility Complex (MHCII) and T cell receptor (TCR) molecules. The two capture molecules are called MHCII and MHCII-D10. The design of the MHCII and TCR-D10 was achieved through searching for the binding site protein sequence of Staphylococcal enterotoxins in the relevant literature, and MHCII-D10 was to link MHCII sequence with TCR-D10 sequence using linker (G4S)3 linking peptide. These capture molecules were shown to effectively bind to seven types of toxins and to capture SEs in various matrices. The digestion time, ratio, and temperature were further optimized, reducing the overall digestion time to just 2 h. The specificity, linearity, sensitivity, precision (RSD%), and recovery of the two methods were verified by liquid chromatography–mass spectrometry. When the MHCII and MHCII-D10 captured the toxins, the limit of quantification (LOD) in the 1 × PBS, plasma, and milk matrices ranged from 1.5625 to 100 fmol/µL, with the recovery rate ranging from 18.4% to 96%. The design of these capture molecules eliminates the need for animal-specific immunization, simplifying the pre-detection process and avoiding ethical concerns. This development holds significant promise for clinical diagnosis and reference. Full article

Staphylococcus aureus stands as one of the most pervasive pathogens given its morbidity and mortality worldwide due to its roles as an infectious agent that causes a wide variety of diseases ranging from moderately severe skin infections to fatal pneumonia and sepsis. S. aureus produces a variety of exotoxins that serve as important virulence factors in S. aureus-related infectious diseases and food poisoning in both humans and animals. For example, staphylococcal enterotoxins (SEs) produced by S. aureus induce staphylococcal foodborne poisoning; toxic shock syndrome toxin-1 (TSST-1), as a typical superantigen, induces toxic shock syndrome; hemolysins induce cell damage in erythrocytes and leukocytes; and exfoliative toxin induces staphylococcal skin scalded syndrome. Recently, Panton–Valentine leucocidin, a cytotoxin produced by community-associated methicillin-resistant S. aureus (CA-MRSA), has been reported, and new types of SEs and staphylococcal enterotoxin-like toxins (SEls) were discovered and reported successively. This review addresses the progress of and novel insights into the molecular structure, biological activities, and pathogenicity of both the classic and the newly identified exotoxins produced by S. aureus. Full article

Certain Staphylococcus aureus strains harbour staphylococcal enterotoxin genes and hence can produce enterotoxin during their growth in food. Therefore, food can be a source of staphylococcal food poisoning, one of the most common food-borne diseases worldwide. Epidemiological data show that S. aureus is often present in raw milk cheeses, and consequently, cheeses are often the source of staphylococcal food poisoning outbreaks. The aim of this study was to determine the phenotypic characteristics of S. aureus isolates from fresh cheese, including antibiotic susceptibility; the presence of classical sea-see enterotoxin genes through molecular methods; and the isolate’s ability to produce SEA-SEE enterotoxins in vitro through reversed passive latex agglutination. A total of 180 coagulase-positive staphylococci were isolated from 18 out of 30 cheese samples, and 175 were confirmed as S. aureus through latex agglutination and API STAPH tests. All isolates possessed phenotypic characteristics typical for S. aureus, with certain variations in the egg yolk reaction (18.3% of the isolates showed a weak reaction and 28% no reaction at all) and haemolysis pattern (36.6% of the isolates produced double-haemolysis and 4.6% were non-haemolytic). Antibiotic resistance was observed in 1.1% of the isolates and to mupirocin only. Real-time PCR detected the sec gene in 34 (19.4%) isolates, but most isolates (80.6%) were not enterotoxigenic. For all 34 (19.4%) strains that carried the sec gene, the RPLA method detected the production of the SEC enterotoxin in vitro. For those enterotoxigenic strains, the possibility of enterotoxin production in fresh cheese could not be ruled out. Full article

A Staphyloccoccus aureus is one of the leading causes of food poisoning outbreaks (FPOs) worldwide. Staphylococcal food poisoning (SFP) is induced by the ingestion of food containing sufficient levels of staphylococcal enterotoxins (SEs). Currently, 33 SEs and SE-like toxins (SEls) have been described in the literature, but only five named “classical” enterotoxins are commonly investigated in FPOs due to lack of specific routine analytical techniques. The aims of this study were to (i) establish the genetic profile of strains in a variety of artisanal cheeses (n = 30) in Belgium, (ii) analyze the expression of the SE(l)s by these strains and (iii) compare the output derived from the different analytical tools. Forty-nine isolates of S. aureus were isolated from ten Belgian artisanal cheeses and were analyzed via microbiological, immunological, liquid chromatography mass spectrometry, molecular typing and genetic methods. The results indicated that classical SEs were not the dominant SEs in the Belgian artisanal cheeses that were analyzed in this study, and that all S. aureus isolates harbored at least one gene encoding a new SE(l). Among the new SE(l)s genes found, some of them code for enterotoxins with demonstrated emetic activity and ecg-enterotoxins. It is worth noting that the involvement of some of these new SEs has been demonstrated in SFP outbreaks. Thus, this study highlighted the importance of the development of specific techniques for the proper investigation of SFP outbreaks. Full article

Staphylococcus argenteus is a novel Staphylococcus species derived from Staphylococcus aureus. Information on the prevalence and genetic characteristics of invasive S. argenteus in Asia is limited. In this study, 275 invasive S. aureus complex strains were retrieved from blood culture specimens in Hong Kong and re-analyzed using MALDI-TOF mass spectrometry and an in-house multiplex real-time PCR for S. argenteus. The prevalence of invasive S. argenteus in Hong Kong was found to be 4.0% (11/275). These strains were primarily susceptible to commonly used antibiotics, except penicillin. Whole-genome sequencing revealed the circulation of three S. argenteus genotypes (ST-2250, ST-1223, and ST-2854) in Hong Kong, with ST-2250 and ST-1223 being the predominant genotypes. The local ST-2250 and ST-1223 strains showed close phylogenetic relationships with isolates from mainland China. Antimicrobial-resistant genes (fosB, tet-38, mepA, blaI, blaZ) could be found in nearly all local S. argenteus strains. The ST-1223 and ST-2250 genotypes carried multiple staphylococcal enterotoxin genes that could cause food poisoning and toxic shock syndrome. The CRISPR/Cas locus was observed only in the ST-2250 strains. This study provides the first report on the molecular epidemiology of invasive S. argenteus in Hong Kong, and further analysis is needed to understand its transmission reservoir. Full article

Staphylococcal food poisoning results from the consumption of food contaminated by staphylococcal enterotoxins. In July 2022, the Turin local health board was notified of a suspected foodborne outbreak involving six children who had consumed döner kebab purchased from a takeaway restaurant. The symptoms (vomiting and nausea) were observed 2–3 h later. A microbiological analysis of the food samples revealed high levels (1.5 × 10⁷ CFU/g) of coagulase-positive staphylococci (CPS). The immunoassay detected a contamination with staphylococcal enterotoxins type B (SEB). The whole genome sequencing of isolates from the food matrix confirmed the staphylococcal enterotoxin genes encoding for type B, which was in line with the SEB detected in the food. This toxin is rarely reported in staphylococcal food poisoning, however, because there is no specific commercial method of detection. The involvement of enterotoxin type P (SEP) was not confirmed, though the corresponding gene (sep) was detected in the isolates. Nasal swabs from the restaurant food handlers tested positive for CPS, linking them to the likely source of the food contamination. Full article

Staphylococcus aureus (S. aureus) is a major foodborne pathogen. Rapid and specific detection is crucial for controlling staphylococcal food poisoning. This study reported a Staphylococcus phage named LSA2302 showing great potential for applications in the rapid detection of S. aureus. Its biological characteristics were identified, including growth properties and stability under different pH and temperature conditions. The genomic analysis revealed that the phage has no genes associated with pathogenicity or drug resistance. Then, the phage-functionalized magnetic beads (pMB), serving as a biological recognition element, were integrated with ATP bioluminescence assays to establish a biosensing method for S. aureus detection. The pMB enrichment brought high specificity and a tenfold increase in analytical sensitivity during detection. The whole detection process could be completed within 30 min, with a broad linear range of 1 × 10⁴ to 1 × 10⁸ CFU/mL and a limit of detection (LOD) of 2.43 × 10³ CFU/mL. After a 2 h pre-cultivation, this method is capable of detecting bacteria as low as 1 CFU/mL. The recoveries of S. aureus in spiked skim milk and chicken samples were 81.07% to 99.17% and 86.98% to 104.62%, respectively. Our results indicated that phage-based biosensing can contribute to the detection of target pathogens in foods. Full article

Enzyme-linked immunosorbent assay (ELISA) is one of the most commonly used method for the detection of staphylococcal enterotoxin B (SEB), the main protein toxin causing staphylococcal food poisoning. However, the traditional ELISA reaction needs to be stopped by sulfuric acid to obtain stable colorimetric signal, and it is easily influenced by a colored sample. In order to address this problem, a new ELISA method using zeolite imidazolate skeleton-8 metal-organic framework (ZIF-8 MOF) as a light scattering (LS) reporter for SEB detection was developed in this work. ZIF-8 MOF has the characteristics of high porosity, large specific surface area, clear pore structure, and adjustable size, which is one of the most representative MOFs constructed from Zn²⁺ and 2-methylimidazole (2-mIM). The 2-mIM ligand of ZIF-8 exhibited antioxidant activity and can strongly react with H₂O₂, which could destroy the structure of ZIF-8, resulting in the obvious decrease in LS intensity. We combined this specific reaction with the sandwich immune reaction to construct the LS ELISA method for the successful detection of SEB. This method is more reliable than commercial tests kits for the detection of colored samples, and it is simple, sensitive, and selective, and has great potential in the detection of other toxins by simply changing the corresponding recognition units. Full article

The aim of this study was to evaluate the biofilm-forming ability and the resistance of planktonic cells and biofilm to methicillin (MIC and MBC, and log CFU/cm² reduction, respectively). Thirty-four isolates were used, including coagulase-positive Staphylococcus (CPS) and coagulase-negative Staphylococcus (CNS), recovered from ready-to-eat (RTE) foods. Biofilm-forming ability was based on enumeration of viable cells from biofilms formed on three types of surfaces, namely stainless steel, polystyrene, and glass. Thirteen of the thirty-four isolates (38.2%) were methicillin-resistant (MIC higher than 4 µg/mL). Staphylococcus aureus (CPS) and Staphylococcus saprophyticus (CNS) were the species that showed the highest percentage of resistance (50% and 71.4%, respectively). Staphylococcus warneri was the only species susceptible to methicillin. In 20 of the 34 isolates, the MBC value was twice the MIC value. The CPS (S. aureus) and the CNS (the other species) were not significantly different (p > 0.05) on biofilm-forming ability on the three surfaces used. However, when comparing the averages obtained for the biofilm-forming ability of the isolates, the values obtained (log CFU/cm²) in polystyrene were significantly higher (p < 0.05) than those obtained with the isolates on stainless steel. On the other hand, data from biofilm-forming ability of the isolates on glass and stainless steel are positively and significantly correlated with each other (r = 0.54; p = 0.02). It was not possible to determine the concentration of methicillin that promotes biofilm removal since log reductions were less than 3 log CFU/cm². The results of this study indicate that foodborne CPS and CNS can form biofilms on different types of material. As these biofilms are resistant to high concentrations of methicillin, their occurrence in food environments and their spread to medical settings can result in staphylococcal food poisoning or, in the worst-case scenario, septicemia, respectively. Good hygiene and good manufacturing practices (GHP and GMP) are therefore mandatory to prevent contamination with Staphylococcus spp. Full article