Search Results (221)

Staphylococcus aureus is a major bacterial pathogen that can cause clinical infections and foodborne illnesses through the production of 25 exotoxin types. The most frequently implicated toxins in food poisoning outbreaks are Staphylococcal enterotoxins type A–E (SEA-SEE), which are the first enterotoxins discovered. While in vitro detection methods are available to identify the presence of enterotoxins, they cannot distinguish between biologically active and inactive forms of the toxins. Detection of biologically active enterotoxins currently relies on in vivo testing, using the emetic response in kittens or monkeys. Here, we show the development of an in vitro assay to detect the active forms of SEB, a potential biological warfare agent and leading cause of food poisoning, and SEC1, a frequent cause of staphylococcal food poisoning. The novel assay involves the implementation of a genetically engineered Jurkat T-cell line expressing TCR Vβ3, resulting in a dose response of IL-2 production when exposed to active toxin. We also show that at a concentration of 100 ng/mL, the biological activity of SEB is significantly decreased at temperatures over 70 °C, while pasteurization at 63 °C only slightly reduces the biological activity of the toxin. Our studies provide an alternative method to animal testing to determine the presence of active toxins and provide possible inactivation methods of the toxins. Full article

Staphylococcus aureus is a major cause of foodborne intoxication through the production of heat-stable enterotoxins (SEs) and is also an important opportunistic pathogen of humans and livestock. Meat and meat products are major vehicles for this pathogen because their protein-rich composition supports bacterial growth and toxin production. However, the combined effects of temperature and nutrient composition on S. aureus growth and virulence expression under food-relevant conditions remain unclear. In this study, we investigated the interactive effects of temperature and nutritional context on the growth and virulence-associated phenotypes under model food-relevant environments with the reference strain S. aureus FRI-S6. Bacterial growth, biofilm formation, staphylococcal enterotoxins A and B (SEA, SEB), and hemolytic activity were evaluated at 25 °C and 37 °C in brain heart infusion (BHI) medium supplemented with NaCl, glucose, or tryptone to simulate diverse food-relevant conditions. Growth was generally faster at 37 °C, whereas glucose-supplemented cultures at 25 °C reached higher cell densities during prolonged incubation. Biofilm formation increased at 37 °C in BHI and glucose conditions. SEA production was enhanced at 37 °C under NaCl and tryptone, but at 25 °C in glucose-rich conditions. In contrast, SEB production and hemolytic activity were consistently higher at 37 °C, particularly in the presence of tryptone and glucose. These findings demonstrate the strong interaction between temperature and nutrient composition in shaping S. aureus virulence in food environments and provide important insights for food safety risk assessment and highlight practical implications for controlling enterotoxin production in meat products and other foods during storage and processing. Full article

Methicillin-resistant Staphylococcus aureus (MRSA) is a major antimicrobial-resistant pathogen affecting both human and animal health. Although historically associated with healthcare settings, MRSA is now established in livestock production and throughout the production chain. Its detection in animals, food products, and processing environments reflects the complex ecology of antimicrobial resistance (AMR) in modern food systems. This narrative review synthesizes current evidence on the molecular basis of methicillin resistance and multidrug resistance determinants, as well as the epidemiology of MRSA in food-associated settings. Particular emphasis is placed on its occurrence in animal-derived foods and key reservoirs within farms, slaughterhouses, and processing environments. Livestock-associated populations are dominated by clonal complex CC398. In contrast, CC9 is prevalent in pig production systems in Asia, while CC5-related lineages occur at the human and animal interface. MRSA has been detected in retail meat and animal-derived foods at low but measurable prevalence, indicating contamination during slaughter and processing. Virulence determinants include staphylococcal enterotoxins linked to food poisoning and Panton–Valentine leukocidin associated with severe infections. Biofilm formation and adhesins further support persistence and colonization. Epidemiological and molecular evidence indicates that livestock, processing environments, and food-contact surfaces act as interconnected reservoirs sustaining MRSA circulation. Human exposure occurs primarily through occupational contact and environmental pathways, whereas foodborne transmission appears less common. Effective control requires integrated surveillance, responsible antimicrobial use in livestock production, and strict hygiene practices throughout the production chain within a One Health framework. Full article

Raw milk dairy products, an integral part of Italian food heritage, are the primary products of small-scale farms in mountain regions where pasture is seasonal. While raw milk dairy products offer potential health benefits, their physicochemical properties make them susceptible to foodborne pathogens. Long-term surveillance of these products is essential to safeguard consumer health. Here, we present a fourteen-year microbiological surveillance of raw milk dairy products and intermediate matrices from northeastern Italy’s alpine areas, analyzing coagulase-positive Staphylococci (CPS), β-glucuronidase-positive Escherichia coli, Listeria monocytogenes, and Shiga toxin-producing E. coli (STEC). The most frequently detected pathogens were CPS and β-glucuronidase-positive E. coli, with up to 19.6% and 51.7% of samples exceeding regulatory limits, respectively. Butter, curd, and fresh cream were the most contaminated matrices. Detection rates of staphylococcal enterotoxins, L. monocytogenes, and STEC aligned with European detection averages (6.7%, 2.6%, and 2.1%, respectively). These findings underscore the necessity of Good Hygiene and Management Practices, together with regular microbiological monitoring to mitigate contamination risks, supporting the safety and quality of traditional raw milk dairy products in alpine regions. Full article

Background/Objectives: Staphylococcus aureus (S. aureus) intramammary infection remains a major global dairy problem due to its contagious nature, its ability to persist and colonize teat/skin and mucosal niches, and the often-limited bacteriological cure achieved with antimicrobial therapy. Beyond udder health, it is relevant to public health because it can enter raw milk chains and serve as a reservoir for antimicrobial resistance determinants that may circulate between dairy animals and humans. Methods: We assessed S. aureus’ ecology in raw ewe milk from 75 sheep farms in Epirus (Greece) by sampling clinically healthy controls (group A) and clinical mastitis cases pre-treatment (group B), followed by resampling at the first post-withdrawal milking after penicillin/streptomycin treatment (group C1—therapeutic protocol 1), oxytetracycline treatment (group C2—therapeutic protocol 2), or enrofloxacin treatment (group C3—therapeutic protocol 3). Results: S. aureus detection was high and comparable across groups (A 23.0%, B 22.0–30.0%, C 20.0–22.0%), and paired analyses showed no significant pre–post shifts in detection/burden within therapeutic protocols (all p > 0.05). Nevertheless, persistence remained evident. The chromosomal gene mecA was detected in S. aureus strains in all groups, ranging from 13.6% in controls to 54.5% post-withdrawal in group C1, and was also present in the pre-treatment group. In paired sampling animals, mecA was mostly stable, with rare emergence or loss. Across antibiotic classes, within-animal resistance transitions were generally uncommon and non-significant (p > 0.05); β-lactam resistance was fully stable (p = 1.00). Descriptively, resistance to protein synthesis inhibitors tended to decline after therapy in protocol 1 and protocol 3, while protocol 3 showed post-treatment gains in fluoroquinolone resistance. By contrast, virulence-associated phenotype traits shifted after therapy: enterotoxigenicity increased post-withdrawal (especially in the C3 group), Staphylococcal Enterotoxin A (SEA) and Staphylococcal Enterotoxin B (SEB) appeared only post-therapy, Staphylococcal Enterotoxin D (SED) increased significantly in paired isolates (p = 0.002), and strong biofilm adherence increased (in C3, p = 1.5 × 10⁻⁵). Conclusions: The detection of S. aureus after therapy suggests that one possibility is that antimicrobial exposure may select for, or otherwise reshape, the residual intramammary population, rather than reliably eliminating it—an outcome that remains clinically relevant for udder health. Moreover, the persistence of mecA/methicillin-resistant Staphylococcus aureus (MRSA)-compatible profiles indicates that milk released to the food chain after withdrawal compliance may still harbor S. aureus with enhanced preservation capacity and significant food safety relevance. Full article

Bacterial toxins, such as Staphylococcal Enterotoxin B (SEB) and Shiga Toxins (STX1, STX2), pose severe public health risks and significant biological threats, demanding rapid and precise qPCR detection. This study reports the initial stages of standardization for uniplex qPCR components, serving as a foundational step toward a future multiplex detection system. The annealing temperature was successfully optimized for the stx1, seb, and 16S rRNA targets, showing high consistency in Cq values and fluorescence intensities at 58.4 °C. The optimized primer ratios ensure maximum amplification efficiency while minimizing potential molecular competition. These optimized assays provide a robust foundation for the multiplex qPCR platform. Once fully validated, this protocol will enhance timely and effective responses, as well as improve preparedness and readiness within the Brazilian biodefense system. Full article

Computer devices in university settings are frequently shared and repeatedly handled, making them potential reservoirs for pathogenic bacteria. This study aimed to investigate the prevalence, virulence determinants, and antimicrobial resistance profiles of Staphylococcus aureus and Escherichia coli isolated from computer devices used by staff and students at a university in Northern Thailand. A total of 400 computer devices were sampled, with each device defined as a single sampling unit comprising both the keyboard and computer mouse. Bacterial identification was performed using PCR, while staphylococcal enterotoxin (se) genes and diarrheagenic E. coli (DEC)-associated virulence genes were detected by PCR. Antimicrobial susceptibility was assessed using the disk diffusion method. Overall, 74 (18.5%) S. aureus isolates and 6 (1.5%) E. coli isolates were recovered. The highest prevalence of S. aureus was observed among personal-use student computer devices (29%; p < 0.001), whereas E. coli was most frequently detected on public-use staff computer devices (4%). Among S. aureus isolates, 24.3% (18/74) carried at least one se gene, with sec being the most prevalent (13.5%). Half of the E. coli isolates harbored the astA gene. Low resistance rates (<10%) were observed among S. aureus; however, four isolates (5.4%) were classified as MRSA, three of which exhibited multidrug resistance. All E. coli isolates were resistant to ampicillin, and 50% displayed multidrug-resistant phenotypes. These findings suggest that computer devices can act as occasional reservoirs of potentially pathogenic and antimicrobial-resistant bacteria in university environments. Full article

The increasing prevalence of antimicrobial resistance, especially in methicillin-resistant Staphylococcus aureus (MRSA), underscores the need for alternative therapies from natural sources. This study investigated the chemical composition, antibacterial activity, and gene expression modulation of Melaleuca cajuputi essential oils. Gas chromatography–mass spectrometry (GC-MS) identified 91 compounds, with naphthalene (23.90%), guaiol (12.92%), caryophyllene oxide (9.69%), D-limonene 98% (8.59%), and gamma terpinene (7.54%) among the most abundant. In Silico molecular docking against MRSA virulence proteins revealed that alloaromadendrene had the strongest binding to toxic shock syndrome toxin-1 (TSST-1) (−7.948 kcal/mol), suggesting high inhibitory potential, while cyclohexane showed weak binding with staphylococcal enterotoxin A (SEA) (−3.532 kcal/mol). Antibacterial assays demonstrated concentration-dependent inhibition, with the zones ranging from 6.33 ± 0.33 mm to 16.67 ± 0.88 mm. MIC and MBC values ranged from 1.56 to 12.5% and 3.13 to 25%, respectively, with most isolates showing bactericidal effects (MBC/MIC ≤ 2). Gene expression analysis of MRSA isolate 4 indicated that sea was moderately upregulated (FC = 1.44), while sec remained unchanged (FC = 1.02). In contrast, fnbA (FC = 0.72), seb (FC = 0.33), and mecA (FC = 0.23) genes were downregulated, and the tsst-1 gene (FC = 0.05) was nearly silent. These findings highlight M. cajuputi essential oils as a promising candidate with both antibacterial efficacy and regulatory effects on MRSA virulence genes. Full article

Colonial cheese production represents a valuable cultural and economic activity in southern Brazil. This study evaluated the effect of oral supplementation of dairy cows with an essential oil blend (EOB)—a combination of eucalyptus oil, peppermint oil, and menthol crystals—on the chemical composition and quality parameters of Colonial cheese during 21 days of ripening. Nine dairy cows were randomly assigned to three groups: control, EOB3.6 (3.6 g/cow/day), and EOB7.2 (7.2 g/cow/day). Milk from each treatment was used to produce Colonial cheeses, which were analyzed for physicochemical composition, texture, color, lipid profile, thiobarbituric acid reactive substances (TBARS), and microbiological quality at different ripening stages. Data were analyzed by analysis of variance (ANOVA) using SAS^® software, following verification of normality and homogeneity of variances. When assumptions were met, repeated-measures ANOVA was applied, and means were compared using Tukey’s test (p < 0.05). Sensory data were evaluated by ANOVA using XLSTAT^® (Addinsoft, Paris, France). EOB supplementation maintained the physicochemical integrity of the cheeses, with a gradual increase in fat content during maturation (40 g/100 g at 21 days, p < 0.05). At seven days, the EOB7.2 treatment showed lower lipid oxidation (TBARS = 0.063, p < 0.05), indicating antioxidant potential. Significant interactions between treatment and maturation were observed for color parameters and polyunsaturated fatty acids (PUFA) (p < 0.05). Cheeses from EOB7.2 presented higher saturated fatty acids (SFA) and lower unsaturated fatty acids (UFA) compared with the control (p < 0.05). No Salmonella spp. or Staphylococcal enterotoxins were detected. Counts of coagulase-positive Staphylococcus, molds, and yeasts remained stable, while Escherichia coli counts were lower in EOB-supplemented cheeses throughout ripening. Overall, EOB supplementation improved oxidative stability and microbiological safety without compromising the technological or compositional quality of Colonial cheese. Full article

Platelet concentrates (PCs) are used to treat patients with platelet deficiencies. PCs are stored at 20–24 °C under agitation for up to 7 days to maintain platelet functionality, but these conditions are amenable for proliferation of contaminants such as Staphylococcus aureus, posing a risk for transfusion-transmitted infections. We investigated the contribution of staphylococcal enterotoxins (SEs) type G (SEG) and type H (SEH) to platelet activation, cytokine release, microRNA (miRNA) modulation, and in vivo virulence. PCs were inoculated with wildtype S. aureus CBS2016-05 or SE-deficient mutants (Δseg, Δseh, ΔΔsegh) and monitored during storage. Flow cytometry revealed progressive elevation of platelet activation markers CD62P and Annexin V in contaminated PCs, with significantly higher expression in wildtype compared to SE-mutant strains. Cytokine profiling demonstrated that SEs modulate pro- and anti-inflammatory mediators, notably CCL2, TGF-β1, IFN-γ, and TNF-α, implicating SEG in their regulation. Next-generation sequencing and RT-qPCR validation identified transient induction of immune-related microRNAs miR-98-5p, miR-146a-5p, miR-221-3p, miR-320a-3p, with SE-dependent expression patterns. In a silkworm infection model, wildtype S. aureus-contaminated PCs exhibited significantly higher lethality than SE-deficient strains, confirming toxin-mediated virulence. Collectively, these findings reveal that SEs exacerbate platelet activation and immune dysregulation during storage, enhancing bacterial pathogenicity. This study identifies platelet-derived cytokine and miRNA signatures as potential biomarkers of bacterial contamination and underscores the need to mitigate SE-driven platelet dysfunction to improve transfusion safety. Full article

Staphylococcus aureus is one of the most important bacterial pathogens affecting both humans and animals. This review discusses the most significant factors that contribute to the pathogenicity of these bacteria and the mechanisms that regulate their expression. We also focus on the factors that play a role in the pathogenesis of skin diseases. S. aureus possesses a wide array of virulence factors that allow it to bypass passive and active mechanisms of the host’s immune system and effectively infect and spread within the infected organism. These include factors that facilitate colonization of the skin (i.e., arginine catabolic mobile element-ACME), mucous membranes and other surfaces, proteins protecting the bacteria from the host’s immune system, superantigens and superantigen-like proteins, surface proteins that promote adhesion and biofilm formation, toxins, enzymes, and iron uptake systems. Additionally, a complex network of regulatory systems (accessory gene regulator -Agr, (staphylococcal accessory regulator -Sar, S. aureus exoprotein expression -Sae, and others) controls the expression of virulence genes at the transcriptional and translational levels. The activity of these regulatory systems is pivotal in determining whether S. aureus initiates an invasive infection, characterized by toxin and enzyme production (e.g., hemolysin alpha -Hla, phenol soluble modulins -PSM, toxic shock syndrome -TSST-1, enterotoxins, Panton-Valentine leukotoxin- PVL). This is indicative of community-acquired S. aureus (CA-Sa, CA-MRSA, CA-MSSA), or a chronic infection, characterized by surface protein expression and biofilm formation, which is indicative of hospital-acquired or healthcare-acquired S. aureus (HA-Sa, HA-MRSA, HA-MSSA). Full article

Staphylococcal enterotoxins (SEs), particularly enterotoxin C (SEC), are potent superantigens primarily known for causing food poisoning, but recent studies have highlighted their potential role in immune-mediated intestinal diseases. Despite the widespread use of food preservatives, their influence on SEC production—especially from coagulase-negative staphylococci (CNS)—remains poorly understood. In this study, we evaluated the effects of commonly used preservatives, including sodium chloride, potassium nitrate, and sorbic acid, on the expression and production of SEC₃ and SEC_epi in Staphylococcus aureus and S. epidermidis, respectively. Using ELISA and RT-qPCR, we analyzed toxin levels at both the protein and mRNA levels. Proliferation assays on human PBMCs assessed the mitogenic potential of culture supernatants. While sodium chloride and potassium nitrate did not significantly alter SEC levels or bacterial growth, only sorbic acid at 0.07% consistently inhibited both mRNA expression and protein production of SEC₃ and SEC_epi. Furthermore, supernatants from sorbic acid-treated cultures induced significantly lower PBMC proliferation. These results suggest that even sub-emetic concentrations of enterotoxins may have immunomodulatory effects, and sorbic acid could be a promising agent in mitigating such risks. Full article

Staphylococcus spp. present a dual role in cheese production as some species are pathogenic, while others bring beneficial characteristics. Coagulase-positive staphylococci (CoPS), particularly Staphylococcus aureus, are of concern due to their ability to produce enterotoxins linked to foodborne outbreaks. These toxins, encoded by staphylococcal enterotoxin (SE) genes, cause gastroenteritis, especially vomiting. Many members of the genus harbor a plethora of virulence genes and are able to form biofilms. The prevalence of antibiotic-resistant strains, including methicillin-resistant S. aureus (MRSA), complicates control. In contrast, some members of the coagulase-negative staphylococci (CoNS) group, such as Staphylococcus carnosus, Staphylococcus condimenti, Staphylococcus equorum, Staphylococcus piscifermentans, Staphylococcus succinus, and Staphylococcus xylosus, contribute to ripening, influencing flavor and texture. Some are even considered safe and studied for their ability to inhibit pathogens. Expression of enterotoxin genes in Staphylococcus, particularly S. aureus, is influenced by environmental factors and can be regulated by different mechanisms including quorum sensing. Understanding gene expression in conditions found during cheese production and ripening can help in formulating effective interventions. Risks posed by enterotoxin-producing Staphylococcus in cheese are evident, with numerous outbreaks reported worldwide. Moreover, several species present risks to both animal and human health. Effective control measures include adherence to microbiological criteria in foods, animal health monitoring, good manufacturing practices (GMP), temperature control, proper ripening conditions and hygiene. This review compiles and discusses existing knowledge on CoPS and CoNS in cheeses, providing a framework for evaluating their risks and benefits and guiding future studies in cheese microbiology. Full article

Staphylococcal food poisoning (SFP) is a common foodborne illness caused by the ingestion of enterotoxins produced by Staphylococcus aureus, posing a persistent global public health concern. Although regional differences in implicated food types and predominant enterotoxins have been reported, the underlying factors remain unclear. In this study, we systematically investigated the effects of nutritional factors on the growth, biofilm formation, and production of two representative enterotoxins, SEA and SEB, by S. aureus. Specifically, we evaluated bacterial responses to different concentrations of NaCl, glucose, and tryptone. NaCl suppressed growth, biofilm formation and enterotoxin production in a dose-dependent manner. Glucose markedly inhibited both bacteria growth and enterotoxin production, with a stronger effect on SEB than SEA. In contrast, tryptone promoted bacterial growth and moderately enhanced biofilm formation but did not significantly affect enterotoxin production. Importantly, even under comparable bacterial counts, the types and amounts of SEs produced varied substantially depending on the nutrient composition. These findings provide new insights into the nutrient-dependent regulation of virulence in S. aureus and highlight the importance of considering environmental and nutritional factors when assessing risks of SFP and designing effective food safety strategies. Full article

Staphylococcus aureus is a major pathogen responsible for staphylococcal food poisoning (SFP), with its pathogenicity primarily dependent on staphylococcal enterotoxins (SEs). Among these, staphylococcal enterotoxin A (SEA) is a critical risk factor due to its high toxicity, high detection rate (accounting for 80% of SFP cases), strong thermal stability, and resistance to hydrolysis. Traditional SEA immunoassays, such as enzyme-linked immunosorbent assay (ELISA), are prone to false-positive results caused by nonspecific binding interference from S. aureus surface protein A (SpA). In recent years, nanobodies (single-domain heavy-chain antibodies) have emerged as an ideal alternative to address SpA interference owing to their small molecular weight (15 kDa), high affinity, robust stability, and lack of Fc regions. In this study, based on a previously developed highly specific monoclonal antibody against SEA (mAb-4C6), four anti-SEA nanobodies paired with mAb-4C6 were obtained through two-part (four-round) of biopanning from a naive nanobody phage display library. Among these, SEA-4-20 and SEA-4-31 were selected as optimal candidates and paired with mAb-4C6 to construct double-antibody sandwich ELISAs. The detection limits for SEA were 0.135 ng/mL and 0.137 ng/mL, respectively, with effective elimination of SpA interference. This approach provides a reliable tool for rapid and accurate detection of SEA in food, clinical, and environmental samples. Full article