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BACKGROUND: Coagulase-negative staphylococci (CoNS) are evolving as major reservoirs and vectors of unusual and critical antimicrobial resistance (AMR) mechanisms. MATERIALS AND METHODS: In this study, the genomic characterization of 26 multidrug-resistant (MDR)-CoNS (S. borealis, S. saprophyticus, S. sciuri, S. hominis, S. epidermidis, S. pasteuri, S. hyicus, S. simulans, S. haemolyticus, and S. arlettae) previously obtained from the nasal cavity of healthy nestling storks, humans who had no contact with animals, pigs, and pig farmers, as well as dogs and dog owners from Spain was performed. High-quality draft genomes obtained by Illumina sequencing technology were used to determine their resistome, virulome, mobile genetic elements, and CRISPR-Cas types. The relatedness of three CoNS species with publicly available genomes was assessed by core-genome single nucleotide polymorphisms (SNPs). RESULTS: AMR genes to all classes of antibiotics in staphylococci were detected including unusual ones (mecC, ermT, and cfr), of which their corresponding genetic organizations were analyzed. About 96.1% of the MDR-CoNS strains harbored diverse adherence or immune evasion genes. Remarkably, one enterotoxin-C and -L-carrying S. epidermidis-ST595 strain from a nestling stork was detected. Moreover, various plasmid bound-biocide resistance genes (qacACGJ) were identified in 34.6% of the MDR-CoNS. Two genes that encode for cadmium and zinc resistance (cadD, czrC) were found, of which czrC predominated (42.3%). Complete CRISPR-Cas system was detected in 19.2% of the CoNS strains, of which cas-1, -2, and -9 predominated, especially in 75% of the S. borealis strains. The phylogenetic analysis identified clusters of related S. epidermidis lineages with those of other countries (SNP < 100). Also, highly related S. borealis isolates (SNP < 10) from pigs was confirmed for the first time in Spain. CONCLUSION: These findings showed that various ecological niches harbor CoNS that presented MDR phenotypes mediated by multiple AMR genes carried by mobile genetic elements with relatively low frequency of intact CRISPR-Cas systems. Furthermore, the transmission of some CoNS species in humans and animals is strongly suggested.

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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.

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Diagnosis of allergic disease is primarily verified by IgE blood serum analysis. Determination in nasal secretions is technically more difficult, particularly due to a low specimen volume and the method of sample collection. Nasal secretions are frequently collected by lavage, which allows qualitative diagnostics, whereas swabs with defined amounts of mucus enable quantitative analyses. In the case of negative skin and serum tests, detection of IgE in nasal mucus combined with nasal provocation testing aids differentiation between local allergic and nonallergic rhinitis.

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During surveillance of Staphylococcus aureus in lesions from patients with atopic dermatitis (AD), we isolated Staphylococcus argenteus, a species registered in 2011 as a new member of the genus Staphylococcus and previously considered a lineage of S. aureus. Genome sequence comparisons between S. argenteus isolates and representative S. aureus clinical isolates from various origins revealed that the S. argenteus genome from AD patients closely resembles that of S. aureus causing skin infections. We previously reported that 17%-22% of S. aureus isolated from skin infections produce staphylococcal enterotoxin Y (SEY), which predominantly induces T-cell proliferation via the T-cell receptor (TCR) Vα pathway. Complete genome sequencing of S. argenteus isolates revealed a gene encoding a protein similar to superantigen SEY, designated as SargEY, on its chromosome. Population structure analysis of S. argenteus revealed that these isolates are ST2250 lineage, which was the only lineage positive for the SEY-like gene among S. argenteus. Recombinant SargEY demonstrated immunological cross-reactivity with anti-SEY serum. SargEY could induce proliferation of human CD4+ and CD8+ T cells, as well as production of TNF-α and IFN-γ. SargEY showed emetic activity in a marmoset monkey model. SargEY and SET (a phylogenetically close but uncharacterized SE) revealed their dependency on TCR Vα in inducing human T-cell proliferation. Additionally, TCR sequencing revealed other previously undescribed Vα repertoires induced by SEH. SargEY and SEY may play roles in exacerbating the respective toxin-producing strains in AD. IMPORTANCE: Staphylococcus aureus is frequently isolated from active lesions of atopic dermatitis (AD) patients. We reported that 17%-22% of S. aureus isolated from AD patients produced a novel superantigen staphylococcal enterotoxin Y (SEY). Unlike many S. aureus superantigens that activate T cells via T-cell receptor (TCR) Vß, SEY activates T cells via TCR Vα and stimulates cytokine secretion. Staphylococcus argenteus was isolated from AD patients during the surveillance for S. aureus. Phylogenetic comparison of the genome indicated that the isolate was very similar to S. aureus causing skin infections. The isolate encoded a SEY-like protein, designated SargEY, which, like SEY, activated T cells via the TCR Vα. ST2250 is the only lineage positive for SargEY gene. ST2250 S. argenteus harboring a superantigen SargEY gene may be a novel staphylococcal clone that infects human skin and is involved in the exacerbation of AD.

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Notification of foodborne outbreaks has been mandatory in Europe since 2005, and surveillance is carried out along the entire food chain. Here we report the results obtained from laboratory investigations about four cases of foodborne outbreaks that occurred in Sicily between 2009 and 2016, deemed to be related to staphylococcal enterotoxins (SEs) and coagulase-positive Staphylococci (CPS) by the Local Public Health Authority. Primosale cheese samples were processed by culture methods for enumeration of CPS and immunoenzymatic assays for detection and differentiation of the SEs possibly contained in food samples. In all cases, the mistrusted foods were found to be contaminated by CPS at bacterial loads between 5 and 8 log CFU/g and contained SE type C (SEC). The reported data confirm the risk of staphylococcal food poisoning associated with the consumption of raw milk cheese. SEC is the most commonly occurring SE in goat milk and dairy products and the most represented enterotoxin in Sicilian dairy products. Our results highlighted the need for improving the current monitoring efficiency and implementing the available laboratory methods to collect more faithful epidemiological data on the current prevalence of staphylococcal toxins in the food chain, including SEs currently not detectable by validated analytical methods.

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Staphylococcus aureus is one of the primary pathogenic agents found in cheeses produced with raw milk. Some strains of S. aureus are enterotoxigenic, possessing the ability to produce toxins responsible for staphylococcal food poisoning when present in contaminated foods. This study aimed to genotypically characterize, assess the antimicrobial resistance profile, and examine the enterotoxigenic potential of strains of S. aureus isolated from artisanal colonial cheese. Additionally, a bacterial diversity assessment in the cheeses was conducted by sequencing the 16S rRNA gene. The metataxomic profile revealed the presence of 68 distinct species in the cheese samples. Fifty-seven isolates of S. aureus were identified, with highlighted resistance to penicillin in 33% of the isolates, followed by clindamycin (28%), erythromycin (26%), and tetracycline (23%). The evaluated strains also exhibited inducible resistance to clindamycin, with nine isolates considered multidrug-resistant (MDR). The agr type I was the most prevalent (62%) among the isolates, followed by agr type II (24%). Additionally, ten spa types were identified. Although no enterotoxins and their associated genes were detected in the samples and isolates, respectively, the Panton-Valentine leukocidin gene (lukS-lukF) was found in 39% of the isolates. The presence of MDR pathogens in the artisanal raw milk cheese production chain underscores the need for quality management to prevent the contamination and dissemination of S. aureus strains.

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Background: Food safety is a serious challenge in the face of increasing population and diminishing resources. Staphylococcus aureus is a critical foodborne pathogen characterized by its capability to secret a diverse range of heat-resistant enterotoxins. Antibiotic usage in dairy herds resulted in the occurrence of antimicrobial resistance (AMR) patterns among bacterial species, which were consequently transmitted to humans via dairy products. Lactic acid bacteria (LAB) produce bacteriocins, which provide an excellent source of natural antimicrobials with the further advantage of being environmentally friendly and safe. Aim: Detection of multidrug resistance (MDR) S. aureus isolates in concerned samples, molecular characteristics, biofilm production, and the inhibitory role of LAB against it. Methods: Random samples of raw milk and other dairy products were analyzed for S. aureus isolation. Phenotypic and genotypic assessment of AMR was performed, in addition to detection of classical enterotoxin genes of S. aureus. Finally, evaluation of the antimicrobial action of some Lactobacillus strains against S. aureus. Results: Incidence rates of presumptive S. aureus in raw milk, Kariesh cheese, and yogurt samples were 50%, 40%, and 60%, respectively. The highest resistance of S. aureus was to Kanamycin (100%) and Nalidixic acid (89.3%), respectively. (78.66%) of S. aureus were MDR. 11.1% of S. aureus carried mecA gene. In concern with enterotoxins genes, PCR showed that examined isolates harbored sea with a percentage of (22.2%), while sed was found in (11.1%) of isolates. Regarding biofilm production, (88.88%) of S. aureus were biofilm producers. Finally, agar well diffusion showed that Lactobacillus acidophilus had the strongest antimicrobial action against S. aureus with inhibition zone diameter ranging from 18 to 22 mm. Conclusion: There is a widespread prevalence of MDR S. aureus in raw milk and dairy products. Production of staphylococcal enterotoxins, as well as biofilm production are responsible for public health risks. Therefore, installing proper hygienic routines and harsh food safety policies at food chain levels is substantial.

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INTRODUCTION: Raw milk is a nutrient-rich food, but it may harbour harmful bacteria, such as enterotoxigenic Staphylococcus aureus (S. aureus), which can cause staphylococcal food poisoning. Antibiotic resistance of S. aureus in raw milk can increase the risk of such infections, particularly among susceptible individuals. OBJECTIVE: This study aimed to investigate the prevalence of enterotoxin genes a, d, g, i and j and the antibiotic resistance of S. aureus isolated from raw milk samples. METHODS: During a 6-month sampling period, 60 raw milk specimens were obtained from diverse locations in Yazd province, Iran. Antibiogram profiling was conducted via the disc diffusion method. In addition, staphylococcal enterotoxin (SE) genes a, d, g, i, and j were detected through real-time PCR analysis. RESULTS: Bacteriological assays confirmed the presence of S. aureus in 11 samples (18.3%). All isolates demonstrated 100% resistance to penicillin G but exhibited sensitivity to vancomycin, while resistance to other antibiotics ranged from 36.4% to 45.5%. The prevalence of enterotoxin genes in these strains showed variable distribution, with sea being the predominant SE (45.5%), followed by sed (36.4%), seg (18.2), sej and sei (9.1% each). CONCLUSIONS: This study discovered the presence of multiple enterotoxins in S. aureus strains obtained from raw milk samples. These strains also demonstrated resistance to a variety of antibiotics. Since enterotoxigenic S. aureus is known to cause human food poisoning, monitoring food hygiene practices, especially during raw milk production, is critical.

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Retail stores maintain fresh rice noodles (FRNs) at room temperature because refrigeration negatively impacts FRNs' texture. The room temperature and high water activity of FRNs help spore-forming Bacillus cereus to grow and produce toxins. In this study, the effect of steam cooking on survival and different storage temperatures on the growth and enterotoxins production of B. cereus in FRNs were investigated. White rice flour was used to make FRNs. Three treatments of FRNs were used in this study; uninoculated, inoculated (with 4.0 log CFU/ml of B. cereus spores), and autoclaved as a negative control. A slurry of rice flour, cornstarch, and water was steam cooked for 4 min at 90°C and incubated for 168 h at 4°C, and for 72 h at 22 and 32°C. Incubated FRNs were tested for pH, B. cereus growth, and enterotoxins production. Steam cooking reduced the total number of B. cereus spores by 0.7 ± 0.3 log CFU/g. Surviving B. cereus spores in inoculated and uninoculated FRNs germinated over 72 h of storage. No B. cereus was detected in negative controls. An interaction was observed across storage temperatures and time (p < 0.05). The B. cereus population in uninoculated FRNs increased by more than 7.0 log CFU/g at 22 and 32°C over 72 h, while inoculated FRNs showed a 5.0 log bacterial increase at these storage temperatures. No growth was observed at 4°C in both inoculated and uninoculated FRNs. The pH of inoculated FRNs was reduced from 6.9 ± 0.1 to 5.7 ± 0.0 at 32°C and to 6.2 ± 0.1 at 22°C, and the pH of uninoculated FRNs was reduced from 7.0 ± 0.1 to 5.8 ± 0.2 at 32°C and to 6.5 ± 0.0 at 22°C, indicative of FRNs spoilage. B. cereus in inoculated FRNs produced enterotoxins after 12 h of storage at 32°C, and over 24 h of storage at 22°C, while no toxin was detected at 4°C. Our findings show that storing FRNs at room temperature for 24 h leads to enterotoxin production, emphasizing the importance of proper FRN storage and their potential risk to consumers. Nevertheless, further research should investigate the effect of other foodborne pathogens on these products.

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The increasing global popularity of ready-to-eat (RTE) foods for their convenience simultaneously brings along a risk, as these products can be contaminated with various microorganisms, including potentially harmful pathogens. We aimed to investigate the food contamination of Staphylococcus aureus (S. aureus) in RTE foods in Guangdong, South China. All S. aureus isolates were subjected to characterization through antimicrobial susceptibility tests, multilocus sequence typing (MLST), and PCR analysis for detecting mec and blaZ genes. A total of 824 RTE food samples were collected from 2017 to 2022, of which 73 (8.9%) were found to be contaminated with S. aureus. Contamination levels were mostly in the range of 0.3-1.0 most probable number (MPN)/g, with 10 samples exceeding 110 MPN/g. Of the 73 S. aureus isolates, 10 were identified as methicillin-resistant S. aureus (MRSA). In MRSA, resistance was most frequently observed to penicillin (100%, 10/10), followed by erythromycin (80.0%, 8/10) and tetracycline (70%, 7/10). And in methicillin-sensitive S. aureus (MSSA), resistance was most frequently observed to penicillin (98.4%, 62/63), followed by tetracycline (30.2%, 19/63) and erythromycin (23.8%, 15/63). Overall, 98.6% (72/73) of the isolates demonstrated resistance to at least one antimicrobial agent, whereas 31.5% (23/73) were resistant to three or more antimicrobials. Fifty-seven S. aureus isolates harbored the penicillin-resistant gene blaZ, and 10 isolates carried the mec gene. In addition, 30.1% of the isolates harbored genes for classical staphylococcal enterotoxins (SEs), with seb being the most frequently detected SE gene. MLST revealed that the 73 isolates belonged to 14 different sequence types (STs), the most prevalent of which was ST7. In MRSA, the most common prevalent clone is ST6, and in MSSA, ST7 was the most common isolates. The prevalent multidrug resistance indicates that the resistance situation of foodborne S. aureus in Guangdong is severe, posing a potential threat to consumer safety and health.

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Staphylococcal enterotoxins (SEs), the major virulence factors of Staphylococcus aureus, cause a wide range of food poisoning and seriously threaten human health by infiltrating the food supply chain at different phases of manufacture, processes, distribution, and market. The significant prevalence of Staphylococcus aureus calls for efficient, fast, and sensitive methods for the early detection of SEs. Here, we provide a comprehensive review of the hazards of SEs in contaminated food, the characteristic and worldwide regulations of SEs, and various detection methods for SEs with extensive comparison and discussion of benefits and drawbacks, mainly including biological detection, genetic detection, and mass spectrometry detection and biosensors. We highlight the biosensors for the screening purpose of SEs, which are classified according to different recognition elements such as antibodies, aptamers, molecularly imprinted polymers, T-cell receptors, and transducers such as optical, electrochemical, and piezoelectric biosensors. We analyzed challenges of biosensors for the monitoring of SEs and conclude the trends for the development of novel biosensors should pay attention to improve samples pretreatment efficiency, employ innovative nanomaterials, and develop portable instruments. This review provides new information and insightful commentary, important to the development and innovation of further detection methods for SEs in food samples.

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This study aimed to identify coagulase-positive staphylococci (CPS) species from 21 samples of clandestine Minas Frescal cheese, investigate the potential for deterioration in psychrotrophic and mesophilic conditions, verify the toxigenic potential of Staphylococcus aureus, and determine the antimicrobial susceptibility profile of toxigenic S. aureus. Species determination was performed based on the detection of ß-hemolysis in 5% ovine blood agar; fermentation of mannitol, maltose, and trehalose sugars; and production of acetoin. After species determination, DNA extraction and analysis was performed for S. aureus colonies for genes encoding staphylococcal toxins (eta, etb, tst, sea, seb, sec, sed, and see) using 2 multiplex PCR assays. Isolates identified as toxigenic S. aureus were tested for antimicrobial susceptibility to tetracycline, erythromycin, clindamycin, gentamicin, ciprofloxacin, sulfazotrim, trimethoprim, streptomycin, cefoxitin, vancomycin and enrofloxacin. Elevated CPS counts were observed with an average of >6 log cfu/g. Of the 355 isolates, 177 (49.86%) were identified as S. aureus. Staphylococcus hyicus, Staphylococcus intermedius, Staphylococcus delphini, and Staphylococcus coagulans were identified in 3 (0.84%), 2 (0.56%), 2 (0.56%), and 1 (0.28%) isolates, respectively. Of the total number of S. aureus, 25 (52.08%) were positive for the gene that encodes for toxic shock toxin (TSST-1). Another 16 (33.33%) were positive for the sea gene, and 4 isolates (8.33%) were positive for see and one isolate each was positive for seb (2.08%), sec (2.08%), and etb (2.08%) genes. All isolates demonstrated lipolytic activity under mesophilic and psychrotrophic conditions. S. intermedius and S. hyicus had the most prominent proteolytic potential. Multidrug resistance was observed in most of the potentially toxigenic isolates, with clindamycin having the lowest efficiency (40%), whereas the aminoglycosides (gentamicin and streptomycin) had the highest effectiveness demonstrating inhibition in all evaluated isolates. Methicillin-resistant S. aureus (MRSA) was detected. Minas Frescal cheeses, marketed in the north of Tocantins in the Brazilian Amazon region, do not comply with legal quality standards and pose a public health risk due to the enterotoxigenic potential of multiresistant isolates, in addition to low shelf life of the samples given the high spoilage potential of this microbiota.

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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.

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BACKGROUND: Staphylococcus aureus is part of normal flora and also an opportunistic pathogen responsible for a wide range of infections in both humans and animals. Livestock-associated S. aureus (LA-SA) has gained importance in recent years due to its increased prevalence in recent years, becoming a worry in public health view. This study aimed to study the epidemiology of LA-SA strains in Madurai district, Tamil Nadu, India. METHODS: A total of 255 samples were collected from bovine and other small ruminants like goats and sheep nares (n = 129 and n = 126 respectively). Nasal swab samples were collected from study animals with sterile sample collecting cotton swabs (Hi-Media, Mumbai). Samples were transported to the lab in Cary-Blair Transport media for further analysis. The samples were tested for S. aureus using antibiotic selection and PCR-based assays. The pathogenicity of the bacteria was assessed using chicken embryo models and liver cross-sections were used for histopathology studies. RESULTS: The prevalence rate in bovine-associated samples was 42.63% but relatively low in the case of small ruminants associated samples with 28.57% only. The overall prevalence of S. aureus is found to 35.6% and MRSA 10.98% among the study samples. The antibiogram results that LA-SA isolates were susceptible to aminoglycosides and tetracyclines but resistant to ß-lactam drugs. The biofilm formation results showed that the LA-SA isolates are weak to high-capacity biofilm formers. The enterotoxigenic patterns revealed that most of the isolated strains are enterotoxigenic and possess classical enterotoxins. The survival analysis of chicken embryos suggested that the Bovine-associated strains were moderately pathogenic. CONCLUSION: The study concluded that economically important livestock animals can act as reservoirs for multi-drug resistant and pathogenic which in-turn is a concern for public health as well as livestock health.

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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.

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Bacillus cereus is estimated to be responsible for 1.4-12% of all food poisoning outbreaks worldwide. The objective of this study was to investigate the toxigenic potential of 181 isolates of B. cereus previously recovered from different types of berries and berry products (strawberries, raspberries, blackberries, and blueberries) by assessing the presence of enterotoxin genes (hblA, hblC, hblD, nheA, nheB, nheC, and cytK) and an emetic toxin cereulide synthetase gene (ces). The cytotoxic activity on Caco-2 cells was also evaluated for the two isolates containing the gene cytK. Twenty-three toxigenic profiles were found. The nheABC (91.7%) and hblACD (89.0%) complexes were the most prevalent among the isolates, while the cytK and ces genes were detected in low percentages, 1.1% and 3.3%, respectively. In addition, the nheABC/hblACD complex and ces genes were detected in isolates recovered throughout the production process of blackberries and strawberries. The cytotoxic activity on Caco-2 cells was also observed to be greater than 60% for isolates containing the cytK gene.

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How foodborne enterotoxigenic Bacillus cereus rewires energy metabolism during intestinal tract infection is still not understood. In this study, we used the Seahorse XFe technology to simultaneously analyze oxygen consumption and acidification rates to estimate bioenergetic changes in the intestinal Caco-2 cell line after exposure to the B. cereus sensu lato (s.l.) enterotoxin-producing pathotypes, American Type Culture Collection (ATCC) 14579 (836), NVH0391-98 (828), and NVH0075/95 (825). Infection of Caco-2 led to a more energetic phenotype due to increased flux through oxidative phosphorylation and glycolysis. Strain 836 caused the most pronounced effects toward the specific energy phenotype, followed by strains 828 and 825. However, the metabolic potential of Caco-2 cells was most strongly induced by the 828 strain. Furthermore, infected cells manifested an increased adenosine triphosphate (ATP) production rate. Strain 828 caused the highest glycolytic and mitochondrial ATP production rates, followed by the 836 and 825 B. cereus s.l. strains. The glycolytic stress assay showed that strains 828 and 826 slightly increased compensatory glycolysis, providing a better understanding of the pathogenicity of this versatile pathogen. The results of this study underline that extracellular flux measurement can be used to accurately estimate bioenergetic perturbations of Caco-2 cells as a consequence of infection. Our findings enhance our understanding of how intestinal cells adjust their metabolism during infection with B. cereus s.l.

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BACKGROUND: Staphylococcus aureus expresses numerous toxins, many of which are strongly believed to be responsible for specific symptoms and even diseases, making it significant in the pathogenesis of human health. Enterotoxins, which are vital toxins, are associated with foodborne illnesses that manifest through symptoms like vomiting and diarrhea. In the present study, 264 S. aureus isolates obtained from various retail foods in Hangzhou, China were further investigated the profiles of enterotoxin genes and genetic backgrounds. RESULTS: Approximately, 64.02% of the isolates from diverse sources contained at least one Staphylococcal Enterotoxin (SE) genes, displaying a total of 36 distinct combinations. Enterotoxin gene cluster (egc) encoded enterotoxin genes, normally designated by seg, sei, sem, sen, seo and selu, plus with sep were more frequently detected (33.73%, each). In contrast, see, ses and set were absent in any of the isolates tested. A total of 44 sequence types (STs), 20 clonal complexes (CCs) and 66 different staphylococcal protein A (spa) types (including six novel types) were identified among those 169 SE-positive isolates. Moreover, nineteen methicillin-resistant Staphylococcus aureus (MRSA) isolates were identified. The majority of those isolates belonged to the CC59-Sccmec IVa cluster and carried the seb-sek-seq gene cluster. The egc cluster, either coexisting with or without other enterotoxin genes, was observed in all isolates allocated into CC5, CC9, CC20, CC25, CC72 and ST672. Irrespective of the spa types and origins of the food, it appeared that seh was a distinct genetic element present in isolates belonging to the CC1 clonal lineage. CONCLUSIONS: The results not only proposed a suspected relationship between distribution of enterotoxigenic strains and genetic backgrounds, but also attributed the presence of novel enterotoxins to potential hazards in food safety.

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Abstract Biofilm formation by Bacillus cereus strains is now recognized as a systematic contaminaron mechanism in foods; the aim of this study was to evaluate the production of submerged and interface biofilms in strains of B. cereus group in different materials, the effect of dex-trose, motility, the presence of genes related to biofilms and the enterotoxigenic profile of the strains. We determine biofilm production by safranin assay, motility on semi-solid medium, toxin gene profiling and genes related to biofilm production by PCR in B. cereus group iso-lated from food. In this study, we observe strains used a higher production of biofilms in PVC; in the BHI broth, no submerged biofilms were found compared to phenol red broth and phenol red broth supplemented with dextrose; no strains with the ces gene were found, the enterotoxin profile was the most common the profile that includes genes for the three enterotoxins. We observed a different distribution of tasA and sipW with the origin of isolation of the strain, being more frequent in the strains isolated from eggshell. The production and type of biofilms are differential according to the type of material and culture medium used.

Resumen La formación de biopelículas por cepas de Bacillus cereus es reconocida como un mecanismo de contaminación sistemática en alimentos; el objetivo del estudio fue evaluar la producción de biopelículas sumergidas y de superficie en cepas del grupo de Bacillus cereus en diferentes materiales, el efecto de la dextrosa, la motilidad, la presencia de genes relacionados a biopelículas y el perfil enterotoxigénico de las cepas. Determinamos la producción de biopelículas por el ensayo de safranina, motilidad en medio sólido, perfil enterotoxigénico y genes relacionados a producción de biopelículas por PCR en aislados del grupo de Bacillus cereus de alimentos. En este estudio, observamos en las cepas utilizadas una alta producción de biopelículas en PVC; en caldo BHI, no se encontraron biopelículas sumergidas en comparación con el caldo rojo de fenol y caldo rojo de fenol suplementando con dextrosa; no se encontraron cepas con el gen ces, el perfil de enterotoxinas más común fue el perfil que incluía los genes de las tres enterotoxinas, también observamos una distribución diferente de tasA y sipW con relación al origen de la cepa, siendo más frecuente estos genes en las cepas aisladas de huevos. La producción y el tipo de biopelículas es diferente de acuerdo con el tipo de material y el medio de cultivo utilizado.

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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.