Occurrence of Antibiotic-Resistant Bacteria in Fish and Seafood from Slovak Market.

The consumption of sushi or poke has grown globally. However, this type of dish often contains raw fish or seafood; therefore, it can pose a microbial risk for consumers. This study deals with the occurrence of total and antibiotic-resistant coliform bacteria and enterococci in fish and seafood as well as sushi and poke bought from Slovak retail (restaurants and fast food). Total coliforms have ranged in sushi, poke samples and samples of fish and seafood from cooling counters from 0.6 to 5.1 log CFU/g. Ampicillin resistance has been predominantly observed in all types of samples. Tetracycline resistance was detected in 16% of all tested samples and gentamicin resistance in 13%. Total enterococci has been detected in 74% of sushi samples, 100% of poke samples and 62% of samples obtained from supermarkets. The majority of enterococci were resistant to ampicillin. Vancomycin resistance was observed in five samples. Forty-eight resistant coliforms were identified mainly as Enterobacter spp. and Klebsiella spp. Antibiotic-resistant isolates were predominantly resistant to gentamicin, chloramphenicol and tetracycline. In 13% of resistant isolates was detected efflux pumps overproduction, and in four isolates was detected the tetA resistance gene. Our results point to poor hygiene in some establishments. The prevention of the antibiotic-resistant bacteria spread would be in better stewardship and improved monitoring of sanitation.

Monophasic Variant of Salmonella Typhimurium 4,[5],12:i:- (ACSSuGmTmpSxt Type) Outbreak in Central Italy Linked to the Consumption of a Roasted Pork Product (Porchetta).

The monophasic variant of S. Typhimurium 4,[5],12:i:- (MVST) is the third most commonly reported Salmonella serovar involved in human infections (8.8%) in the EU and ranks after S. Enteritidis (54.6%) and S. Typhimurium (11.4%). In Italy, in contrast, the MVST has achieved peculiar epidemiological and ecological success which has allowed it to be, since 2011, the serovar most frequently isolated from humans. In the summer of 2022, a foodborne outbreak of the MVST involving 63 people occurred in the Marche Region (Central Italy). A common food exposure source among some human cases was a roasted, ready-to-eat (RTE) pork product, porchetta, which is a typical product of Central Italy. This paper describes the results of investigations conducted to clarify this outbreak. The porchetta was produced by a local manufacturing plant and distributed to at least two local retail stores, one of which was the retail outlet for the manufacturing plant. The MVST was isolated from surface samples collected at the porchetta manufacturing plant and at both local retail stores via bacterial analysis, and the porchetta sampled at one store contained the MVST. These data confirm this type of RTE pork product can be a source of Salmonella infection in humans.

Sterilizing Ready-to-Eat Poached Spicy Pork Slices Using a New Device: Combined Radio Frequency Energy and Superheated Water.

In this study, a new device was used to inactivate G. stearothermophilus spores in ready-to-eat (RTE) poached spicy pork slices (PSPS) applying radio frequency (RF) energy (27.12 MHz, 6 kW) and superheated water (SW) simultaneously. The cold spot in the PSPS sample was determined. The effects of electrode gap and SW temperature on heating rate, spore inactivation, physiochemical properties (water loss, texture, and oxidation), sensory properties, and SEM of samples were investigated. The cold spot lies in the geometric center of the soup. The heating rate increased with increasing electrode gap and hit a peak under 190 mm. Radio frequency combined superheated water (RFSW) sterilization greatly decreased the come-up time (CUT) compared with SW sterilization, and a 5 log reduction in G. stearothermophilus spores was achieved. RFSW sterilization under 170 mm electrode gap reduced the water loss, thermal damage of texture, oxidation, and tissues and cells of the sample, and kept a better sensory evaluation. RFSW sterilization has great potential in solid or semisolid food processing engineering.

Microbiological Safety and Quality of Meals and Work Surfaces in Collective Catering Systems in Central Italy: A Five-Year Monitoring Study.

Ready-to-eat (RTE) meals produced and served by the catering system still represent one of the major causes of foodborne outbreaks, especially for susceptible consumers. Despite the great progress in food hygiene and safety, the systematic monitoring of microbial contamination of foodstuff is the most effective tool to ensure food safety and protect consumers' health. The aim of this study was to perform a thorough assessment of the microbial safety and quality of meals and work surfaces of collective catering systems in central Italy, over a five-year period (2014−2018). In total 11,012 microbiological analytical determinations were performed in food matrices (80.1%) and environmental samples (19.9%). The results obtained show a low level of non-conformities ranging from 2.2% to 6.3% of total samples, concerning both hygiene and safety parameters. A decreasing trend of non-conformities during the years was also highlighted (p-value < 0.05), especially for environmental samples. This study suggests that the implementation of Good Manufacturing Practices (GMPs) and the proper definition of Hazard Analysis and Critical Control Point (HACCP) plans, combined with a thorough evaluation of microbiological monitoring, are able to ensure high levels of food safety and hygiene.

Contamination of ready-to-eat street food in Pakistan with Salmonella spp.: Implications for consumers and food safety.

OBJECTIVES: Ready-to-eat (RTE) food sold in Quetta, Pakistan was assessed for microbial contamination. METHODS: Equal numbers of samples were collected from four categories of RTE food - burgers, shawarma, pizza and sandwiches - from January 2018 to December 2018. Microbial contamination of individual food samples was assessed by quantifying the total aerobic count obtained from plating samples on bacterial growth medium. Salmonella spp. serovars were identified using polymerase chain reaction. RESULTS: Approximately 38% (121/320) of RTE food samples were not fit for human consumption. The most contaminated type of RTE food was shawarma (49%). Microbial contamination of food samples was higher in summer compared with the other seasons. Approximately 40% (49/121) of food samples that were not fit for human consumption were contamined with Salmonella spp. Salmonella enteritidis (69%) and Salmonella typhimurium (31%) were the only serovars among the samples testing positive for Salmonella spp. Of the 49 samples with high microbial counts, S. enteritidis was present in 34 samples and S. typhimurium was present in 15 samples. The antibiotic sensitivity results demonstrated that both S. enteritidis and S. typhimurium were resistant to amoxicillin. In addition, S. enteritidis was resistant to chloramphenicol and erythromycin, and S. typhimurium presented high resistance to erythromycin. Both S. typhimurium and S. enteritidis were highly sensitive to kanamycin. CONCLUSION: RTE food sold by street vendors in Quetta was found to be contaminated with Salmonella spp. and poses a great health risk to consumers. As such, consumption should be avoided, and the health authorities should take stringent action to ensure the quality of street food in order to reduce the healthcare burden.

Prevalence of Listeria monocytogenes and Salmonella spp. in Different Ready to Eat Foods from Large Retailers and Canteens over a 2-Year Period in Northern Italy.

This study aims to give an overview of the prevalence of Listeria monocytogenes and Salmonella spp. in 9727 samples (2996 for L. monocytogenes and 6731 for Salmonella spp.) from different categories of ready-to-eat (RTE) foods, collected over 2 years from 28 large retailers and 148 canteens in the regions of northern Italy. The RTE samples were classified into two groups according to the preparation methods: (i) multi-ingredient preparations consisting of fully cooked food ready for immediate consumption, or with minimal further handling before consumption (Group A), and (ii) multi-ingredient preparations consisting of cooked and uncooked food, or preparations consisting of only raw ingredients (Group B). L. monocytogenes and Salmonella spp. were investigated in both of these categories. The overall prevalence of L. monocytogenes and Salmonella spp. was 0.13% and 0.07%, respectively. More specifically, L. monocytogenes was found in 0.04% of 2442 analysed RTE food samples belonging to group A and in 0.54% of 554 samples belonging to group B. Furthermore, 0.03% of 5367 RTE food samples from group A and 0.21% of 1364 samples from group B tested positive for Salmonella spp. In conclusion, the results obtained in this study can provide a significant contribution to L. monocytogenes and Salmonella spp. risk analysis in RTE foods.

Characterization of class 1 integrons harboring blaVEB-1 in Vibrio parahaemolyticus isolated from ready-to-eat foods in China.

The aim of this study is to investigate the prevalence of integrons and integron-associated antibiotic resistance in V. parahaemolyticus strains collected from RTE foods in China, and to carry out a comprehensive analysis on the molecular characterization of V. parahaemolyticus strains carrying blaVEB-1-positive class 1 integron. Of the 51 V. parahaemolyticus strains isolated from RTE food samples, none of the isolates was found to carry integrase genes intI2 and IntI3. However, all 51 strains were positive to integrase gene intI1, and only 2 of 51 (3.92%) intI1-positive isolates yielded polymerase chain reaction (PCR) products of gene cassette amplification. Sequence data and BLAST analysis indicated the gene cassette arrays of class 1 integron in VP007 is dfrA14-blaVEB-1-aadB, while the gene cassette arrays of class 1 integron in V187 is blaVEB-1-aadB-arr2-cmlA-blaOXA-10-aadA1. Antimicrobial susceptibility testing showed that the two V. parahaemolyticus isolates harboring class 1 integrons exhibited multi-drug resistance to various antibiotics. S1-PFGE and Southern blot analysis confirmed the class 1 integron harboring blaVEB-1 gene in V187 was located on the plasmid of ~175 kb and transferrable to the recipient strain by conjugation. This is the first detection of class 1 integrons harboring the ESBL gene blaVEB-1 in V. parahaemolyticus. To the best of our knowledge, this is also the first report of VEB-producing V. parahaemolyticus from RTE foods. Our findings revealed that class 1 integron on conjugative plasmid contributes significantly to the dissemination of VEB-producing V. parahaemolyticus, which warrants further investigation because of the public health threat it poses.

Resistance to ß-lactams in bacteria isolated from different types of Portuguese cheese.

The purpose of this study was to investigate the presence of beta-lactam-resistant bacteria in six different types of Portuguese cheese. The numbers of ampicillin resistant (AMP(r)) bacteria varied from 4.7 x 10(2) to 1.5 x 10(7) CFU/g. Within 172 randomly selected beta-lactam-resistant bacteria, 44 resistant phenotypes were found and 31.4% were multidrug resistant. The majority (85%) of the isolates identified belonged to the Enterobacteriaceae family. The presence of the bla(TEM) gene was detected in 80.9% of the tested isolates. The results suggest that without thermal processing of the milk and good hygienic practices, cheese may act as a vehicle of transfer of beta-lactam-resistant bacteria to the gastrointestinal tract of consumers.

Experimental accelerated shelf life determination of a ready-to-eat processed food.

The most direct way to estimate the shelf life of a product is to conduct simulation tests which are time consuming and expensive. Conversely, accelerated shelf life tests can be successfully used for stable products having long expected shelf life. The aim of the study was directed to verify the possibility to apply an accelerated shelf life test to perishable food products having a short-expected shelf life, such as a new ready-to-eat processed food preparation, composed mainly by cereals, tuna and chicken, packed in thermo-sealed trays and pasteurised. Different samples of the product were stored in thermal abuse conditions, collected periodically and subjected to determinations of TVB-N, pH and sensorial characteristics. Q10 and activation energy were calculated allowing to obtain a predictive evaluation of the product shelf life at the 4°C recommended temperature. The product shelf life was assessed at 26 days vs the 30 days expected by the manufacturer, showing the possibility to apply successfully ASLT for products having short shelf life, saving both time and money.

Development of prototypes of bioactive packaging materials based on immobilized bacteriophages for control of growth of bacterial pathogens in foods.

Due to lack of adequate control methods to prevent contamination in fresh produce and growing consumer demand for natural products, the use of bacteriophages has emerged as a promising approach to enhance safety of these foods. This study sought to control Listeria monocytogenes in cantaloupes and RTE meat and Escherichia coli O104:H4 in alfalfa seeds and sprouts under different storage conditions by using specific lytic bacteriophage cocktails applied either free or immobilized. Bacteriophage cocktails were introduced into prototypes of packaging materials using different techniques: i) immobilizing on positively charged modified cellulose membranes, ii) impregnating paper with bacteriophage suspension, and iii) encapsulating in alginate beads followed by application of beads onto the paper. Phage-treated and non-treated samples were stored for various times and at temperatures of 4°C, 12°C or 25°C. In cantaloupe, when free phage cocktail was added, L. monocytogenes counts dropped below the detection limit of the plating technique (<1 log CFU/g) after 5 days of storage at both 4°C and 12°C. However, at 25°C, counts below the detection limit were observed after 3 and 6h and a 2-log CFU/g reduction in cell numbers was seen after 24h. For the immobilized Listeria phage cocktail, around 1-log CFU/g reduction in the Listeria count was observed by the end of the storage period for all tested storage temperatures. For the alfalfa seeds and sprouts, regardless of the type of phage application technique (spraying of free phage suspension, bringing in contact with bacteriophage-based materials (paper coated with encapsulated bacteriophage or impregnated with bacteriophage suspension)), the count of E. coli O104:H4 was below the detection limit (<1 log CFU/g) after 1h in seeds and about a 1-log cycle reduction in E. coli count was observed on the germinated sprouts by day 5. In ready-to-eat (RTE) meat, LISTEX™ P100, a commercial phage product, was able to significantly reduce the growth of L. monocytogenes at both storage temperatures, 4°C and 10°C, for 25 days regardless of bacteriophage application format (immobilized or non-immobilized (free)). In conclusion, the developed phage-based materials demonstrated significant antimicrobial effect, when applied to the artificially contaminated foods, and can be used as prototypes for developing bioactive antimicrobial packaging materials capable of enhancing the safety of fresh produce and RTE meat.

The Role of Stress and Stress Adaptations in Determining the Fate of the Bacterial Pathogen Listeria monocytogenes in the Food Chain.

The foodborne pathogen Listeria monocytogenes is a highly adaptable organism that can persist in a wide range of environmental and food-related niches. The consumption of contaminated ready-to-eat foods can cause infections, termed listeriosis, in vulnerable humans, particularly those with weakened immune systems. Although these infections are comparatively rare they are associated with high mortality rates and therefore this pathogen has a significant impact on food safety. L. monocytogenes can adapt to and survive a wide range of stress conditions including low pH, low water activity, and low temperature, which makes it problematic for food producers who rely on these stresses for preservation. Stress tolerance in L. monocytogenes can be explained partially by the presence of the general stress response (GSR), a transcriptional response under the control of the alternative sigma factor sigma B (σB) that reconfigures gene transcription to provide homeostatic and protective functions to cope with the stress. Within the host σB also plays a key role in surviving the harsh conditions found in the gastrointestinal tract. As the infection progresses beyond the GI tract L. monocytogenes uses an intracellular infectious cycle to propagate, spread and remain protected from the host's humoral immunity. Many of the virulence genes that facilitate this infectious cycle are under the control of a master transcriptional regulator called PrfA. In this review we consider the environmental reservoirs that enable L. monocytogenes to gain access to the food chain and discuss the stresses that the pathogen must overcome to survive and grow in these environments. The overlap that exists between stress tolerance and virulence is described. We review the principal measures that are used to control the pathogen and point to exciting new approaches that might provide improved means of control in the future.

Toxin production and growth of pathogens subjected to temperature fluctuations simulating consumer handling of cold cuts.

It is crucial for the quality and safety of ready-to-eat (RTE) foods to maintain the cold chain from production to consumption. The effect of temperature abuse related to daily meals and elevated refrigerator temperatures on the growth and toxin production of Bacillus cereus, Bacillus weihenstephanensis and Staphylococcus aureus and the growth of Listeria monocytogenes and Yersinia enterocolitica was studied. A case study with temperature loggings in the domestic environment during Easter and Christmas holidays was performed to select relevant time and temperature courses. A model for bacterial surface growth on food using nutrient agar plates exposed to variations in temperatures was used to simulate food stored at different temperatures and exposed to room temperature for short periods of time. The results were compared with predicted growth using the modeling tool ComBase Predictor. The consumers exposed their cold cuts to room temperatures as high as 26.5°C with an average duration of meals was 47 min daily for breakfast/brunch during the vacations. Short (≤ 2 h) daily intervals at 25°C nearly halved the time the different pathogens needed to reach levels corresponding to the levels associated with human infection or intoxication, compared with the controls continuously stored at refrigerator temperature. Although the temperature fluctuations affected growth of both B. weihenstephanensis and S. aureus, toxin production was only detected at much higher cell concentrations than what has been associated with human intoxications. Therefore, growth of L. monocytogenes and Y. enterocolitica was found to be the limiting factor for safety. In combination with data on temperature abuse in the domestic environment, modeling programs such as ComBase Predictor can be efficient tools to predict growth of some pathogens but will not predict toxin production.