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Listeria monocytogenes (Lm) is responsible for listeriosis, a serious foodborne disease, with high hospitalization and mortality rates worldwide. The main cause of listeriosis in humans is the consumption of ready-to-eat (RTE) foods; Commission Regulation (EC) No 2073/2005 establishes microbiological criteria for Lm in RTE foods. Raw fish products are widely consumed, e.g., in sushi and various seafood recipes (e.g., carpaccio, sashimi, maki, nigiri, tartare, etc.), but are not subjected to RTE food safety criteria. The aim of our study was to assess the growth potential of Lm in raw sea bass fillets obtained from a leading aquaculture company in Greece. In order to assess the growth of Lm in raw sea bass fillets, we applied the “challenge test”, a scientific experiment designed to assess the growth of Lm within a specific food product under controlled conditions. According to our results, and taking into consideration the health risk for the listeriosis-vulnerable population, raw fish products utilized in the preparation of RTE foods, including sushi and an array of seafood dishes, should be incorporated in the Category of Safety Criteria of Regulation (EC) No 2073/2005 “Ready-to-eat food able to support the growth of Listeria monocytogenes”.

Hummus is a traditional and very popular Mediterranean ready-to-eat (RTE) food, with growing popularity worldwide. However, it has a high water activity and is susceptible to microbial growth and post-process contamination that limit its quality and shelf-life. For this purpose, the present study compared the use of several antimicrobials, alone or in combination, for hummus preservation during storage (4 °C), for up to 45 days. The chemical preservative potassium sorbate 0.09% (S) was evaluated, along with three natural antimicrobials: garlic 1.25% (G); vinegar 5% (V); natamycin 0.002% (N); or their combination: garlic 1.25%–vinegar 5% (GV); vinegar 5%–natamycin 0.002% (VN); garlic 1.25%–natamycin 0.002% (GN); and garlic 1.25%–vinegar 5%–natamycin 0.002% (GVN) to increase the shelf-life of hummus. A thymol and carvacrol mixture 0.2% (O) was also assessed to preserve and develop a new oregano-flavored hummus. All treatments that included vinegar used alone or in combination had significantly higher antimicrobial effectiveness than the other treatments. They achieved 2.2–3.2, 1.8–3.1, and 1.4–2.1 log reductions in total aerobic counts (TAC), Pseudomonas spp., and lactic acid bacteria (LAB), respectively, as compared to the control samples © at day 21. Therefore, the shelf-life of C, S, N, G, GN, and O was around (ca.) 19 days, compared to an extended one of ca. 25 days for V and VN, and ca. 30 days for GV and GVN. Sensory analysis showed the highest acceptability for C, N, S, V, and VN, followed by GV and GVN, and the lowest was for G, GN, and finally O. The findings provide potential alternatives to chemical preservatives, which could be used for natural hummus preservation and shelf-life extension.

Vegetables are an essential component of a balanced diet. The consumption of ready-to-eat foods may lead to the risk of infections and illnesses due to microbial contamination. To mitigate the potential of microbial contamination risks, it is critical to promote safe handling practices among consumers. In this study, our research evaluated the efficacy of different vegetable washing methods, specifically with lettuce, tomato, and cucumber, to establish optimal practices for reducing microbial contamination. This study consisted of two phases. Initially, a survey was distributed to 150 volunteers using snowball sampling to assess everyday vegetable handling and washing methods. The survey's results identified four predominant methods: washing with a 5% vinegar solution for 3 min followed by tap water rinse (37.3% of participants), rinsing with tap water for 1 min (29.3%), washing with a 5% salt solution (vegetable soap) for 3 min followed by a tap water rinse (16.6%), and a 3 min tap water rinse (14%). A minor segment (3.33%) reported not washing their vegetables at all. The survey's findings guided the second phase, which tested the aforementioned washing protocols' effectiveness in reducing Escherichia coli ( E. coli ) levels on spiked contaminated salad vegetables. The tested vegetables were sterilized using UV light, inoculated with 0.5 McFarland E. coli , and then washed using the four identified methods. After that, E. coli enumeration after washing was performed using 3M™ Petrifilm and the comparison was analyzed via one-way ANOVA. During this study, it was revealed that the cucumbers had the highest E. coli contamination levels in comparison to the lettuce and tomato after washing. Interestingly, by comparing the three washing methods, it was found that washing the vegetables with vinegar proved to be the most effective solution for reducing microbial presence on both lettuce and cucumbers. Notably, the natural smoothness of tomato skin led to no significant differences in contamination levels across washing methods. In summary, vinegar washing effectively reduces microbial contamination from salad vegetables, highlighting the need for informed consumer practices to prevent foodborne outbreaks. This study emphasizes the importance of monitoring contamination sources and using safe washing techniques.

The present study aimed to characterize and assess the diversity of CoNS strains as potential vectors for the spread of resistance to antimicrobial agents from RTE foods served in bars and restaurants. Eighty-five CoNS strains, obtained from 198 RTE food samples, were investigated. Sixty-seven CoNS isolates (78.8%) were resistant to at least one antibiotic tested, and 37 (43.5%) were multidrug resistant (MDR-CoNS). Moreover, CoNS strains contained genes conferring resistance to antibiotics critically important in medicine, i.e., β—lactams [mecA (29.4%); blaZ (84.7%)], aminoglycosides [aac(6′)-Ie-aph(2″)-Ia (45.9%); aph(2″)-Ic (3.5%)], macrolides, lincosamides and streptogramin B-MLSB [msrA/B (68.2%); ermB (40%) and mphC (4.7%)], tetracyclines [tetK (31.8%); tetM (16.5%) and/or tetL (2.35%)]. We also found the fusB/C/D genes responsible for the acquired low-level fusidic acid resistance (17.6%) and streptogramin resistance determinant vgaA in 30.6% of isolates. In three linezolid resistant strains (2 S. epidermidis and 1 S. warneri), mutation was detected, as demonstrated by L101V and V188I changes in the L3 protein amino acid sequences. The high frequency in RTE food of MDR-CoNS including methicillin-resistant (MR-CoNS) strains constitutes a direct risk to public health as they increase the gene pool from which pathogenic bacteria can pick up resistance traits.

Foodborne listeriosis is one of the most serious and severe foodborne diseases, with a high mortality rate worldwide. Listeria monocytogenes’ (Lm) ability to survive under a diversity of conditions makes it a threat for food safety. Soft and semisoft cheeses are common RTE foods that support survival and growth of Lm due to their high moisture content and favorable pH. The aim of the present study was to assess, after artificial inoculation, the growth potential (Δ) of Lm in vacuum packed RTE soft Greek Anthotyros cheese. Growth potential of Lm was determined according to the new version of “EURL Lm Technical Guidance Document on challenge tests and durability studies for assessing shelf life of ready-to-eat foods related to Lm”. The results of our study revealed that the growth potential of Lm was (Δ) = 4.93 log10 CFU/g, indicating that the specific soft cheese “anthotyros” is a “Ready to eat food able to support growth of Lm and classified in food safety category 1.2 in the (EC) Regulation 2073/2005”. That means that the food business operator (FBO) must comply with the criteria that define that the bacterium must not be detected in 25 g (n = 5, c = 0) at the time of leaving the production plant. Through this study, we try to highlight the need for FBO to conduct relevant research to ensure that the ready-to-eat products which they produce, during their shelf life, do not support the growth of Lm.

Staphylococcus aureus is a pathogenic microorganism of humans and animals, able to cause foodborne intoxication due to the production of staphylococcal enterotoxins (SEs) and to resist antibiotic treatment as in the case of methicillin-resistant S. aureus (MRSA). In this study, we performed a genomic characterisation of 12 genetically diverse S. aureus strains isolated from ready-to-eat foods in Algiers (Algeria). Moreover, their ability to produce some classical and new staphylococcal enterotoxins (SEs) was investigated. The 12 S. aureus strains resulted to belong to nine known sequence types (STs) and to the novel ST7199 and ST7200. Furthermore, S. aureus SA46 was assigned to the European clone MRSA-ST80-SCCmec-IV. The 12 strains showed a wide endowment of se and sel (staphylococcal enterotoxin-like toxin) genes (sea, seb, sed, seg, seh, sei, selj, sek, sem, sen, seo, seq, ser, selu2, selw, selx, sey, sel30; ψent1-ψent2), including variants and pseudogenes, and harboured the enterotoxin gene cluster (egc) types 1 and 5. Additionally, they produced various amounts of SEA (64.54–345.02 ng/mL), SEB (2871.28–14739.17 ng/mL), SED (322.70–398.94 ng/mL), SEH (not detectable–239.48 ng/mL), and SER (36,720.10–63,176.06 ng/mL) depending on their genotypes. The genetic determinants related to their phenotypic resistance to β-lactams (blaZ, mecA), ofloxacin (gyrA-S84L), erythromycin (ermB), lincomycin (lmrS), kanamycin (aph(3′)-III, ant(6)-I), and tetracyclin (tet(L), tet(38)) were also detected. A plethora of virulence-related genes, including major virulence genes such as the tst gene, determinant for the toxic shock syndrome toxin-1, and the lukF-PV and lukS-PV genes, encoding the panton-valentine leukocidin (PVL), were present in the S. aureus strains, highlighting their pathogenic potential. Furthermore, a phylogenomic reconstruction including worldwide foodborne S. aureus showed a clear clustering based on ST and geographical origin rather than the source of isolation.

Listeriosis is almost entirely transmitted through foods contaminated with Listeria monocytogenes. Ready-to-eat foods present a particular challenge due to their long refrigerated shelf-life, not requiring any heat treatment before consumption. In this work, a shelf-life assessment of an industrially produced ready-to-eat salad was performed using conventional culture-based and molecular methods. L. monocytogenes isolates were confirmed and serogrouped using multiplex PCR, and genetic subtyping was performed by pulsed-field gel electrophoresis (PFGE). PMAxx-qPCR was used as an alternative method for L. monocytogenes quantification in foods. Salad samples were kept at 4 °C, 12 °C, and 16 °C for eight days and analysed. At 4 °C, acceptable results were obtained considering hygiene indicators, i.e., Enterobacteriaceae (ranging from 3.55 ± 0.15 log cfu/g to 5.39 ± 0.21 log cfu/g) and aerobic mesophilic colony counts (5.91 ± 0.90 log cfu/g to 9.41 ± 0.58 log cfu/g) throughout the study, but the same did not happen at 12 °C and 16 °C. L. monocytogenes culture-based quantification exhibited low numbers (L. monocytogenes with the majority belonging to serogroup IVb. PFGE subtyping showed that 7 of the 10 L. monocytogenes isolates had 100% of pulsotype similarity, suggesting a possible common contamination source. PMAxx-qPCR revealed a statistically higher L. monocytogenes quantification (>3 log cfu/g) when compared to the conventional culture-based method, suggesting viable but non-culturable forms. Taken together, results underline the need to combine conventional methods with more sensitive, specific, and rapid ones for L. monocytogenes assessment in ready-to-eat foods shelf-life studies to reduce the potential risk for consumers.

Ready-to-eat food microbiota are the microorganisms present in the dishes that are currently consumed during meals. These microorganisms include those that may have a health benefit, are potentially pathogenic or have not yet been given a function. Foods suitable for consumption are not free of microorganisms; however, within the food industry only yeasts have been given a beneficial function, while other microorganisms such as filamentous fungi and bacteria have been studied for their negative effects on food. We determined the bacterial diversity in samples of highly demanded, freshly prepared, unspoiled ready-to-eat dishes by high-throughput DNA sequencing of 16S rDNA libraries. We found a great bacterial diversity, whereby the most abundant bacterial phyla were Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, TM7 and Thermi, among others. These phyla included bacteria with remarkable abundances in some dishes. The alfa diversity analyses showed that the main dishes had the largest diversity. The beta-diversity analyses clustered the bacterial communities of soups, side plates, desserts, and beverages, and some main dishes. Based on our results we conclude that unspoiled ready-to-eat Mexican dishes contain a rich bacterial diversity, which may contribute to the organoleptic properties of the dishes without representing a sanitary risk for the consumers.

Although coagulase-positive staphylococci are considered to be the main factor responsible for food poisoning, an increasing role for the coagulase-negative staphylococci in the production of enterotoxins has been observed in recent years. This study was conducted to assess the occurrence of genes responsible for the production of staphylococcal enterotoxins (SE), enterotoxin-like toxins (SEI) and toxic shock syndrome toxin-1 (TSST-1) in coagulase-negative staphylococci (CoNS) isolated from ready-to-eat food from bars and restaurants. One hundred and eighteen CoNS strains were tested using polymerase chain reaction (PCR) to five superantigenic toxin genes, including five different types of classical enterotoxins (sea, seb, sec, sed and see) and the toxic shock syndrome toxin-1 (tsst-1) as well as to supertoxin-like genes. PCR-positive isolates were then tested using immunoenzymatic methods (SET-RPLA, Vidas SET 2) for toxin expression. Out of 118 CoNS strains, the presence of staphylococcal enterotoxins was confirmed in 72% of them. The most frequently found enterotoxin-like genotype was ser, selu. Two of the tested strains had up to ten different enterotoxin genes in the genome at the same time. Although no production of enterotoxins was detected in the CoNS, which means that their possible role in the epidemiology of food-borne diseases is minimal, the data demonstrated that the toxigenic capacity of the CoNS should not be ignored, and that this group of microorganisms should be continuously monitored in food.

The prevalence of Staphylococcus aureus in 2160 bulk ready-to-eat foods from the Sichuan province of China during 2013–2016 was investigated. The antibiotic resistance and the associated genes, as well as biofilm formation capacity of the S. aureus isolates were measured. Furthermore, the relationship between the antibiotic resistance and the resistant genes was discussed. It was found that 54 S. aureus isolates were recovered, and their prevalence in meat products, dairy, fruit and vegetables, and desserts were 31 (2.6%), six (3.0%), nine (2.2%) and eight (2.3%), respectively. Most strains (52/54) were resistant to at least one of the antibiotics, and 21 isolates were identified as multidrug-resistant (MDR) S. aureus. Three isolates were found to be methicillin-resistant S. aureus. Penicillin, erythromycin, clindamycin, tetracycline and inducible clindamycin resistance were determined as the predominant antibiotics, and the isolates with the phenotypic resistance on these five antibiotics were all determined positive for the resistant gene associated. In total, 33 of 54 S. aureus isolates showed biofilm formation capacity, including two strong biofilm producers, one moderate and 30 weak ones. Two S. aureus isolates with strong biofilm formation abilities showed multi-drug resistance, and one moderate biofilm producer was resistant to two categories of antibiotics.

Simultaneous determination of twenty-seven mycotoxins in ready-to-eat food samples using “Quick Easy Cheap Rough and Safe” (QuEChERS) extraction and chromatographic methods coupled to mass spectrometry in tandem is described in this study. Mycotoxins included in this survey were aflatoxins (B1, B2, G1, G2), enniatins (A, A1, B, B1), beauvericin (BEA), fumonisins (FB1, FB2), sterigmatocystin (STG), deoxynivalenol (DON), 3-acetyl-deoxynivalenol (3-ADON), 15-acetyl-deoxynivalenol (15-ADON), nivalenol (NIV), neosolaniol (NEO), diacetoxyscirpenol (DAS), fusarenon-X (FUS-X), zearalenone (ZEA), α-zearalanol (αZAL), β-zearalenone (βZAL), α-zearalenol (αZOL), β-zearalenol (βzol), T2, and HT-2 toxin. The method showed satisfactory extraction results with recoveries ranging from 63 to 119% for the different food matrix samples. Limits of detection (LODS) and quantification (LOQs) were between 0.15–1.5 µg/kg and 0.5–5 µg/kg, respectively. The method was successfully applied to the analysis of 25 ready-to-eat food samples. Results showed presence of deoxynivalenol at 36% of samples (2.61–21.59 µg/kg), enniatin B at 20% of samples (9.83–86.32 µg/kg), HT-2 toxin at 16% of samples (9.06–34.43 µg/kg), and aflatoxin G2 at 4% of samples (2.84 µg/kg). Mycotoxins were detected mainly in ready-to-eat food samples prepared with cereals, vegetables, and legumes, even at levels below those often obtained from raw food.