Milk, due to its diversity in terms of its nutritional content, is an important element of the human diet, as well as a good medium for the development of bacteria. The genus Bacillus contains ubiquitous aerobic, rod-shaped, endospore-producing gram-positive bacteria. Representatives of the Bacillus cereus group and the Bacillus subtilis group contribute to shortening the shelf life of milk and dairy products by degrading milk components and its additives. They also produce a number of heat-stable toxins and can cause a number of ailments, mainly in the digestive system. The aim of this research was to identify Bacillus sp. strains isolated from raw milk and to determine their antibiotic resistance. Strains isolated from raw milk samples (n = 45) were identified by MALDI-TOF MS. Ninety strains of Bacillus sp. were identified, for which the antibiotic resistance phenotype was determined. A total of 90 strains of Bacillus were classified in five groups (the Bacillus cereus group (n = 35), B. licheniformis (n = 7), the B. subtilis group (n = 29), B. pumilus (n = 16), and Bacillus sp. (n = 3). All isolates were susceptible to chloramphenicol and meropenem. The antibiotic resistance profiles of the tested groups of Bacillus spp. differed from each other, which is of particular concern in relation to multidrug-resistant representatives of the B. cereus group resistant to cefotaxime (94.29%), ampicillin (88.57%), rifampicin (80%), and norfloxacin (65.71%). Our study provides data on the prevalence and antibiotic sensitivity of Bacillus sp. In raw milk, suggesting a potential risk to health and the dairy industry.

Bacterial biofilm is one of the major hazards facing the food industry. Biofilm-forming ability is one of the most important virulence properties of enterococci. The genus Enterococcus includes pathogenic, spoilage, and pro-technological bacteria. The presence of enterococci in milk and dairy products is usually associated with inadequate hygiene practices. The study examined the isolates’ capacity for biofilm formation and identification of the genetic determinants of its formation among 85 Enterococcus strains isolated from raw milk (n = 49) and soft-ripened cheeses made from unpasteurized milk (n = 36). E. faecalis and E. faecium were the dominant species. The obtained results showed that 41.4% isolates from milk and 50.0% isolates from cheeses were able to form biofilm. All of the isolates analyzed had at least one of the studied genes. As regards the isolates from raw milk, the most prevalent gene was the gelE (85.6%), followed by the asa1 (66.7%). None of the isolates from cheeses showed the presence of cylA and sprE. The most prevalent gene among the strains from this source was the epbC (94.4%), followed by the gelE (88.9%). In isolates from both sources, the presence of proteins from the Fsr group was noted the least frequently. Nevertheless, results showed that were no significant differences between the biofilm-producing Enterococcus spp. and non-biofilm-producing isolates in term of occurrences of tested virulence genes. The ability to produce a biofilm by enterococci isolated from raw milk or ready-to-eat products emphasizes the need for continuous monitoring of the mechanisms of microbial adhesion.

Dairy products from unpasteurized milk produced using traditional production methods may contain many groups of microorganisms, including Staphylococcus aureus and Listeria monocytogenes. Therefore, the use of postbiotics as an alternative preservation method may be important for improving the safety of these products. Therefore, the main aim of the research conducted was to isolate and identify lactic acid bacteria and prepare postbiotics from selected strains using four different methods, sterilization, pasteurization, sonication and pascalization, to determine their antibacterial properties. The antilisterial and antistaphylococcal activities of the prepared postbiotics were investigated in vitro and in a fresh cheese model. The obtained results showed that the most effective method of postbiotic preparation was pascalization. Both, the results of the MRS medium and the challenge test confirmed that postbiotics produced through pascalization exhibited antistaphylococcal activity. This study may help identify more effective biopreservation compounds to combat pathogens in food products, particularly in dairy products.

Coagulase-negative staphylococci (CoNS), including Staphylococcus epidermidis, are commonly occurrence in a variety of food products. Historically considered non-pathogenic, these microorganisms were excluded from routine food safety monitoring protocols. However, their increasing involvement in nosocomial infections underscores their pathogenic potential. Emerging evidence suggests that the food chain may serve as a reservoir and transmission route for antibiotic-resistant bacteria. In this study, 26 S. epidermidis isolates obtained from ready-to-eat food were subjected to whole-genome sequencing and comprehensive bioinformatics analyses. The antimicrobial susceptibility of the isolates was also evaluated against a broad spectrum of agents including aminoglycosides, β-lactams, fluoroquinolones, glycopeptides, lincosamides, macrolides, nitrofurantoins, oxalidinones, phenicols, steroids, sulphonamides and tetracyclines. Sequence typing revealed the presence of 17 distinct sequence types (STs), with ST329 being the most frequently identified (8/26, 30.77 %), followed by ST88 and ST152 (each 2/26; 7.69 %). Notably, one isolate harbored a novel multi-locus sequence type. Phenotypically resistance to erythromycin was most prevalent (21/26, 80.77 %), followed by resistance to clindamycin (19/26, 73.08 %). Genomic analysis confirmed the presence of multiple antimicrobial resistance genes including norA/C, vanT, mecA, dfrC and multidrug resistance genes. The carrying of mobile genetic elements was demonstrated by 25/26 (96.15 %) strains. These findings indicate that S. epidermidis strains isolated from ready-to-eat foods not only exhibit multidrug resistance but also carry a diverse array of antimicrobial resistance genes. The potential for horizontal gene transfer to commensal or pathogenic bacteria highlights the need for increased surveillance and risk assessment concerning CoNS in the food supply.

Antimicrobial resistance is spreading rapidly throughout the world. The food chain can be one of the routes of transmission for microorganisms containing drug-resistance genes and thus serve as a channel for their transmission. Environmental stress and methods of preventing the spread of microorganisms trigger adaptive responses in bacterial cells. The aim of the present study was to determine the effect of the stress induced by sub-inhibitory concentrations (SICs) of cadmium chloride and benzalkonium chloride on antibiotic resistance and the expression of selected virulence factors in Staphylococcus isolates from food. The study was conducted on strains of the species S. epidermidis, S. heamolyticus, S. saprophyticus, and S. aureus. The values of the minimum inhibitory concentration against erythromycin, tetracycline, and oxacillin were determined before and after the incubation of the tested strains under stress conditions. The ability to form biofilm and slime production was also investigated. The expression levels of the genes responsible for antibiotic resistance (blaZ, tetK, tetM, ermB, and mecA) and virulence (eno) were conducted using Real-Time PCR. The MIC values of the antibiotics tested against the strains analyzed were found to be elevated in the presence of SICs of benzalkonium chloride and cadmium chloride. Furthermore, the intensity of biofilm production was also increased. SICs of benzalkonium chloride induced the expression of the tetM, tetK, mecA, and blaZ genes in 75%, 66.6%, 33.3%, and 40% of the isolates tested, respectively. Similar treatment with cadmium chloride induced the expression of the same genes in 75%, 100%, 66.6%, and 40% of the strains. In both cases, the expression of the ermB gene was reduced in 100% of the isolates. The eno gene was found to be overexpressed in 66.6% of the strains following benzalkonium chloride stress, and in 100% of the strains following cadmium chloride stress. These findings suggest that in Staphylococcus spp. strains, changes in the expression of the genes encoding antibiotic resistance and virulence factors may occur in response to the applied stress factors. The results indicate the possibility of selecting more resistant and virulent strains due to the use of too low concentrations of disinfectants, which emphasizes the need to use appropriate inhibitory doses of disinfectants in the food industry.