Food Processing Environments Research Articles

Antimicrobial activity of magnolol against Bacillus cereus and its application as food preservative

The aim of this study was to explore the antibacterial properties and mechanism of magnolol against Bacillus cereus. Our findings demonstrated that magnolol exhibited potent antimicrobial activity against B. cereus, with minimum inhibitory concentrations (MICs) of 2 μg/mL. Additionally, magnolol could inhibit spore germination and biofilm formation in a dose-dependent manner. Electron microscopy and membrane potential analysis indicated that the cytomembranes were disrupted, resulting in leakage of cellular contents. Transcriptomic profiling showed that magnolol treatment (1/2 × MIC) resulted in 585 differentially expressed genes (DEGs), including 295 down-regulated and 290 up-regulated genes. Bioinformatic analysis indicated that magnolol affected many potential cellular responses, including cellular metabolic process, locomotion, oxidative stress, and cell membranes. The pathway analysis revealed that DEGs mainly associated with flagellar assembly, bacterial chemotaxis, amino acids, and energy metabolism. Moreover, magnolol obviously reduced the amount of B. cereus in boiled rice and beef at 4 °C storage. These results provided insights into the molecular mechanism of magnolol against B. cereus and suggested that magnolol might be a promising natural antimicrobial agent for controlling the B. cereus in foods. Industrial relevanceBacillus cereus is considered as an important foodborne pathogenic microorganism that can exist as vegetative cells, spores, and biofilm in various food-processing environment, posing a big challenge for the food industry. Magnolol is a natural compound with potent antimicrobial activity against various pathogens and has been certified as safe by food safety organizations. Consequently, magnolol holds great promise as a natural antimicrobial agent for controlling B. cereus contaminations in foods.

The hidden threat: Comprehensive assessment of antibiotic and disinfectant resistance in commercial pig slaughterhouses

The presence of antibiotic resistance genes (ARGs), disinfectant resistance genes (DRGs), and pathogens in animal food processing environments (FAPE) poses a significant risk to human health. However, knowledge of the contamination and risk profiles of a typical commercial pig slaughterhouse with periodic disinfectant applications is limited. By creating the overall metagenomics-based behavior and risk profiles of ARGs, DRGs, and microbiomes in a nine-section pig slaughterhouse, an important FAPE in China. A total of 454 ARGs and 84 DRGs were detected in the slaughterhouse with resistance genes for aminoglycosides and quaternary ammonium compounds, respectively. The entire slaughtering chain is a hotspot for pathogens, including 83 human pathogenic bacteria (HPB), with 47 core HPB. In addition, 68 high-risk ARGs were significantly correlated with 55 HPB, 30 of which were recognized as potential bacteria co-resistant to antibiotics and disinfectants, confirm a three-fold risk of ARGs, DRGs, and pathogens prevailing throughout the chain. Pre-slaughter pig house (PSPH) was the major risk source for ARGs, DRGs, and HPB. Moreover, 75 Escherichia coli and 47 Proteus mirabilis isolates showed sensitivity to potassium monopersulfate and sodium hypochlorite, suggesting that slaughterhouses should use such related disinfectants. By using whole genome multi-locus sequence typing and single nucleotide polymorphism analyses, genetically closely related bacteria were identified across distinct slaughter sections, suggesting bacterial transmission across the slaughter chain. Overall, this study underscores the critical role of the PSPH section as a major source of HPB, ARGs, and DRGs contamination in commercial pig slaughterhouses. Moreover, it highlights the importance of addressing clonal transmission and cross-contamination of antibiotic- and disinfectant-resistant bacteria within and between slaughter sections. These issues are primarily attributed to the microbial load carried by animals before slaughter, carcass handling, and content exposure during visceral treatment. Our findings provide valuable insights for One Health-oriented slaughterhouse management practices.

Levels of Different Microbial Groups on Inert Surfaces of Poultry Slaughterhouses: Identification Using Matrix-Assisted Laser Desorption Ionization Time-of-Flight and Detection of Extended-Spectrum Beta-Lactamase- and Carbapenemase-Producing Enterobacteria.

Knowledge of the microbiota present in food processing environments is a significant advance that will allow for better evaluation of the risk of food contamination and a better design of the procedures for sanitization. The levels of microbial group indicators of hygienic quality were determined in different areas of the slaughter lines of two poultry slaughterhouses in the northwest of Spain (22 surfaces in each slaughterhouse were studied). The average microbial levels (cfu/cm2) were 2.15 × 102 ± 4.26 × 102 (total aerobic counts, TAC), 1.99 × 102 ± 5.00 × 102 (psychrotrophic microorganisms), 3.10 × 100 ± 1.37 × 101 (enterobacteria), 3.96 × 100 ± 2.55 × 101 (coliforms), 1.80 × 10-1 ± 7.79 × 10-1 (enterococci), and 1.12 × 10-1 ± 3.35 × 10-1 (vancomycin-resistant enterococci, VRE). TAC and psychrotrophic microorganisms were the most abundant groups in all samples (p < 0.05). The counts of both microbial groups were higher (p < 0.05) in samples of Slaughterhouse A than in those of Slaughterhouse B. Microbial loads for the rest of the bacteria were not influenced by the slaughterhouse sampled (p > 0.05). All 44 samples showed TAC and psychrotrophic microorganisms. Colonies of the rest of the microbial groups were only found in 26 samples (59.1% of the total). The isolates (one from each sample) were identified with MALDI-TOF and PCR. Gram-negative bacteria (all Enterobacteriaceae) were isolated in 23 samples, and Gram-positive bacteria were isolated in 16 (9 Enterococcus spp., 2 Enterococcus spp. and VRE, 3 VRE, 1 Enterococcus spp. and Listeria spp., and 1 Listeria spp.). The resistance of the strains to 11 (Enterococcus spp.) or 17 (Enterobacteriaceae) antibiotics was determined (disk diffusion, CLSI), finding an average of 2.05 ± 2.06 resistances per strain (3.46 ± 2.27 if reduced susceptibility reactions are included). A total of 37.3% of the Enterobacteriaceae isolates had a gene for resistance to beta-lactam antibiotics (blaTEM, blaCTX-M-15, blaKPC, blaCMY-2 or blaNDM). The high prevalence of resistant bacteria and resistance genes highlights the need to establish measures to control the spread of antibiotic resistance in poultry slaughterhouses. The findings of this work could contribute to the design of more effective sanitation procedures.

Draft genome sequences of two wide-host-range phages of Listeria monocytogenes from food processing environments in the United States.

Listeria monocytogenes is notorious for persistence in food facilities. Phages can significantly impact the ecology of Listeria, but there is a dearth of genome sequence data for Listeria phages from food processing ecosystems. We report the genome sequences of two Listeria phages from turkey processing facilities in the USA.

Mozzarella cheese in Italy: Characteristics and occurrence of Listeria monocytogenes and coagulase-positive staphylococci at retail

Mozzarella cheese is a popular type of fresh cheese, originating from Mediterranean countries and worldwide consumed. As for other soft cheeses, its characteristics might support the growth of Listeria monocytogenes; therefore, the research's aim was to determine the occurrence of L. monocytogenes strains and Listeria spp. on mozzarella cheese at the time of purchase (T0) and at the end of shelf life (Tf). Given the relevance in dairy products, occurrence of coagulase-positive staphylococci was also explored. Retail batches (n = 106) collected in Italy, in supermarkets, discounts and small retailers, were analysed for pH and to detect L. monocytogenes, other Listeria spp. and coagulase-positive staphylococci. Three batches were positive for L. monocytogenes (2.8%; 3/106) and nine (9/106) for other species of Listeria spp. with an occurrence of 10.4% for Listeria spp. Coagulase-positive staphylococci were never detected. Mozzarella cheese showed variability in terms of pH (from 4.30 to 6.24 at T0) and residual shelf life (from 3 to 22 days). This research confirms that L. monocytogenes can be found in mozzarella cheese at retail. However, a low concentration of the pathogen (below the limit of quantification of 10 cfu g−1) was found in the contaminated products at the time of purchase and/or at the end of shelf life. This evidence is relevant since mozzarella cheese is not undergoing further treatment before consumption therefore thus represent the level of contamination to which the consumer might be exposed. Considering the technological steps of mozzarella production, preventing food contamination in food-processing and retail-handling environments is crucial to reduce the level of exposure to the consumer.

Virulence Potential and Antimicrobial Resistance of Listeria monocytogenes Isolates Obtained from Beef and Beef-Based Products Deciphered Using Whole-Genome Sequencing

Listeria monocytogenes is a ubiquitous bacterial pathogen that threatens the food chain and human health. In this study, whole-genome sequencing (WGS) was used for the genomic characterization of L. monocytogenes (n = 24) from beef and beef-based products. Multilocus Sequence Type (MLST) analysis revealed that ST204 of CC204 was the most common sequence type (ST). Other sequence types detected included ST1 and ST876 of CC1, ST5 of CC5, ST9 of CC9, ST88 of CC88, ST2 and ST1430 of CC2, and ST321 of CC321. Genes encoding for virulence factors included complete LIPI-1 (pfrA-hly-plcA-plcB-mpl-actA) from 54% (13/24) of the isolates of ST204, ST321, ST1430, and ST9 and internalin genes inlABC that were present in all the STs. All the L. monocytogenes STs carried four intrinsic/natural resistance genes, fosX, lin, norB, and mprF, conferring resistance to fosfomycin, lincosamide, quinolones, and cationic peptides, respectively. Plasmids pLGUG1 and J1776 were the most detected (54% each), followed by pLI100 (13%) and pLM5578 (7%). The prophage profile, vB_LmoS_188, was overrepresented amongst the isolates, followed by LP_101, LmoS_293_028989, LP_030_2_021539, A006, and LP_HM00113468. Listeria genomic island 2 (LGI-2) was found to be present in all the isolates, while Listeria genomic island 3 (LGI-3) was present in a subset of isolates (25%). The type VII secretion system was found in 42% of the isolates, and sortase A was present in all L. monocytogenes genomes. Mobile genetic elements and genomic islands did not harbor any virulence, resistance, or environmental adaptation genes that may benefit L. monocytogenes. All the STs did not carry genes that confer resistance to first-line antibiotics used for the treatment of listeriosis. The characterization of L. monocytogenes in our study highlighted the environmental resistance and virulence potential of L. monocytogenes and the risk posed to the public, as this bacterium is frequently found in food and food processing environments.

Biofilms, mobile genetic elements and the persistence of pathogens on environmental surfaces in healthcare and food processing environments.

Biofilms are the natural state for bacterial and fungal species. To achieve surface hygiene in commercial facilities, the presence of biofilms must be adequately considered. However, standard disinfectant and sanitizer efficacy tests required by the US-EPA and the European Committee for Standardization (CEN) do not currently consider the role of environmental biofilms. This selective review will discuss what biofilms are and why they are important. We will also cover where they are commonly found in healthcare and food processing facilities and explore how current antimicrobial test methods required for product registration do not test for the presence of biofilms. Additionally, we will explore how a lack of efficacy against biofilms may play a role in the development of antimicrobial resistance in healthcare facilities due to the exchange of mobile genetic elements that occur readily in a biofilm matrix.

Antimicrobial Resistance of Listeria monocytogenes Strains Isolated in Food and Food-Processing Environments in Italy

Listeria monocytogenes, along with various other pathogenic bacteria, may show resistance against a broad spectrum of antibiotics. Evaluating the extent of resistance in harmful microorganisms like Listeria monocytogenes holds significant importance in crafting novel therapeutic strategies to mitigate or combat the rise of infections stemming from antibiotic-resistant bacteria. The present work aims to investigate the occurrence of antimicrobial resistance among Listeria monocytogenes strains in meat products (n = 173), seafood (n = 54), dairy products (n = 19), sauces (n = 2), confectionary products (n = 1), ready-to-eat rice dishes (n = 1), and food-processing environments (n = 19). A total of 269 Listeria monocytogenes strains belonging to eight different serovars were tested against 10 antimicrobials. In the classes of antibiotics, most of the strains were resistant antibiotics belonging to the family of β-lactams (92.94%). High proportions of L. monocytogenes isolates were resistant to oxacillin (88.48%), followed by fosfomycin (85.87%) and flumenique (78.44%). The lowest level of resistance was observed against gentamycin (1.49%). A total of 235 strains (n = 87.36%) showed a profile of multidrug resistance. In conclusion, a high occurrence of resistant and multidrug-resistant strains of Listeria monocytogenes was observed among the examined serotypes isolated from different food sources. This understanding enables the adoption of suitable measures to avert contamination and the spread of resistant bacteria via food.

Impact of surface topography and hydrodynamic flow conditions on single and multispecies biofilm formation by Escherichia coli O157:H7 and Listeria monocytogenes in presence of promotor bacteria

Biofilm formation of E. coli O157:H7 and L. monocytogenes in the presence of promotor bacteria R. insidiosa was evaluated on equipment surfaces under different hydrodynamic flow conditions in a Centers for Disease Control and Prevention biofilm reactor (CBR). The CBR containing 330 ml bacterial suspension (6 log CFU/ml in 10% TSB) was used to grow the biofilms on SS 316 L, PTFE, Polycarbonate and EPDM rubber surface coupons under 100, 200 and 300 rpm (shear stresses 0.368, 1.218 and 2.462 N/m2, respectively) for 24 h in batch flow and additional 24 h in continuous flow conditions. Surface roughness and topography of coupons were measured by electron microscopy. Bacterial populations in biofilms were determined by scrapping of coupon surfaces and plating on selective media. Multispecies E. coli O157:H7 biofilm formations were affected by the surface and shear stress. SS, PTFE, and PC exhibited a significant increased biofilm growth at higher shear stress of 2.462 N/m2 compared to 0.368 N/m2. Multispecies L. monocytogenes biofilms formation decreased with increasing shear stress on all the surface materials. Our study emphasizes the importance of surface skewness and kurtosis besides surface's roughness while selecting the material for food processing environments.

Core-shell nanofibers based on microalgae proteins/alginate complexes for enhancing survivability of probiotics

In this study, a novel one-step coaxial electrospinning process is employed to fabricate shell-core structure fibers choosing Chlorella pyrenoidosa proteins (CP) as the core material. These nanofibers, serving as the wall material for probiotic encapsulation, aimed to enhance the stability and antioxidant activity of probiotics in food processing, storage, and gastrointestinal environments under sensitive conditions. Morphological analysis was used to explore the beads-on-a-string morphology and core-shell structure of the electrospun fibers. Probiotics were successfully encapsulated within the fibers (7.97 log CFU/g), exhibiting a well-oriented structure along the distributed fibers. Compared to free probiotics and uniaxial fibers loaded with probiotics, encapsulation within microalgae proteins/alginate core-shell structure nanofibers significantly enhanced the probiotic cells' tolerance to simulated gastrointestinal conditions (p < 0.05). Thermal analysis indicated that microalgae proteins/alginate core-shell structure nanofibers displayed superior thermal stability compared to uniaxial fibers. The introduction of CP resulted in a 50 % increase in the antioxidant capacity of probiotics-loaded microalgae proteins/alginate nanofibers compared to uniaxial alginate nanofibers, with minimal loss of viability (0.8 log CFU/g) after 28 days of storage at 4 °C. In summary, this dual-layer carrier holds immense potential in probiotic encapsulation and enhancing their resistance to harsh conditions.

Listeria monocytogenes from Food Products and Food Associated Environments: Antimicrobial Resistance, Genetic Clustering and Biofilm Insights.

Listeria monocytogenes, a foodborne pathogen, exhibits high adaptability to adverse environmental conditions and is common in the food industry, especially in ready-to-eat foods. L. monocytogenes strains pose food safety challenges due to their ability to form biofilms, increased resistance to disinfectants, and long-term persistence in the environment. The aim of this study was to evaluate the presence and genetic diversity of L. monocytogenes in food and related environmental products collected from 2014 to 2022 and assess antibiotic susceptibility and biofilm formation abilities. L. monocytogenes was identified in 13 out of the 227 (6%) of samples, 7 from food products (meat preparation, cheeses, and raw milk) and 6 from food-processing environments (slaughterhouse-floor and catering establishments). All isolates exhibited high biofilm-forming capacity and antibiotic susceptibility testing showed resistance to several classes of antibiotics, especially trimethoprim-sulfamethoxazole and erythromycin. Genotyping and core-genome clustering identified eight sequence types and a cluster of three very closely related ST3 isolates (all from food), suggesting a common contamination source. Whole-genome sequencing (WGS) analysis revealed resistance genes conferring resistance to fosfomycin (fosX), lincosamides (lin), fluoroquinolones (norB), and tetracycline (tetM). In addition, the qacJ gene was also detected, conferring resistance to disinfecting agents and antiseptics. Virulence gene profiling revealed the presence of 92 associated genes associated with pathogenicity, adherence, and persistence. These findings underscore the presence of L. monocytogenes strains in food products and food-associated environments, demonstrating a high virulence of these strains associated with resistance genes to antibiotics, but also to disinfectants and antiseptics. Moreover, they emphasize the need for continuous surveillance, effective risk assessment, and rigorous control measures to minimize the public health risks associated to severe infections, particularly listeriosis outbreaks. A better understanding of the complex dynamics of pathogens in food products and their associated environments can help improve overall food safety and develop more effective strategies to prevent severe health consequences and economic losses.

Characterization and tolerance of foodborne pathogenic bacteria in microplastic biofilm

Microplastics have the ability to selectively accumulate microorganisms from their surrounding environments, leading to the formation of biofilms. The potential for microplastic biofilm in food processing environments to contaminate the processing environment and food matrices poses a threat to food safety. This study validated the biofilm formation of three commonly encountered foodborne pathogens, Salmonella enterica serovar Typhimurium (ST), Staphylococcus aureus (SA), and Listeria monocytogenes (LM) on microplastics, and investigated their tolerance to disinfectant and gastrointestinal tract. Three strains showed distinct biofilm formation patterns, with biomass reaching a steady state at 2 d. Meanwhile, the particle size of microplastics exhibited a specific impact on biofilm formation, with smaller microplastics sizes leading to increased biofilm formation. Sodium hypochlorite (NaClO) can effectively eliminate most biofilm-forming bacteria, except for ST. After treatment with 50 ppm NaClO, 34% cells in ST biofilm survived. Only SA survived in simulated gastric fluid, with around 3 log reduction after 1 h. ST and LM were not detected. All three strains showed a slight increase after treatment with simulated intestinal fluid for 6 h. These results highlighted the capacity of microplastics to transport pathogens through biofilms, emphasizing the potential synergistic effects between microplastics and pathogens in food-associated environments.

Phenotypic and proteomic differences in biofilm formation of two Lactiplantibacillus plantarum strains in static and dynamic flow environments

Lactiplantibacillus plantarum is a Gram-positive non-motile bacterium capable of producing biofilms that contribute to the colonization of surfaces in a range of different environments. In this study, we compared two strains, WCFS1 and CIP104448, in their ability to produce biofilms in static and dynamic (flow) environments using an in-house designed flow setup. This flow setup enables us to impose a non-uniform flow velocity profile across the well. Biofilm formation occurred at the bottom of the well for both strains, under static and flow conditions, where in the latter condition, CIP104448 also showed increased biofilm formation at the walls of the well in line with the higher hydrophobicity of the cells and the increased initial attachment efficacy compared to WCFS1. Fluorescence and scanning electron microscopy showed open 3D structured biofilms formed under flow conditions, containing live cells and ∼30 % damaged/dead cells for CIP104448, whereas the WCFS1 biofilm showed live cells closely packed together. Comparative proteome analysis revealed minimal changes between planktonic and static biofilm cells of the respective strains suggesting that biofilm formation within 24 h is merely a passive process. Notably, observed proteome changes in WCFS1 and CIP104448 flow biofilm cells indicated similar and unique responses including changes in metabolic activity, redox/electron transfer and cell division proteins for both strains, and myo-inositol production for WCFS1 and oxidative stress response and DNA damage repair for CIP104448 uniquely. Exposure to DNase and protease treatments as well as lethal concentrations of peracetic acid showed highest resistance of flow biofilms. For the latter, CIP104448 flow biofilm even maintained its high disinfectant resistance after dispersal from the bottom and from the walls of the well. Combining all results highlights that L. plantarum biofilm structure and matrix, and physiological state and stress resistance of cells is strain dependent and strongly affected under flow conditions. It is concluded that consideration of effects of flow on biofilm formation is essential to better understand biofilm formation in different settings, including food processing environments.

Covalent conjugate of pea protein induced by cyanidin-3-O-glucoside quinone: The structural formation and functional properties

The unique structural of anthocyanins makes them vulnerable to degradation within food processing environments. Proteins commonly coexist with polyphenols in natural food matrices on a non-covalent and covalent basis, serving to stabilize protection for polyphenols. Here, three natural pea-extracted proteins, including albumin, vicilin, and legumin were bound to cyanidin-3-O-glucoside (C3G), and binding modes, conjugate structures and functions were characterized. Isothermal titration calorimetry showed a strongly exothermic interaction between C3G and proteins. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fourier transform infrared spectroscopy revealed that the C3G-quinone (C3GQ) formed by C3G oxidation established covalent cross-links with C–S and C–N in the amino acid residues of proteins, and could form more large molecular weight aggregates at an addition amount of 1 mM. The free sulfhydryl and amino contents of the three protein conjugates decreased with the addition of C3GQ. Due to the presence of 4–5 times more sulfhydryl groups in albumin as compared to globulin, albumin showed a 20% higher binding capacity for C3GQ than globulin. This consequently led to the highest antioxidant activity in the C3GQ-albumin conjugate. In contrast to native pea proteins, the introduction of covalent modifications induces changes in the secondary structure of the pea proteins. Specifically, the structure of albumin tends to exhibit increased orderliness, while the molecular flexibility of globular proteins was augmented. Thus, the conjugates have enhanced thermal stability. Our results affirmed that the stability of C3G can be more effectively improved through designing it to covalently interact with proteins relative to non-covalent interactions. This study aimed at offering novel theoretical underpinnings for the functional utilization of C3G and pea proteins in the food industry.

Prevalence of Staphylococcus argenteus among food handlers, kitchen utensils, and food samples in Japan.

Staphylococcus argenteus has received increased attention from an aspect of food safety since several food poisoning outbreaks caused by the bacterium were reported in Japan. However, S. argenteus prevalence among food handlers and utensils has not yet been investigated. In this study, we investigated S. argenteus prevalence among a collection of coagulase-positive staphylococci (CPS) that were isolated during food sanitary inspections in Japan. Out of a total of 191 CPS isolates, 14 were identified as S. argenteus. One was isolated from shelled shrimp, nine were isolated from food handlers' hand swabs, and four were isolated from kitchen utensils. Whole-genome sequencing revealed that transmission of S. argenteus from human hands to utensils was possible. Though all 14 isolates were negative for the pvl and tst-1 genes, 6 harbored the seb gene. Only 21.4% of S. argenteus isolates were resistant to antibiotics, while 62.1% of the S. aureus isolates from the same sources were confirmed to be resistant. To the best of our knowledge, this is the first report to demonstrate possible transmission of S. argenteus from food handlers to utensils in food-processing environments.

Virulence properties of Listeria monocytogenes isolated from meat and meat contact surfaces in a slaughterhouse.

Listeria monocytogenes is a ubiquitous microorganism that is isolated from a variety of sources such as soil, water, decaying vegetation, sewage, animal feeds, silage, farm environments and food-processing environments. This study aimed to determine the prevalence, serogroups, biofilm formation, virulence factor genes, and genetic relationships of L. monocytogenes strains isolated from beef meat and meat contact surfaces obtained from a slaughterhouse in Burdur, Turkey. In this study, a total of 179 beef meat and meat contact surface samples were analyzed for the presence of L. monocytogenes by polymerase chain reaction (PCR). Out of a total of 179 beef meat and meat contact surface samples, 83 (46.37%) were found to be contaminated with L. monocytogenes, with the highest incidence (53.01%) occurring in beef meat. In the present study, most of the isolated strains belonged to serogroups IIB and IVB (lineage I). The L. monocytogenes strain also contained monoA-B, prfA, plcA, plcB, mpl, hlyA, actA, gtcA, dltA, Fri, flaA, InlA, InlC, InlJ, and iap genes. Biofilm formation was not determined in the tested samples at pH 5.5 and different temperatures (4°C, 10°C, 25°C, and 37°C). However, strong biofilm formation was observed in 6.45% (2/31) of the strains at pH 7.0 after 48 h incubation at 37°C, and in 3.22% (1/31) of the strains at pH 7.0 after 48 h incubation at 4°C and 10°C. Pulsed-field gel electrophoresis (PFGE) results showed that L. monocytogenes isolates were clonally related, and cross-contamination was present. In addition, PFGE results also revealed that AscI had more distinguishing power than the ApaI restriction enzyme. These results indicate that L. monocytogenes detected from meat and meat contact surfaces in the slaughterhouse pose a potential risk to public health.

The impact of different acidic conditions and food substrates on Listeria monocytogenes biofilms development and removal using nanoencapsulated carvacrol

Listeria monocytogenes biofilms present a significant challenge in the food industry. This study explores the impact of different acidic conditions of culture media and food matrices on the development and removal of biofilms developed on stainless steel surfaces by wild-type (WT) L. monocytogenes strains as well as in two mutant derivatives, ΔsigB and ΔagrA, that have defects in the general stress response and quorum sensing, respectively. Additionally, the study investigates the efficacy of nanoencapsulated carvacrol as an antimicrobial against L. monocytogenes biofilms developed in Tryptic Soy Broth (TSB) culture media acidified to different pH conditions (3.5, 4.5, 5.5, 6.5), and in food substrates (apple juice, strained yogurt, vegetable soup, semi-skimmed milk) having the same pH levels. No biofilm formation was observed for all L. monocytogenes strains at pH levels of 3.5 and 4.5 in both culture media and food substrates. However, at pH 5.5 and 6.5, increased biofilm levels were observed in both the culture media and food substrates, with the WT strain showing significantly higher biofilm formation (3.04–6.05 log CFU cm−2) than the mutant strains (2.30–5.48 log CFU cm−2). For both applications, the nanoencapsulated carvacrol demonstrated more potent antimicrobial activity against biofilms developed at pH 5.5 with 2.23 to 3.61 log reductions, compared to 1.58–2.95 log reductions at pH 6.5, with mutants being more vulnerable in acidic environments. In food substrates, nanoencapsulated carvacrol induced lower log reductions (1.58–2.90) than the ones in TSB (2.02–3.61). These findings provide valuable insights into the impact of different acidic conditions on the development of L. monocytogenes biofilms on stainless steel surfaces and the potential application of nanoencapsulated carvacrol as a biofilm control agent in food processing environments.

Screening of E. coli O157:H7 AI-2 QS inhibitors and their inhibitory effect on biofilm formation in combination with disinfectants

Food processing environments contaminated with E. coli O157:H7 and forming biofilms will be a continuous source of contamination, leading to foodborne illness and food spoilage. Now, the main countermeasure is chemical disinfectants, but the use of disinfectants may cause the development of drug-resistant bacteria and other negative effects. Therefore, there is an urgent need to find novel antimicrobial agents. Quorum-sensing inhibitors (QSI) are a promising strategy to control biofilm formation. This study aimed to screen the optimal QSI of E. coli O157:H7 from plant-derived small molecules by biofilm and QS inhibitory activities and to validate their effectiveness in combination with disinfectants. In addition, the inhibition mechanism of gallic acid on biofilm formation was further explored. Gallic acid was screened as the QSI of E. coli O157:H7, and its AI-2 inhibitory activity against LuxS/AI-2 QS system reached 95.54% and 98.89% after treatment with 1/2 MIC gallic acid for 4 h and 6 h, respectively. It inhibited biofilm formation by affecting flagellar assembly, chemotaxis, and expression of adhesion factor-related genes, which interfered with the transition of bacteria from reversible to irreversible adhesion. In beef broth simulation system, the inhibition rate was greater than 90.00% in all gallic acid and benzalkonium chloride combination groups. Therefore, gallic acid can be a potential antimicrobial synergist in the food industry, combined with disinfectants to control E. coli O157:H7 biofilms while avoiding the development of resistance.

Protein-Based Rechargeable and Replaceable Antimicrobial and Antifouling Coatings on Hydrophobic Food-Contact Surfaces.

The growing concerns regarding foodborne illnesses related to fresh produce accentuate the necessity for innovative material solutions, particularly on surfaces that come into close contact with foods. This study introduces a sustainable, efficient, and removable antimicrobial and antifouling coating ideally suited for hydrophobic food-contact surfaces such as low-density polyethylene (LDPE). Developed through a crosslinking reaction involving tannic acid, gelatin, and soy protein hydrolysate, these coatings exhibit proper stability in aqueous washing solutions and effectively combat bacterial contamination and prevent biofilm formation. The unique surface architecture promotes the formation of halamine structures, enhancing antimicrobial efficacy with a rapid contact killing effect and reducing microbial contamination by up to 5 log10 cfu·cm-2 against both Escherichia coli (Gram-negative) and Listeria innocua (Gram-positive). Notably, the coatings are designed for at least five recharging cycles under mild conditions (pH6, 20 ppm free active chlorine) and can be easily removed with hot water or steam to refresh the depositions. This removal process not only conveniently aligns with existing sanitation protocols in the fresh produce industry but also facilitates the complete eradication of potential developed biofilms, outperforming uncoated LDPE coupons. Overall, these coatings represent sustainable, cost-effective, and practical advancements in food safety and are promising candidates for widespread adoption in food processing environments.

Anti-Listerial Activity of Bacteriocin-like Inhibitory Substance Produced by Enterococcus lactis LBM BT2 Using Alternative Medium with Sugarcane Molasses.

Listeria monocytogenes is a foodborne pathogen that contaminates food-processing environments and persists within biofilms on equipment, thus reaching final products by cross-contamination. With the growing demand for clean-label products, the search for natural antimicrobials as biopreservants, such as bacteriocins, has shown promising potential. In this context, this study aimed to evaluate the anti-listerial action of bacteriocins produced by Enterococcus lactis LBM BT2 in an alternative medium containing sugarcane molasses (SCM). Molecular analyses were carried out to characterize the strain, including the presence of bacteriocin-related genes. In the kinetic study on SCM medium E. lactis, LBM BT2 showed biomass and bacteriocin productions similar to those observed on a sucrose-based medium (control), highlighting the potential of the sugarcane molasses as a low-cost substrate. Stability tests revealed that the molecule remained active in wide ranges of pH (4-10) and temperature (60-100 °C). Furthermore, the proteolytic treatment reduced the biomolecule's antimicrobial activity, highlighting its proteinaceous nature. After primary purification by salting out and tangential flow filtration, the bacteriocin-like inhibitory substance (BLIS) showed bacteriostatic activity on suspended L. monocytogenes cells and against biofilm formation at a concentration of 0.625 mg/mL. These results demonstrate the potential of the produced BLIS as a biopreservative in the food industry.