Foodborne Pathogens Research Articles

Development and characterization of sodium alginate and β-cyclodextrin nanoemulsions encapsulating betel leaf (Piper betle L.) extract for enhanced antimicrobial efficacy against foodborne pathogen

This study aims to investigate the physical stability, droplet size, zeta potential, and antimicrobial properties of nanoemulsions formulated with betel leaf extract using β-cyclodextrin (CD) and sodium alginate (SA) biopolymers. Nanoemulsions with β-cyclodextrin exhibit superior stability at lower temperatures, with limited droplet size, and strong electrostatic repulsion. Morphological images demonstrate the successful encapsulation of betel leaf extract within both biopolymers, highlighting their potential for antimicrobial applications. Both CD and SA nanoemulsions display inhibitory effects on bacterial strains (E. coli, P. aeruginosa, L. monocytogenes, S. aureus, and B. cereus) and fungal growth (A. brasiliensis, R. stolonifer, F. oxysporum, and C. albicans). SA nanoemulsions show higher antimicrobial activity due to H+ ion release, particularly against A. brasiliensis and C. albicans. These findings underscore the potential of betel leaf extract nanoemulsions, especially those with SA, for various antimicrobial applications for sustainable food packaging, highlighting their significance in addressing microbial challenges.

Influence of fractionation of polyphenols by membrane chromatography on antioxidant, antimicrobial and proliferation-inhibiting effects of red fruit juices

Interest in fruit juice extracts as nutraceuticals is constantly increasing due to their health-beneficial properties, mainly caused by polyphenols. However, the correlation between the various effects of fruit juice extracts and their individual composition, including anthocyanins and copigments, is unknown. Therefore, in the present study, eight red fruit juice extracts were prepared using XAD-7 column chromatography, followed by fractionation and identification of the different compounds as well as characterization of their health-promoting effects. The fruit juice extract of pomegranate, chokeberry, and cranberry showed the highest antimicrobial potential against food-borne pathogens. The highest antioxidant and cell proliferation-inhibiting potential was also found in the pomegranate extract. It can be assumed that pomegranate extracts, which are rich in copigments, especially hydrolyzable tannins, are suitable natural antioxidants and antimicrobial agents. Pomegranate extracts could be used as nutraceuticals or natural preservatives.

Biantennary N-glycans As Receptors for MARTX Toxins in Vibrio Pathogenesis.

Multifunctional Autoprocessing Repeats-in-Toxin (MARTX) toxins are a diverse effector delivery platform of many Gram-negative bacteria that infect mammals, insects, and aquatic animal hosts. The mechanisms by which these toxins recognize host cell receptors for translocation of toxic effectors into the cell have remained elusive. Here, we map the first surface receptor-binding domain of a MARTX toxin from the highly lethal foodborne pathogen Vibrio vulnificus . This domain corresponds to a 273-amino acid sequence with predicted symmetrical immunoglobulin-like folds. We demonstrate that this domain binds internal N -acetylglucosamine on complex biantennary N-glycans with select preference for L1CAM and other N-glycoproteins with multiple N-glycans on host cell surfaces. This receptor binding domain is essential for V. vulnificus pathogenesis during intestinal infection. The identification of a highly conserved motif universally present as part of all N-glycans correlates with the V. vulnificus MARTX toxin boasting broad specificity and targeting nearly all cell types.

Development and application of a bacteriophage cocktail for Shigella flexneri biofilm inhibition on the stainless steel surface

Food contamination and biofilm formation by Shigella in food processing facilities are major causes of acute gastrointestinal infection and mortality in humans. Bacteriophages (phages) are promising alternatives to antibiotics in controlling plankton and biofilms in food matrices. This study isolated two novel phages, S2_01 and S2_02, with lytic activity against various Shigella spp. From sewage samples. Transmission electron microscopy revealed that phages S2_01 and S2_02 belonged to the Caudovirales order. On characterizing their lytic ability, phage S2_01 initially exhibited relatively weak antibacterial activity, while phage S2_02 initially displayed rapid antibacterial activity after phage application. A combination of these phages in a 1:9 ratio was selected, as it has been suggested to elicit the most rapid and sustained lysis ability for up to 24 h. It demonstrated lytic activity against various foodborne pathogens, including six Shigella spp. The phage cocktail exhibited biofilm inhibition and disruption abilities of approximately 79.29% and 42.55%, respectively, after 24 h in a 96-well microplate. In addition, inhibition (up to 23.42%) and disruption (up to 19.89%) abilities were also observed on stainless steel surfaces, and plankton growth was also significantly suppressed. Therefore, the phage cocktail formulated in this study displays great potential as a biological control agent in improving food safety against biofilms and plankton.

Genomic insights into prevalence of virulence and multi drug resistance genes in milk borne Klebsiella pnuemoniae: Face of emerging resistance to last resort antibiotics

Spread of hypervirulent and multi-drug resistant Klebsiella pneumoniae in raw milk is public health concern due to its potential impact on food safety and public health. Therefore, this study investigated antibiotic susceptibility test (AST), antibiotic resistance genes (ARGs), mutations conferring ARGs, virulence factor and plasmid replicons to check prevalence of fosfomycin resistant MDR K. pneumoniae isolated from raw milk samples collected from Saurashtra region of Gujarat, India. K. pneumoniae isolated from raw milk and subjected to disk diffusion assay. From that, MDR along with fosfomycin resistant isolates were analysed for multi locus sequence typing, presence of ARGs, mutations conferring resistance, virulence factors and plasmid replicon types by using its whole genome sequence. Results shows that, among 32 K. pneumoniae, 8 were phenotypically resistant to fosfomycin. As per WGS analysis, 8 MDR isolates were assigned into different sequence types such as ST3321, ST37, ST2715, ST1087, ST3157, ST299 and ST29. Among that, ST37 is well recognized MDR high risk clone reported worldwide and first time reported from raw milk of Saurashtra region of Gujarat, India. ARGs responsible for resistance to fosfomycin (fosA) were found in all 8 isolates. Other ARGs such as blaSHV, kdeA, OqxA, OqxB, dfrA1, sul1, qnrB4, aadA2 and ere(A) were also detected. High diversity of virulence factors was also identified by detection of genes encoding virulence factors related to iron uptake such as entE, fepD, entA, entB, Irp2, fepG, ybtU, ybtP, fepC, ybtA, ybtE, fepB, ybtS, fyuA, ybtQ, ybtT, ybtX, Irp1, adherence such as yagZ/ecpA, yagV/ecpE, yagX/ecpC, yagV/ecpE, ykgK/ecpR and invasion such as fimA, pla, fimC, fimH, fimB, fimE were detected in eight genomes. Mutations in murA, uhpT and glpT conferring a fosfomycin resistance were also present in genomes of 8 K. pneumoniae. IncF was the most common plasmid replicon type detected in all 8 genomes. The study reports high diversity of virulent and multidrug resistant K. pneumoniae in raw milk. Hence, genomic surveillance plans are urgently required for food borne pathogens.

CRISPR/Cas12a and Hybridization Chain Reaction-Coregulated Magnetic Relaxation Switching Biosensor for Sensitive Detection of Viable Salmonella in Animal-Derived Foods.

We combined a CRISPR/Cas12a system with a hybridization chain reaction (HCR) to develop an ultrasensitive magnetic relaxation switching (MRS) biosensor for detecting viable Salmonella typhimurium (S. typhimurium). Magnetic nanoparticles of two sizes (30 and 1000 nm: MNP30 and MNP1000, respectively) were coupled through HCR. The S. typhimurium gene-activated CRISPR/Cas12a system released MNP30 from the MNP1000-HCR-MNP30 complex through a trans-cleavage reaction. After magnetic separation, released MNP30 was collected from the supernatant and served as a transverse relaxation time (T2) signal probe. Quantitative detection of S. typhimurium is achieved by establishing a linear relationship between the change in T2 and the target gene. The biosensor's limit of detection was 77 CFU/mL (LOD = 3S/M, S = 22.30, M = 0.87), and the linear range was 102-108 CFU/mL. The accuracy for detecting S. typhimurium in real samples is comparable to that of qPCR. Thus, this is a promising method for the rapid and effective detection of foodborne pathogens.

Integrating lateral flow device with controllable gold in situ growth for sensitive detection of staphylococcal enterotoxin A in milk

Gold nanoparticle-based lateral flow immunoassays (AuNP-LFIA) are widely used for pathogen monitoring to prevent foodborne illness outbreaks. However, conventional AuNP-LFIA exhibits poor sensitivity and limited quantitative capacity due to the low colorimetric signal intensity of AuNPs. Herein, we introduced a low-background gold in situ growth (GISG) strategy by lowering the pH of the growth solution to weaken the reducibility of hydroxylamine, thereby enhancing the sensitivity of AuNP-LFIA. Additionally, we developed a universal and manufacturable lateral flow device to streamline the GISG process. We applied this device to detect staphylococcal enterotoxin A (SEA), an exotoxin produced by Staphylococcus aureus. Under optimal conditions, the proposed device demonstrated superior practicality and excellent sensitivity for SEA detection, achieving a detection limit of 0.061 ng/mL with the total detection time of 37 min, showing 311 times more sensitive than the unamplified AuNP-LFIA. Furthermore, SEA detection in milk samples showed a strong correlation (R2 = 0.8845) with results obtained from a conventional ELISA kit. Therefore, this promising LFIA device offers a novel strategy with high sensitivity and practicality for in-field detection of Staphylococcus aureus and can be easily adapted for screening other foodborne pathogens.

Determination of the combined effect of grape seed extract and cold atmospheric plasma on foodborne pathogens and their environmental stress knockout mutants.

The study aimed to explore the antimicrobial efficacy of grape seed extract (GSE) and cold atmospheric plasma (CAP) individually or in combination against L. monocytogenes and E. coli wild type (WT) and their isogenic mutants in environmental stress genes. More specifically, we examined the effects of 1% (wt/vol) GSE, 4 min of CAP treatment, and their combined effect on L. monocytogenes 10403S WT and its isogenic mutants ΔsigB, ΔgadD1, ΔgadD2, ΔgadD3, as well as E. coli K12 and its isogenic mutants ΔrpoS, ΔoxyR, and ΔdnaK. In addition, the sequence of the combined treatments was tested. A synergistic effect was achieved for all L. monocytogenes strains when exposure to GSE was followed by CAP treatment. However, the same effect was observed against E. coli strains, only for the reversed treatment sequence. Additionally, L. monocytogenes ΔsigB was more sensitive to the individual GSE and the combined GSE/CAP treatment, whereas ΔgadD2 was more sensitive to CAP, as compared to the rest of the mutants under study. Individual GSE exposure was unable to inhibit E. coli strains, and individual CAP treatment resulted in higher inactivation of E. coli in comparison to L. monocytogenes with the strain ΔrpoS appearing the most sensitive among all studied strains. Our findings provide a step toward a better understanding of the mechanisms playing a role in the tolerance/sensitivity of our model Gram-positive and Gram-negative bacteria toward GSE, CAP, and their combination. Therefore, our results contribute to the development of more effective and targeted antimicrobial strategies for sustainable decontamination.IMPORTANCEAlternative approaches to conventional sterilization are gaining interest from the food industry, driven by (i) the consumer demand for minimally processed products and (ii) the need for sustainable, environmentally friendly processing interventions. However, as such alternative approaches are milder than conventional heat sterilization, bacterial pathogens might not be entirely killed by them, which means that they could survive and grow, causing food contamination and health hazards. In this manuscript, we performed a systematic study of the impact of antimicrobials derived from fruit industry waste (grape seed extract) and cold atmospheric plasma on the inactivation/killing as well as the damage of bacterial pathogens and their genetically modified counterparts, for genes linked to the response to environmental stress. Our work provides insights into genes that could be responsible for the bacterial capability to resist/survive those novel treatments, therefore, contributing to the development of more effective and targeted antimicrobial strategies for sustainable decontamination.

Sustainable Recovery of Polyphenols and Carotenoids from Horned Melon Peel via Cloud Point Extraction.

Using natural plant extracts as food additives is a promising approach for improving food products' quality, nutritional value, and safety, offering advantages for both consumers and the environment. Therefore, the main goal of this study was to develop a sustainable method for extracting polyphenols and carotenoids from horned melon peel using the cloud point extraction (CPE) technique, intending to utilize it as a natural food additive. CPE is novel promising extraction method for separation and pre-concentration of different compounds while being simple, inexpensive, and low-toxic. Three parameters within the CPE approach, i.e., pH, equilibrium temperature, and equilibrium time, were investigated as independent variables through the implementation of Box-Behnken design and statistical analyses. The optimized conditions for the maximum recovery of both polyphenols and carotenoids, reaching 236.14 mg GAE/100 g and 13.80 mg β carotene/100 g, respectively, were a pH value of 7.32, an equilibrium temperature of 55 °C, and an equilibrium time of 43.03 min. The obtained bioactives' recovery values under the optimized conditions corresponded to the predicted ones, indicating the suitability of the employed RSM model. These results highlight the effectiveness of CPE in extracting bioactive compounds with varying polarities from agricultural by-products, underscoring its potential for enhancing the value of food waste and advancing sustainable practices in food processing. According to microbiological food safety parameters, the optimal CPE extract is suitable for food applications, while its storage under refrigerated and dark conditions is particularly beneficial. The CPE extract's enhanced stability under these conditions makes it a more viable option for long-term storage, preserving both safety and quality.

Effects of X‐ray irradiation on pathogen inactivation, quality changes, and hydrocarbon profiles of pork cutlets

SummaryPork cutlets are susceptible to cross‐contamination with foodborne pathogens during processing. Food irradiation effectively improves food hygiene by eliminating microbes. We investigated the application of X‐ray irradiation (0.5–10 kGy) on pathogen inactivation (D10 value), quality changes [total aerobic bacteria (TAB), yeast and mould (Y&M), total coliform (TC), colour, thiobarbituric acid reactive substance (TBARS), and pH], and the hydrocarbon profile of pork cutlets. D10 values of Escherichia coli O157:H7, Salmonella typhimurium, Listeria monocytogenes, and Staphylococcus aureus were 0.37, 0.62, 0.52, and 0.36 kGy, respectively. At least 3.10 kGy (5‐D10 values) is required to ensure the microbiological safety of pork cutlets. X‐ray irradiation dose‐dependently reduced all microbial counts, and up to 7 kGy had no adverse effects on colour, TBARS, and pH. Furthermore, 1,7‐hexadecadiene and 8‐heptadecene were potential markers for irradiated pork cutlets. X‐ray irradiation of up to 7 kGy is effective in improving hygiene levels while maintaining pork cutlet quality.

Conditional expression of flagellar motility, curli fimbriae, and biofilms in Shiga toxin- producing Escherichia albertii.

Escherichia albertii is an emerging foodborne pathogen. We previously reported that some avian Shiga toxin-producing E. albertii strains exhibited higher or comparable cytotoxicity in Vero-d2EGFP cells with several enterohemorrhagic E. coli (EHEC) outbreak strains. To better understand the environmental persistence of this pathogen, comparative genomics and phenotypic assays were applied to assess adhesion capability, motility, and biofilm formation in E. albertii. Among the 108 adherence-related genes, those involved in biogenesis of curli fimbriae, hemorrhagic E. coli pilus, type 1 fimbriae, and Sfm fimbriae were conserved in E. albertii. All 20 E. albertii strains carried a complete set of primary flagellar genes that were organized into four gene clusters, while five strains possessed genes related to the secondary flagella, also known as lateral flagella. Compared to EHEC strain EDL933, the eight chemotaxis genes located within the primary flagellar gene clusters were deleted in E. albertii. Additional deletion of motility genes flhABCD and motBC was identified in several E. albertii strains. Swimming motility was detected in three strains when grown in LB medium, however, when grown in 5% TSB or in the pond water-supplemented with 10% pigeon droppings, an additional four strains became motile. Although all E. albertii strains carried curli genes, curli fimbriae were detected only in four, eight, and nine strains following 24, 48, and 120 h incubation, respectively. Type 1 fimbriae were undetectable in any of the strains grown at 37°C or 28°C. Strong biofilms were detected in strains that produced curli fimbriae and in a chicken isolate that was curli fimbriae negative but carried genes encoding adhesive fimbriae K88, a signature of enterotoxigenic E. coli strains causing neonatal diarrhea in piglets. In all phenotypic traits examined, no correlation was revealed between the strains isolated from different sources, or between the strains with and without Shiga toxin genes. The phenotypic variations could not be explained solely by the genetic diversity or the difference in adherence genes repertoire, implying complex regulation in expression of various adhesins. Strains that exhibited a high level of cytotoxicity and were also proficient in biofilm production, may have potential to emerge into high-risk pathogens.

Identification and characterization of a small molecule BFstatin inhibiting BrpR, the transcriptional regulator for biofilm formation of Vibrio vulnificus.

Many pathogenic bacteria form biofilms that are resistant to not only host immune defenses but also antibiotics, posing a need for the development of strategies to control biofilms. In this study, to prevent biofilm formation of the fulminating foodborne pathogen Vibrio vulnificus, chemical libraries were extensively screened to identify a small molecule inhibiting the activity of BrpR, a transcriptional regulator for biofilm genes. Accordingly, the BrpR inhibitor BFstatin [N1-(2-chloro-5-fluorophenyl)-N3-propylmalonamide], with a half-maximal effective concentration of 8.01 μM, was identified. BFstatin did not interfere with bacterial growth or exhibit cytotoxicity to the human epithelial cell line. BFstatin directly bound to BrpR and interrupted its binding to the target promoter DNAs of the downstream genes. Molecular dynamics simulation of the interaction between BFstatin and BrpR proposed that BFstatin modifies the structure of BrpR, especially the DNA-binding domain. Transcriptomic analyses revealed that BFstatin reduces the expression of the BrpR regulon including the cabABC operon and brp locus which contribute to the production of biofilm matrix of V. vulnificus. Accordingly, BFstatin diminished the biofilm levels of V. vulnificus by inhibiting the matrix development in a concentration-dependent manner. Altogether, BFstatin could be an anti-biofilm agent targeting BrpR, thereby rendering V. vulnificus more susceptible to host immune defenses and antibiotics.

Prevalence and genomic characteristics of becAB-carrying Clostridium perfringens strains

Clostridium perfringens, as a foodborne pathogen, can cause various intestinal diseases in both humans and animals according to its repertoire of toxins. In recent years, a multitude of studies have highlighted its threat to infants and young children. C. perfringens carries numerous toxins, with the newly identified BEC toxin confirmed as the second toxin to cause diarrheal illness, after CPE. However, the global dissemination of C. perfringens strains carrying becAB genes, which encode BEC toxins, has not been extensively studied. Following epidemiological surveillance of the prevalence of C. perfringens from different sources in various provinces of China, we identified two becAB-carrying strains and one strain carrying a sequence similar to becAB from distinct provinces and sources. When combined with genomic analysis of other becAB-carrying C. perfringens strains from public databases, we found that becAB was present in strains from different lineages. Our analysis of the plasmid and genetic environment corroborates previous findings on becAB-carrying strains, confirming that it currently achieves horizontal transmission through one type of evolutionarily conserved Pcp plasmid. This study provides a comprehensive analysis of the prevalence and transmission patterns of the newly emerged toxin gene locus, becAB, in C. perfringens. Despite the relatively low identification rate of becAB-carrying strains, their potential impact requires ongoing surveillance and investigation of their features, particularly their antimicrobial resistance.

Monitoring foodborne pathogens of food served at an institutional cafeteria

The provision of food by cafeterias is an opportunity to promote health through a balanced diet and safe food, offering the prospect of sustainably developing the well-being a community and the local economy. This study assessed the hygiene conditions of an institutional cafeteria. The presence of foodborne pathogens and indicators of general hygiene in 143 food and environmental samples were analyzed using the TEMPO and VIDAS food safety testing machines. All food samples tested negative for Salmonella sp., and Staphylococcus aureus counts were below the GCC Standardization Organization (GSO) limit. Escherichia coli was detected at level above the set limit in six food samples. Sixteen food samples showed aerobic bacterial counts above the set limit. Total coliforms and Enterobacteriaceae were each detected above the set limits in eight food samples. Yeasts and molds were detected above the set limit in 14 food samples. Coliform bacteria, S. aureus, and mold were detected on the hands and white coats of cafeteria workers and on work surfaces, machines, and cutting boards. The hygiene conditions of the cafeteria are not suitable for food preparation, and to improve hygienic conditions in cafeterias, regular inspection of food and food preparation facilities is strongly recommended.

Low potential of fish as a source of infection with Angiostrongylus cantonensis

Abstract Objectives Fish are hypothesized to act as paratenic hosts for the zoonotic nematode Angiostrongylus cantonensis, cause of human eosinophilic meningitis. There is a lack of data confirming the relevance of fish in A. cantonensis life cycle and their contribution to human infection. Materials and methods We conducted a series of experiments to investigate survival and infectivity of A. cantonensis larvae in Clarias gariepinus (catfish; n=30) and Oreochromis niloticus (tilapia; n=24). Each fish was inoculated with 10,000 third-stage larvae (L3). Larval survival was assessed through artificial digestion of fish tissues one, two, and three weeks post-infection. To investigate early stages of infection, four catfish were inoculated with 10,000 L3 each and sacrificed 3 days post-infection. qPCR and histopathological examinations were performed to evaluate larval distribution and tissue reactions. Two infected catfish, sacrificed one week post-infection, were used to feed Wistar rats. Results After 45 days, the rats were not shedding L1, indicating the absence of infection. One week post-infection, dead larvae were present in digested tissues of both fish species, and the same was observed two and three weeks after exposure. qPCR analysis revealed that intestine was the most heavily infected organ. Histopathology identified dead larvae within granulomas in intestines and liver. Early-stage infection experiment showed that fish sacrificed three days post-inoculation contained viable L3 which were infective to Wistar rats. Conclusions While A. cantonensis L3 can survive and remain infective in fish for a short period, they typically die within first few days post-infection. This suggests that fish may not be significant long-term paratenic hosts for A. cantonensis but may play a temporary role in its transmission to mammals (including humans) and birds. These results are consistent with previous studies on freshwater shrimps and highlight the importance of understanding aquatic host interactions in the epidemiology of this zoonotic food-borne pathogen.

Innovative Multiplex PCR Assay for Detection of tlh, trh, and tdh Genes in Vibrio parahaemolyticus with Reference to the U.S. FDA's Bacteriological Analytical Manual (BAM).

Vibrio parahaemolyticus is an important foodborne bacterium that causes severe gastroenteritis following the consumption of contaminated seafood. To identify V. parahaemolyticus and determine its pathogenicity, the U.S. Food and Drug Administration (FDA)'s Bacteriological Analytical Manual (BAM) recommends a multiplex polymerase chain reaction (PCR) protocol to simultaneously detect the species-specific thermolabile hemolysin (tlh) gene and the pathogenic thermostable-related hemolysin (trh) and thermostable-direct hemolysin (tdh) genes. However, this assay has shown two limitations: difficulty in separating the amplicons of the trh (486 bp) and tlh (450 bp) genes due to their highly similar sizes, and the weaker band exhibited by the tdh gene amplicon (270 bp). The present study aimed to improve the BAM's multiplex PCR assay by separating the three amplicons with similar intensity. A new primer set was applied for the tlh gene (369 bp) alongside the existing primers for the trh and tdh genes. The amplicons for the three genes were effectively separated by electrophoresis on a 2% tris-borate-EDTA (TBE) agarose gel within 45 min. Primer concentrations of 0.25 µM for three genes produced a significant amount of amplicons among various combinations of primer concentrations with 35 PCR cycles. This assay exhibited a detection limit of 10 pg of bacterial DNA, demonstrating its high sensitivity. It did not display amplicons from nine Vibrio species known to be human pathogens or from 18 well-documented foodborne pathogens. Therefore, the present multiplex PCR protocol could help overcome the limitations of existing assays and provide a more reliable method for detecting the three genes of V. parahaemolyticus.

In the Depths of Wash Water: Isolation of Opportunistic Bacteria from Fresh-Cut Processing Plants.

The fruit and vegetable industry in post-harvest processing plants is characterized by a substantial consumption of water resources. Wash waters may serve as an environment for the periodic or permanent habitation of microorganisms, particularly if biofilm forms on the inner walls of tanks and flushing channels. Despite the implementation of integrated food safety monitoring systems in numerous countries, foodborne pathogens remain a global public health and food safety concern, particularly for minimally processed food products such as vegetables and fruits. This necessitates the importance of studies that will explore wash water quality to safeguard minimally processed food against foodborne pathogen contamination. Therefore, the current study aimed to isolate and identify bacteria contaminating the wash waters of four fresh-cut processing plants (Poland) and to evaluate the phenotypic antibiotic resistance profiles in selected species. Bacteria were isolated using membrane filtration and identified through mass spectrometry, followed by antibiotic susceptibility testing according to EUCAST guidelines. The results revealed that the level of contamination with total aerobic bacteria in the water ranged from 1.30 × 106 cfu/mL to 2.54 × 108 cfu/mL. Among the isolates, opportunistic pathogens including Enterococcus faecalis, Pseudomonas aeruginosa, Klebsiella oxytoca, Klebsiella pneumoniae, Serratia marcescens, and Proteus vulgaris strains were identified. An especially noteworthy result was the identification of cefepime-resistant K. oxytoca isolates. These findings highlight the importance of monitoring the microbial microflora in minimally processed foods and the need for appropriate sanitary control procedures to minimize the risk of pathogen contamination, ensuring that products remain safe and of high quality throughout the supply chain.

Efficient metabolic pathway modification in various strains of lactic acid bacteria using CRISPR/Cas9 system for elevated synthesis of antimicrobial compounds

Lactic acid bacteria (LAB) are known to exhibit various beneficial roles in fermentation, serving as probiotics, and producing a plethora of valuable compounds including antimicrobial activity such as bacteriocin-like inhibitory substance (BLIS) that can be used as biopreservative to improve food safety and quality. However, the yield of BLIS is often limited, which poses a challenge to be commercially competitive with the current preservation practice. Therefore, the present work aimed to establish an optimised two-plasmid CRISPR/Cas9 system to redirect the carbon flux away from lactate towards compounds with antimicrobial activity by disrupting lactate dehydrogenase gene (ldh) on various strains of LAB. The lactic acid-deficient (ldhΔ) strains caused a metabolic shift resulting in increased inhibitory activity against selected foodborne pathogens up to 78 % than the wild-type (WT) strain. The most significant effect was depicted by Enterococcus faecalis-ldh∆ which displayed prominent bactericidal effects against all foodborne pathogens as compared to the WT that showed no antimicrobial activity. The present work provided a framework model for economically important LAB and other beneficial bacteria to synthesise and increase the yield of valuable food and industrial compounds. The present work reported for the first time that the metabolism of selected LAB can be manipulated by modifying ldh to attain metabolites with higher antimicrobial activity.

Comparative assessment of Rhazya stricta and Azadirachta indica methanolic leaf extracts for their antimicrobial efficacy against the selected foodborne pathogens

Comparative assessment of Rhazya stricta and Azadirachta indica methanolic leaf extracts for their antimicrobial efficacy against the selected foodborne pathogens

The application and reflection of emerging molecular detection technologies in Salmonella detection

Salmonella is an important foodborne pathogen and one of the main causes of diarrhea. Every year, about 550 million people suffer from diarrhea due to Salmonella infection, of which about 230 000 die. It has become a major global public safety issue. The application fields of Salmonella detection involve food safety, water quality monitoring, animal husbandry, public health monitoring, and medical diagnosis. The detection requirements mainly come from three aspects: pathogen identification, serotype identification, drug resistance and virulence identification. In recent years, the detection technology for Salmonella has made rapid progress, especially the emergence and development of emerging molecular detection technologies, providing new perspectives for Salmonella detection in different scenarios. However, due to the diversity of Salmonella serotypes and the complexity of detection scenarios, existing detection technologies still have some pain points (such as long detection time, cumbersome operation steps, low scene adaptability, etc.). This article will elaborate on the application of several emerging molecular detection technologies with distinct characteristics, such as CRISPR Cas technology, digital PCR technology, sequencing technology, and microfluidic technology, in Salmonella detection. It aims to provide a reference for the development and improvement of Salmonella detection technology and the establishment of infection warning and control systems.