Brettanomyces bruxellensis is a major spoilage yeast in winemaking due to its persistence in winery environments and its ability to produce volatile phenols responsible for sensory defects. This study evaluated the biocontrol potential of selected Metschnikowia pulcherrima strains, focusing on the role of pulcherriminic acid production and its application in winery-relevant conditions. Twelve M. pulcherrima strains were screened for pulcherriminic acid production under different iron conditions, revealing strong strain-dependent variability. Three strains (1521, 1558, and DSM 70336) were identified as the most efficient producers and exhibited significant iron depletion capacity. The produced metabolite remained stable over an 18-day period. Cell-free supernatants from selected strains inhibited the growth of B. bruxellensis in a strain- and concentration-dependent manner, with the strongest effect observed for strain M. pulcherrima DSM 70742. Biofilm assays on oak wood demonstrated that the supernatants reduced viable cell counts by up to 80-86% when applied at the initial stage of colonization, whereas significantly lower reductions (10-55%) were observed for pre-established biofilms. Application of a mixed supernatant under winery conditions successfully prevented Brettanomyces contamination in treated barrels without affecting wine sensory properties. These findings indicate that M. pulcherrima-derived metabolites primarily act by limiting iron availability and interfering with early biofilm formation. The results highlight the potential of M. pulcherrima as a practical and sustainable biocontrol agent for reducing spoilage yeast contamination and limiting the need for chemical preservatives in winemaking.

The antibacterial activities of numerous medium and long-chain unsaturated fatty acids are well documented. However, the natural short-chain unsaturated fatty acid, 4-pentenoic acid (C5:1 Δ4) has not been systematically evaluated for its antibacterial activity, mechanism of action, or application potential. This study investigated these to provide a basis for novel preservatives. 4-Pentenoic acid showed broad-spectrum activity against 21 strains of pathogenic bacteria, with MICs 1.5-3 mg/mL, MBCs 3-6 mg/mL, and low resistance risk. It disrupted bacterial membrane integrity, inhibited biofilm formation, and regulated the pdu, nar and RND efflux pump genes, thereby affecting bacterial metabolism and stress responses. Checkerboard broth microdilution susceptibility assays confirmed that the combination of 4-pentenoic acid with antibiotics (such as meropenem) exerted additive antibacterial effects. At effective antibacterial concentrations, oral administration of 4-pentenoic acid caused no observable toxicity. It can markedly inhibit the proliferation of pathogenic bacteria in milk, beef, and peanut butter, while dosages close to the minimum inhibitory concentration exerted minimal adverse effects on the sensory properties and pH of these foods. In summary, this study provides theoretical and experimental basis for the development of new food preservatives and antibacterial agents.

Vibrio parahaemolyticus remains a major cause of seafood-associated gastroenteritis, yet routine surveillance still relies largely on the canonical hemolysin markers thermostable direct hemolysin (tdh) and tdh-related hemolysin (trh). To determine whether this framework overlooks accessory virulence determinants in trh-positive lineages, we analyzed 193 V. parahaemolyticus isolates collected between 2022 and 2025 from clinical, environmental, and seafood-associated sources in the Hangzhou Bay region of China. Serotyping identified 45 serotypes, with O10:K4 predominating among clinical isolates. Both clinical and non-clinical populations showed open pan-genomes, although the non-clinical group carried a larger accessory gene pool. We identified a complete hlyCABD-like RTX operon in 10 trh-positive isolates with T3SS2-associated virulence backgrounds. These RTX-positive isolates were distributed across seven sequence types and three of five phylogenetic groups. This distribution was lineage-restricted but non-clonal. In the representative hybrid-assembled genome, the operon occurred within a mosaic genomic region containing additional virulence- and mobility-associated genes, indicating a composite pathogenicity island-like element. In the tested subset, RTX-positive isolates showed significantly greater hemolytic activity than RTX-negative trh-positive isolates. This significant difference was consistently observed in both plate-based and liquid assays, and within the RTX-positive subset, hlyA expression correlated with hemolytic activity, whereas the trh gene and the tlh (thermolabile hemolysin) gene did not. A complete hlyCABD-like RTX operon therefore identifies a hemolysis-associated subset of trh-positive V. parahaemolyticus and supports its further evaluation as an additional target for food safety surveillance.

The increasing significance of non-Saccharomyces strains in fermented beverages comes from their ability to modulate sensory properties and produce diverse metabolites. This study evaluated the oenological potential and fermentation capacity of five Schizosaccharomyces species (S. pombe var. pombe, S. pombe var. malidevorans, S. japonicus, S. octosporus, and S. cryophilus). While all strains assimilate and ferment glucose, fructose, and sucrose, they show no utilization of sweeteners such as xylitol, erythritol, or stevia. Notably, the strains exhibited tolerance to 15-30% glucose, and most of them could grow at 15 °C. Fermentation trials revealed that all species successfully produced 4-6% (v/v) ethanol in apple juice, characterized by high glycerol production and efficient malic acid degradation, while maintaining acetic acid levels comparable to the Saccharomyces bayanus control. Volatile organic compound (VOC) analysis identified ester-rich aroma profiles-featuring fruity, banana-like, and pineapple-like notes-comprising both common and species-specific components. These findings demonstrate that Schizosaccharomyces species are viable candidates for cider production, offering a unique combination of deacidification, glycerol enrichment, and novel aromatic complexity, particularly for use in mixed-culture fermentation strategies.

From 2020 to 2022, an outbreak of listeriosis occurred in Austria, resulting in ten cases, three of which proved fatal. The outbreak was traced to a traditional southern European dairy product known as Kajmak, a lightly salted cream product. The subsequent sequencing of clinical and food-related isolates with clonal complex (CC)1 clonal type (CT) 6568 led to the identification of the outbreak's origin. CT6568 is classified under CC1, which exhibits notable diversification among human isolates, with 14 distinct CTs identified between 2020 and 2022. The clientele of the cheese factory primarily comprised local Balkan restaurants, delicatessen counters, and small bakeries. The existing self-monitoring system and the Hazard Analysis and Critical Control Points (HACCP) program did not meet the requisite standard of adequacy. Listeria monocytogenes was identified in the production and ripening room, indicating potential recontamination of Kajmak during the manual filling and ripening process. The highly perishable product in question was also incorrectly labeled with a best-before date (>21 days) instead of a use-by date. Furthermore, crucial labeling information for customers, such as the necessity of strict refrigeration at 4 °C, was absent. The outbreak was likely precipitated by the discontinuous production of Kajmak, which resulted in inadequate environmental hygiene. The retrospective report on the outbreak highlights the lack of risk analysis in research conducted on food safety risks associated with small-scale production of traditional foods. It is essential that manufacturers align their understanding with the principles of the HACCP concept and effective self-monitoring systems.

Malolactic fermentation (MLF) by lactic acid bacteria (LAB) is essential for most red wine production, yet commercial starter cultures are largely limited to Oenococcus oeni, a species constrained by slow growth, fastidious nutritional requirements, and poor co-inoculation competitiveness. To identify novel MLF candidates from diverse ecological niches, we assembled 714 wild LAB strains from seven Australian geographic regions and eight ecological sources, including insects, flowers, honey, and fermented foods, spanning 21 genera. Four hundred strains were screened in a microplate assay quantifying nine metabolites in Shiraz grape juice and classified into six functional categories based on thresholds for malic acid reduction, growth, and spoilage compound production. Fourteen strains (3.5%) achieved High Performance, dominated by Lactiplantibacillus (9/14). Two Apilactobacillus strains, a genus known primarily for wine spoilage, also completed MLF without spoilage compound production. Twenty-three strains were validated in co-fermentation with Saccharomyces cerevisiae using an automated robotic platform. An insect-derived Lactiplantibacillus plantarum from Kangaroo Island completed MLF within 48 h, outperforming the commercial Lp. plantarum control, while all 14 Fructilactobacillus strains failed in co-fermentation despite demonstrating MLF activity in juice, indicating that wine conditions rather than metabolic capacity determined MLF outcome. Volatile profiling of five selected candidates confirmed wild strains preserved yeast-derived aroma profiles. Geographic origin significantly predicted functional category (P < 0.001), with grape-associated environments enriched for MLF-competent strains. This study validates environmental bioprospecting as a strategy for wine starter culture discovery and provides a curated collection of 714 LAB for fermentation applications.

In recent years, the number of omics studies aimed at characterizing fungal communities in fermented foods, such as table olives, has steadily increased. Moreover, advances in bioinformatics and artificial intelligence have provided new resources for analysing these datasets. Among these tools, supervised machine learning (ML) models, particularly tree-based algorithms, are especially valuable for their interpretability and their ability to identify underlying patterns. In this study, we evaluated three target variables of table olive samples (processing type, country of origin, and olive cultivar) by implementing three models built with tree-based algorithms, specifically Classification and Regression Trees (CART), Random Forest (RF), and eXtreme Gradient Boosting (XGB). The initial dataset consisted of 872 samples of fungal metataxonomic data obtained from diverse table olive sources. The RF models were the most accurate, achieving an overall accuracy above 75% and a kappa coefficient greater than 0.65. The highest accuracy and robustness were obtained for the model classifying country of origin, with 89% accuracy and a kappa coefficient of 0.86. Furthermore, the use of tree-based models enabled the identification of the fungal genera that contributed most to sample classification. Among the most important genera detected, we noticed Citeromyces, Candida, Pichia, Zygotorulaspora, Taphrina, Wickerhamomyces, Saccharomyces, Starmerella, Aureobasidium, and Dekkera. This approach demonstrates the potential for application of this methodology in the table olive sector and other fermented foods, where the industrial implementation of ML techniques based in omic data could enhance traceability, authenticity, and quality control.

Ochratoxin A (OTA) is a widespread mycotoxin found in food and feed, posing serious risks to human and animal health due to its nephrotoxic, immunotoxic, and carcinogenic effects. Strategies to reduce OTA exposure are therefore essential, and biological approaches based on probiotic microorganisms have gained increasing attention. The objective of this study was to identify probiotic bacteria suitable as food and feed additives capable of mitigating the effects of the mycotoxin OTA in the gastrointestinal tract, either through adsorption or metabolic degradation. A total of twenty probiotic bacterial strains belonging to eight different genera were evaluated. The experimental design included an initial screening phase in which the bacterial strains (n = 20) were cultivated in appropriate media. Based on their performance, three strains were selected for a subsequent in vitro digestion assay. OTA and its potential degradation product, ochratoxin α (OTα), were quantified using high-performance liquid chromatography with fluorescence detection (HPLC-FLD). To gain insights into the metabolic capacity of the bacteria, a proteolytic activity test was performed in parallel, followed by the analysis of OTα formation during both the screening and digestion phases. Among the tested bacterial genera, Pediococcus, Lactiplantibacillus, and Limosilactobacillus showed greater potential than Bacillus, Priestia, and Cytobacillus, with the highest efficacy observed for P. pentosaceus 4, Lactiplantibacillus plantarum 5, and Limosilactobacillus fermentum 8. However, all strains exhibited lower detoxification rates under in vitro digestion conditions compared to the screening phase. Although further validation is required, these findings suggest that the evaluated probiotic bacteria represent a promising strategy for OTA mitigation in food and feed, even if complete detoxification has not yet been achieved.

Contamination of milk by psychrotrophic Pseudomonas species represents a primary cause of dairy product spoilage. Antimicrobial peptides (AMPs) present unique advantages for food applications due to their favorable safety profiles, low toxicity, and limited potential to promote antimicrobial resistance. However, conventional AMP screening approaches remain time-consuming and resource-demanding. This study developed an LSTM-based deep learning framework for predictive identification of AMPs with inhibitory activity against Pseudomonas species. Four candidate AMPs were identified through computational screening and subsequently chemically synthesized, with three exhibiting significant inhibitory effects against Pseudomonas fluorescens W3. NPN and PI probe labeling indicated that the most effective peptide, designated WY-1, could disrupt the permeability of the outer membrane and the integrity of the inner membrane of Pseudomonas fluorescens W3, causing intracellular substance leakage and subsequent growth inhibition. This peptide significantly reduced both microbial biomass and extracellular protease activity in contaminated milk models, thereby demonstrating substantial translational potential for dairy preservation applications.

Antimicrobial resistance is a growing global public health concern, and carbapenem-resistant Enterobacterales (CRE) represent one of the most urgent threats due to limited treatment options. While CRE are primarily associated with healthcare settings, increasing evidence suggests that food systems may serve as additional reservoirs and transmission interfaces for clinically significant antibiotic-resistant bacteria. This study aimed to characterize CRE from vegetables, with a focus on isolates carrying carbapenem resistance genes on mobile genetic elements and evaluating their transferability to a recipient bacterium. Commercial vegetable samples were collected from retail grocery stores or farmers' markets and screened for CRE using selective enrichment and CHROMagar™ mSuperCARBA™. Antimicrobial susceptibility testing was performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Whole-genome sequencing using Illumina short-read and Oxford Nanopore long-read platforms was conducted to characterize resistance genes and the plasmid sequence. Conjugation assays were performed to assess plasmid transferability to Escherichia coli DH5α. The results indicated that a carbapenem-resistant Kluyvera sichuanensis isolate (CA-CRE-C23) was recovered from a spinach sample. Genomic analysis identified a 51,477-bp hybrid ColKP3-IncX3 plasmid carrying the carbapenemase gene blaOXA-181 and quinolone resistance gene qnrS1. Conjugation experiments, disk diffusion, and sequencing confirmed successful plasmid transfer from K. sichuanensis isolate (CA-CRE-C23) to E. coli DH5α. The plasmid structure closely resembled globally disseminated IncX3 hybrid plasmids carrying blaOXA-181, which have been predominantly reported in clinical isolates. In conclusion, the detection of a transferable hybrid plasmid harboring blaOXA-181 in a fresh produce-associated K. sichuanensis isolate expands the known host range of this resistance gene and underscores the importance of food-chain surveillance in understanding the environmental dissemination of high-risk antimicrobial resistance genes.

Enterococcus faecium, a member of the human gut microbiota and the second most abundant enterococcal species after E. faecalis, is an important agent of healthcare-associated infection. Its high capacity to acquire antimicrobial resistance genes (ARGs) makes infections difficult to treat. This study investigated ready-to-eat (RTE) meat products, typical of central Italy, as potential vectors of antibiotic-resistant enterococci, focusing on E. faecium. A total of 148 enterococcal strains, 36 of which were identified as E. faecium, were isolated. Among 22 distinct clones, eight were resistant to tetracycline (TET), two also to linezolid (LZD), and one showed a multidrug-resistance phenotype. Antibiotic susceptibility was determined by minimum inhibitory concentration testing, supported by whole-genome sequence analysis. Both LZD-resistant strains harbored LZD resistance genes (optrA or a combination of optrA and poxtA), while only one isolate demonstrated horizontal gene transfer via conjugation. The isolates were further characterized for virulence factors and biofilm formation capacity, both under basal conditions and following exposure to simulated upper gastrointestinal transit. All eight TET resistant strains presented genotypic virulence traits. Following exposure to simulated upper gastrointestinal transit, these isolates demonstrated high survival rates, maintaining both their antibiotic resistance phenotypes and biofilm-forming capacity. These findings highlight the potential role of the analyzed RTE meat products as vehicles for virulent and antibiotic-resistant E. faecium strains capable of surviving upper gastrointestinal exposure. The study underscores the need to implement enterococci surveillance in the food sector to improve monitoring of resistant E. faecium and to limit its possible dissemination and association in clinical risks.

Traditional Chinese sourdough (CTS) is mainly used for the fermentation of steamed pastries, providing a unique natural fluffiness and distinctive flavor, and thus holds important culinary value. However, the microbial mechanisms underlying the diversity of its regional characteristic flavors remain poorly understood. In this study, we integrated metagenomic sequencing with multi-platform flavor profiling-including high-performance liquid chromatography (HPLC), electronic nose, gas chromatography-mass spectrometry (GC-MS), and gas chromatography-ion mobility spectrometry (GC-IMS)-to characterize the physicochemical properties, microbial composition, and flavor compounds of 10 CTS samples collected from five provinces across China. A total of 1231 genera and 3358 species were identified, with Fructilactobacillus sanfranciscensis, Saccharomyces cerevisiae and Lactiplantibacillus plantarum being the dominant species. Flavor profiling analysis revealed 109 volatile organic compounds (VOCs), of which 11 key aroma-active compounds (e.g., 1-nonanol, phenethyl alcohol) were identified based on odor activity values (OAV ≥ 1). Using orthogonal partial least squares (O2PLS) modeling, we established associations between 25 potential flavor-producing microorganisms and specific metabolites. Notably, S. cerevisiae exhibited a significant positive correlation with acetic acid, 1-nonanol and glutamic acid, while L. plantarum showed a strong positive correlation with phenethyl alcohol. This study reveals the correlation patterns between microbial communities and flavor compounds in CTS, offering foundational insights for starter culture design, flavor standardization, and industrial application of traditional fermented doughs.

High-temperature Daqu (HTD), the core starter for sauce-flavor Baijiu, is highly susceptible to quality fluctuations because it is produced through open solid-state fermentation and depends on heterogeneous Muqu. Here, we evaluated whether defined synthetic microbial communities (SynMCs) could improve process controllability and inter-site reproducibility during HTD production. SynMC biofortification reduced physicochemical variability and reshaped microbial community succession in a stage- and composition-dependent manner. Among the tested designs, inter-kingdom community, particularlySynMC4 and SynMC6, showed comparatively stable physicochemical performance and higher pyrazine accumulation. Integrated multi-omics analysis associated pyrazine enrichment with coordinated shifts in specific low-abundance taxa, suggesting that rare members may participate in VOC differentiation under SynMC regulation. Ecological analyses further indicated that SynMCs altered the balance between stochastic and deterministic assembly and reorganized co-occurrence networks by modifying connectivity and modularity, although these effects varied across fermentation stages and subcommunities. Cross-site validation showed that SynMC6 exerted partially consistent regulatory effects on bacterial communities across geographically independent production systems, while the magnitude and specific metabolic outcomes remained site-dependent. These findings provided an application-oriented basis for improving HTD standardization and highlight the need to incorporate both abundant and rare taxa into further SynMC design.

A novel photodynamic ice glaze with high antibacterial ability based on curcumin (Cur)/β-cyclodextrin (β-CD) inclusion complex was developed, and its active cold-chain preservation efficiency for Pacific white shrimp was investigated. Results showed that β-CD/Cur inclusion complex significantly enhanced the aqueous solubility and photostability of curcumin by facilitating the formation of a novel crystalline phase, which enabled efficient reactive oxygen species (ROS) generation in its frozen aqueous state. Under blue LED irradiation during frozen storage, the photodynamic ice exhibited broad-spectrum antibacterial ability against foodborne pathogens, notably with a reduction of >5.98 Log CFU/mL for Gram-positive Staphylococcus aureus and an undetectable level for Gram-negative Vibrio parahaemolyticus. Mechanistic studies confirmed that the generated ROS disrupted cellular structural integrity, and inflicted severe oxidative damage to bacterial DNA and proteins. During 15 days of frozen storage of shrimp, the photodynamic ice glaze slowed microbial proliferation (<5.20 Log CFU/g), inactivated endogenous proteases, retarded water migration, and maintained muscle quality. Therefore, the photodynamic ice glaze is a green and feasible strategy for improving the microbial safety and quality of aquatic products.

To uncover the microbial underpinnings of quality and flavor differences between Xuanwei ham (XWH) and Iberian ham (IRH), this study integrated high-throughput sequencing with metabolomics and volatilomics to establish microbiota-metabolite-flavor linkages. Physicochemical characterization revealed that XWH showed higher moisture, water activity, and salt content, whereas IRH had higher fat and protein. These matrix differences imposed selective pressures on microbial colonization. The IRH surface exhibited higher bacterial diversity, while the XWH interior showed higher bacterial diversity; XWH also had higher fungal diversity overall. Staphylococcus dominated both hams, whereas Debaryomyces yeasts were enriched inside hams. Metabolomics identified 312 differential metabolites, mainly enriched in amino acid pathways. XWH was enriched in sweet-tasting amino acids and short-chain fatty acids, while IRH featured glutamic acid, polyunsaturated fatty acids, and phenolic acid esters. Volatilomics revealed 24 key differential odorants: IRH was enriched in medium-to-long chain ketones and aldehydes from lipid oxidation, and XWH accumulated short-chain ketones and alcohols from carbohydrate fermentation. Correlation analysis showed that Staphylococcus and Debaryomyces were strongly positively correlated with most differential metabolites and key volatiles, indicating their driving role in shaping the metabolic landscape. Tight positive correlations between flavor compounds and their precursor metabolites further support a microbiota-mediated cascade from substrate degradation to volatile generation. These findings demonstrate that region-specific processing conditions sculpt distinct microbial consortia and that core microbiota are primary drivers of quality differentiation. This work provides a mechanistic framework for flavor-directed regulation and starter culture design in dry-ham production.