This study investigated the effects of cocultivation and pH on the metabolite profile of Chaetomium globosum SCF-24. Axenic and coculture fermentation systems were established at pH 5-8, and resulting extracts were analyzed by gas chromatography-mass spectrometry (GC-MS). A hierarchical clustered heatmap was used to compare metabolite profiles based on presence-absence data. GC-MS detected 51 putatively identified primary and secondary metabolites, some of which were detected across all treatments, whereas others were condition-specific. Metabolite profiles also differed between axenic cultures and cocultures, even at identical pH, with numerous metabolites detected exclusively in cocultures, demonstrating that microbial interactions influenced metabolite profiles beyond the effects of pH. Antibacterial assays showed that only extracts from axenic cultures and cocultures at pH 6 inhibited both Gram-positive and Gram-negative test bacteria. Notably, treatments with similar metabolite profiles did not exhibit comparable antibacterial activity, and the treatment with the highest number of detected metabolites was not the most bioactive, suggesting that bioactivity depends more on metabolite composition than on the number of detected metabolites. Overall, these findings demonstrate the combined influence of biotic and abiotic cultivation factors on metabolite composition and associated antibacterial activity.
Listeria monocytogenes is a major foodborne pathogen in ready-to-eat smoked fish products. This study evaluated whether clonal complex affiliation contributes to variability in growth responses to stresses representative of smoked salmon and trout processing. Ten strains were studied, including strains from CC121, CC26 and CC204, the three major clonal complexes reported in the French smoked salmon and trout sectors, together with the EGDe reference strain. Strains were exposed to salt, cold, smoke-derived phenolic compounds and combined stress conditions. Growth responses were compared with whole-genome-based phylogeny, and the impaired phenotype observed under phenolic exposure was further investigated using viable counts, live/dead microscopy and comparative genomics. Growth profiles were partly structured by clonal complex, with strains from the same clonal complex showing similar behaviour across stress conditions. Salt and cold reduced growth globally, while smoke-derived phenolic compounds were the most discriminating conditions. CC204 strains showed markedly lower growth rates under phenolic exposure than CC121, CC26 and EGDe. This phenotype was not associated with loss of cultivability or significant loss of membrane integrity. Comparative genomics did not identify a clear gene-content determinant explaining the CC204 phenotype. These results suggest that CC204 has an impaired adaptive response to smoke-derived compounds, likely involving regulatory or physiological mechanisms.
Foodborne outbreaks have been associated with both pasteurised and unpasteurised milk cheeses in the UK in recent years. This study aimed to assess and compare the microbiological quality of cheese made from pasteurised and unpasteurised milk on sale in England and Northern Ireland. Overall, 2032 cheese samples were collected from retailers and producers throughout England and Northern Ireland. Samples originated from 27 countries, with the UK (53%) and France (15%) being the most common. When compared to UK legislation and guidance, 96% were considered to be of satisfactory microbiological quality, with a greater proportion of unsatisfactory results in unpasteurised (6%) than pasteurised milk cheeses (2%). Five samples (0.2%), all made from unpasteurised milk, were potentially injurious to health due to the detection of shiga toxin-producing E. coli (STEC) in two, >104 CFU/g of coagulase positive staphylococci in two and > 100 CFU/g of Listeria monocytogenes in one. Neither the L. monocytogenes nor the STEC isolates were genetically closely related to those from recent cases of human illness. While the majority of samples were of satisfactory microbiological quality, a small number of results of public health concern were highlighted, particularly where cheeses were made with unpasteurised milk.
This study aimed to identify kefir-derived microorganisms, evaluate their probiotic potential to select the most promising strain, and assess the selected strain's antibacterial and antibiofilm activities against foodborne isolates, Escherichia coli, and Klebsiella pneumoniae. Kefir grains were cultivated in selective media, and isolates were identified using MALDI-TOF mass spectrometry. Their acid and NaCl tolerance, auto- and co-aggregation, and antimicrobial activity were assessed to identify the most effective probiotic strain. Subsequently, the selected strain was subjected to spot overlay tests, broth microdilution assays, and biofilm inhibition experiments against E. coli and K. pneumoniae isolates. Enterococcus durans, Leuconostoc pseudomesenteroides, Lacticaseibacillus paracasei, and Kluyveromyces marxianus were identified. Among these, L. paracasei (Lpk 01) demonstrated the best probiotic characteristics, with 50% survival at pH 5, tolerance to 2% NaCl, and high co-aggregation with K. pneumoniae (40%). It inhibited pathogens (inhibition zones >14 mm), and its cell-free supernatant (Lpk-cf) reduced biofilm biomass by approximately 50%, with effects minimized upon neutralization, suggesting the involvement of organic acids. These findings affirm the antimicrobial potential of L. paracasei, highlighting its application as a natural alternative against bacterial pathogens.
Traditional and industrial soybean fermentations harbor distinct microbial communities that influence product quality and safety. In this study, long-read rRNA operon sequencing was applied to investigate the bacterial communities of traditional and commercial doenjang and cheonggukjang. The 16S-23S rRNA operon (∼4.2 kb) was amplified and sequenced using the Oxford Nanopore MinION platform to obtain high-resolution taxonomic profiles. Traditional doenjang and cheonggukjang exhibited greater microbial diversity than commercial products, as revealed by alpha and beta diversity analyses. Bacillus species, particularly Bacillus velezensis and Bacillus subtilis, were dominant across all samples, while Loigolactobacillus coryniformis, Caldifermentibacillus hisashii, and Tetragenococcus halophilus were more abundant in traditionally fermented samples. These genera are associated with proteolysis, amino acid metabolism, and flavor compound formation during fermentation. Overall, these findings provide insights into the microbial ecology of fermented soybean foods and suggest that non-starter lactic acid bacteria and thermophilic species may contribute to the unique sensory characteristics of traditionally fermented doenjang.
Benzalkonium chloride (BAC) is a widely used disinfectant, but its improper application can induce Salmonella Typhimurium (S. Typhimurium) to develop tolerance to subsequent stressors. In this study, we demonstrated that adaptation of S. Typhimurium to BAC conferred both direct protection and cross-protection against cold stress, resulting in significantly enhanced survival at -20°C compared with non-adapted cells. BAC-adapted S. Typhimurium also exhibited a survival advantage on chicken meat during frozen storage at -20°C. Membrane fatty acid analysis indicated that adaptation of S. Typhimurium to BAC led to a 2.2-fold increase in the levels of cyclopropane fatty acids. Moreover, BAC-adapted S. Typhimurium showed a significant 8.0% increase in cell surface hydrophobicity. RT-qPCR analysis demonstrated upregulation of efflux pump genes tolC, acrB, and mdfA (3.6-, 3.4-, and 3.0-fold, respectively) and cold shock-related genes cspA, cspE, and rpoS (2.4-, 4.0-, and 2.6-fold, respectively). These findings suggest that upregulation of cold shock genes in BAC-adapted cells was associated with the observed cross-protection. Taken together, adaptation of S. Typhimurium to BAC promoted the development of direct and cross-protective phenotypes, which may increase the risk of cross-contamination and salmonellosis in the food industry, posing a serious threat to food safety and public health.
The combination of sophorolipids and copaiba essential oil in a multifunctional facial formulation aimed at the treatment of conditions such as acne and herpetic folliculitis was evaluated. Sophorolipids produced by Starmerella bombicola reached 38.37 g/l, predominantly in the lactonic form. Their antimicrobial activity, alone and combined with copaiba oil, was evaluated against Cutibacterium acnes, Staphylococcus aureus, and Staphylococcus epidermidis. The combination reduced the minimum inhibitory concentration against C. acnes from 62.50 to 31.25 μg/ml, demonstrating an additive effect. Antiviral assays showed strong anti-HSV-1 activity, with 95.58% inhibition in the virucidal test and 63.99% in the antiviral assay. Three dermocosmetic formulations were developed, and the optimized formulation achieved 91.45% inhibition of C. acnes and 26.17% antioxidant activity, confirming its multifunctional potential. The application of a formulation combining sophorolipids and copaiba oil shows promising potential for the treatment of skin conditions, demonstrating significant antimicrobial activity against C. acnes as well as adequate physicochemical stability.
Streptococcus agalactiae is a significant opportunistic pathogen causing severe infections, including urinary tract infections and pyelonephritis in diabetic individuals. To explore the molecular mechanisms behind this susceptibility, we used a streptozotocin-induced diabetic murine model to evaluate S. agalactiae renal infection and performed the first comparative proteomic analysis of S. agalactiae COH1 isolates recovered from the kidneys of diabetic and non-diabetic mice. Diabetic mice exhibited significantly higher bacterial loads and more severe renal histopathological damage. Comparative proteomic profiling identified 2751 proteins, with 82 significantly upregulated and 91 downregulated proteins in S. agalactiae COH1 isolate recovered from the kidneys of diabetic animals. Upregulated virulence factors included serine-rich repeat 2, type VII secretion, BibA, C5a peptidase, and PI-1. Functional analysis revealed a metabolic shift toward increased glycolysis, PTS system activity, and translation machinery, suggesting that the hyperglycemia promoted bacterial proliferation and energy production. Conversely, proteins related to proteolysis were downregulated. Our findings demonstrated that hyperglycemia induced a metabolic reprogramming and enhanced the expression of virulence-associated proteins in S. agalactiae. These adaptations contributed to increased colonization and tissue damage in diabetic kidney, providing potential molecular targets for future therapeutic interventions.
Slow-transit constipation (STC) is characterized by reduced colonic motility, impaired smooth muscle function, and altered mucosal signaling. Among these pathways, mucosal serotonin (5-HT), mainly produced by enterochromaffin cells, contributes to intestinal motility regulation. In a loperamide-induced constipation model in male C57BL/6 J mice, we evaluated the effects of Lactiplantibacillus plantarum NCHBL-004 on bowel function and associated molecular changes. Compared with loperamide-treated mice, live NCHBL-004 at 1 × 10⁹ CFU improved fecal output and gastrointestinal transit and partially preserved fecal water content, whereas the lower live dose and heat-killed preparation showed no significant effects. These functional changes were accompanied by preservation of colonic muscularis thickness and increased expression of smooth muscle-associated markers, including Acta2 and Myh11. NCHBL-004 also modulated mucosal serotonergic gene expression, as indicated by increased Tph1, which encodes a key enzyme for 5-HT synthesis, and decreased Slc6a4, which encodes the serotonin transporter involved in 5-HT reuptake. Overall, these findings suggest that live NCHBL-004 improves intestinal motility in loperamide-induced constipation, with associated changes in colonic smooth muscle phenotype and mucosal serotonergic regulation.
Quinolone-resistant Escherichia coli are increasing in humans and livestock; however, their distribution and underlying mechanisms in shared environments remain incompletely characterized. We analyzed quinolone resistance in E. coli isolates from swine (n = 100) and humans (n = 65) associated with an agroecological farm in Morelos, Mexico. Antimicrobial susceptibility testing was performed for quinolones [nalidixic acid (NAL) and ciprofloxacin (CIP)], and the minimum inhibitory concentrations were determined. Phylogenetic grouping and genotypic analyses targeting gyrA mutations and plasmid-mediated quinolone resistance (PMQR) genes were conducted. High frequencies of NAL and moderate CIP resistance were observed in both swine (NAL, 79%; CIP, 23%) and human (NAL, 65%; CIP, 15%) isolates. The predominant phenotype was NAL-resistant/CIP-susceptible, particularly among commensals of phylogroups A and B1. High-level resistance to both antimicrobials was associated with double mutations in gyrA (S83L + D87N), whereas PMQR determinants, including qnr variants and aac(6')-Ib-cr, were widely distributed and linked to lower-level resistance. These findings indicate that quinolone resistance is shaped by the combined contribution of chromosomal mutations and plasmid-mediated mechanisms. The presence of PMQR determinants in isolates with reduced susceptibility highlights their potential role in the early stages of resistance development, emphasizing the urgent need for integrated surveillance of antimicrobial resistance in agricultural and community settings.
The growing complexity of food-safety systems and the increasing emergence of multidrug-resistant (MDR) foodborne pathogens demonstrate the importance of enhanced genomic surveillance. This study employed whole genome sequencing (WGS) to characterise the genomes of Bacillus safensis NWU MK_WT, Enterococcus lactis ENT7_CNKT_NWU, and ENT3_CNKT_NWU, isolated from food sources. Phenotypic antibiotic susceptibility testing revealed that all strains displayed MDR phenotypes, with resistance to erythromycin, ampicillin, and meropenem. Genome assemblies ranged from 2.6 to 3.7 Mb, exhibiting high completeness (100%) and diverse functional gene profiles. Furthermore, antibiotic resistance genes (ARGs), including vanT and aac(6'), mediating antibiotic inactivation, efflux, and target modification, were identified. Virulence factors, including adhesion, invasion, and biofilm formation, were detected across genomes, indicating pathogenic potential. Mobile genetic element profiling revealed the presence of insertion sequences, plasmids, and an intact prophage in B. safensis NWU MK_WT, demonstrating genomic plasticity and the potential for horizontal gene transfer (HGT). Phylogenomic comparison showed close relatedness between the isolates and strains from Asia, suggesting possible transboundary movement of genetic material. These findings highlight the growing relevance of WGS for monitoring opportunistic foodborne bacteria that harbour and disseminate resistance and virulence determinants, provide foundational data for improving food safety surveillance, and support antimicrobial resistance (AMR) mitigation strategies.
Rising levels of antimicrobial resistance (AMR) among foodborne pathogens is an increasing threat to food safety and product quality, driving the search for effective natural alternatives to conventional preservatives. Antimicrobial peptides (AMPs) have emerged as promising candidates for food preservation because of their broad-spectrum activity, diverse mechanisms of action and lower propensity for resistance development. Within this group, the macin family represents a promising yet underexplored class of cysteine-rich AMPs characterised by eight conserved cysteine residues that form four disulfide bonds. Macins have been identified across diverse taxa, including leeches, bivalves, gastropods, and hydrozoans and exhibit diverse antimicrobial mechanisms ranging from bacterial aggrelgation mediated by hydrophobic and electrostatic interactions and membrane permeabilization. This review provides a comprehensive synthesis of current knowledge of the macin family within the broader landscape of AMPs, highlighting their structural diversity, mechanisms of action and potential applications. Collectively, these characteristics suggest that macins may represent promising candidates for controlling resistant foodborne pathogens while maintaining product quality.
The process of lignocellulosic biofuel production needs enzymes that are resistant to high temperatures and low pH. The mangrove sediments, which are typified by variable conditions, can contain bacteria that synthesize intrinsically steady enzymes. We selected 193 bacterial isolates of 12 mangrove sites in Goa, India and tested them to produce lignocellulolytic enzymes (cellulase, laccase, xylanase, xylose isomerase) under the conditions of neutral (37°C, pH 7), acidic (37°C, pH 5), thermophilic (50°C, pH 7), and combined stress (50°C, pH 5). Bacillus and Vibrio dominated, with 22 genera identified. There were no significant differences in alpha diversity following Benjamini-Hochberg FDR correction (all P_adj = 1.00, Cohen d < 0.5) but significant compositional differentiation in beta diversity (PERMANOVA: R² = 0.243, P = 0.017). Salinity (R² = 0.903, P_adj = 0.003) and temperature (R² = 0.722, P_adj = 0.006) were major structuring factors. Site-type differentiation was the most significant factor in xylanase-producing communities (R² = 0.272, P = 0.013). Although there was limited replication of dead sites (n = 3), the results confirmed that candidates undergo biochemical characterization and that ecosystem degradation does not decrease diversity but alters community composition.
Cinnamon essential oil (CEO), one of volatile plant compounds, exhibits potent inhibitory effects on bacterial growth. Pseudomonas tolaasii (P. tolaasii) is the primary spoilage bacterium that causes the decay of Agaricus bisporus. This study investigated the effects of CEO on quorum sensing (QS) and biofilm formation in Pseudomonas tolaasii. Whole-genome analysis revealed 102 QS-related genes and their association with 10 virulence protein clusters. AHL signaling molecules (C4-HSL, C6-HSL, C8-HSL, and 3-Oxo-C10-HSL) were identified using biosensor methods and LC-MS/MS. CEO significantly inhibited virulence factor secretion, including protease, elastase, and rhamnolipid. It also reduced swarming and swimming motility by 77.21% and 71.38%, respectively, and effectively suppressed biofilm formation by lowering bacterial viscosity and surface hydrophobicity. At a concentration of 0.4 μL/mL, CEO reduced biofilm formation by 40.89% within 24 h, as measured by the crystal violet method. Microscopic and FTIR analysis confirmed CEO's effect on biofilm and extracellular polymer secretion. Thus, it may serve as a novel type of anti-biofilm agent.
Cronobacter sakazakii is an opportunistic foodborne pathogen associated with severe infections in infants, linked to powdered infant formula (PIF) and related products. We conducted genomic profiling of C. sakazakii (n=209) from infant and toddler food in the United States, comprising all publicly available genomes for this source, through the integration of antimicrobial resistance (AMR) gene (ARG), plasmid replicon, virulence gene, phylogenetic, and pan-genome analyses. We further applied a machine learning (ML)-driven isolation source classification approach based on pan-genome features to distinguish food and clinical isolates. AMR analysis revealed a conserved resistome dominated by three β-lactam resistance genes (blaCSA, blaCSA-1, and blaCSA-2). Independent co-occurrence and pairwise association analysis of ARGs and plasmid replicons indicated sparse and gene-specific relationships, suggesting that observed AMR patterns were more consistent with conserved resistance determinants than extensive plasmid-mediated dissemination. Phylogenetic analysis identified two major clades, while pan-genome assessment demonstrated an open genome dominated by accessory genes. Using gene presence/absence profiles, a random forest classifier achieved high accuracy in distinguishing food and clinical isolates, highlighting the classification power of pan-genome signatures within the dataset. These findings provide insights into the genomic structure of food-associated C. sakazakii and the utility of integrating comparative genomics with ML for food safety surveillance.
Asymptomatic carriage of diarrheagenic Escherichia coli (DEC) in kindergarten children without diarrhoea symptoms represents an important yet underexplored component of community transmission. This study aimed to investigate the prevalence, genomic epidemiology, and antimicrobial susceptibility of DEC recovered from kindergarten children without diarrhoea symptoms in Hengyang, China. Faecal samples (n=150) were collected from kindergarten children without diarrhoea symptoms in Hengyang City in 2024. Molecular screening of these samples was performed using multiplex real-time polymerase chain reaction (PCR) targeting 16 enteric pathogens. PCR-positive samples were subjected to bacterial isolation, antimicrobial susceptibility testing, and whole-genome sequencing. Overall, 37 samples (24.7%) were PCR-positive for at least 1 enteric pathogen. DEC was the only pathogen group successfully recovered by culture, yielding 20 isolates, including 12 enteroaggregative Escherichia coli (EAEC), 7 enteropathogenic Escherichia coli and 1 enterotoxigenic Escherichia coli. Core genome multilocus sequence typing identified a highly conserved EAEC cluster comprising four isolates with zero allelic differences, alongside additional closely related isolates. Antimicrobial susceptibility testing revealed resistance to nalidixic acid, streptomycin, and ampicillin, while most isolates remained susceptible to critically important antibiotics. Genomic surveillance revealed that kindergarten children without diarrhoea asymptomatically carry genetically diverse DEC, including closely related EAEC lineages, underscoring the epidemiological relevance of asymptomatic carriage in community settings beyond clinically apparent diarrhoeal disease.
Canonical and non-canonical isoprenoids offer structurally diverse scaffolds for examining how carbon chain length, methyl-substitution pattern, molecular skeleton, and functional group chemistry influence bacterial metabolic activity. In this study, canonical C5/C10 terpenes and novel non-canonical, methyl-substituted C6/C7/C11 isoprenoids were systematically evaluated against the clinical isolate Pseudomonas aeruginosa ESP050. The comparative analysis showed that antimetabolic activity was not governed by carbon number alone. Short-chain non-canonical C6-C7 isoprenoids displayed comparatively stronger and more discriminating suppression profiles than C10-C11 compounds, which exhibited greater compound-specific heterogeneity. Among C7 dimethyl-isoprenol isomers, the position of the methyl substitution significantly influenced activity threshold, recovery dynamics and sustained suppression, indicating that positional isomerism is a key determinant of the antimetabolic phenotype. In the C10 class, the oxygenated linear compound geraniol demonstrated a more prolonged tendency for metabolic suppression compared to the hydrocarbon bicyclic monoterpenes 3-carene and β-pinene. Furthermore, C11 derivatives demonstrated that methylation modulates activity in a scaffold- and position- dependent manner rather than uniformly enhancing efficacy. Overall, these findings indicated that isoprenoid antimetabolic activity arises from the combined effects of chain length, methyl substitution position, molecular skeleton and oxygenation state, supporting the prioritization of selected C6-C7 scaffolds for further antimicrobial evaluation against P. aeruginosa.
The genus Streptomyces encompasses diverse strains capable of producing antifungal compounds that combat various soil pathogens, responsible for reduced rice yields. The primary aim of this research was to investigate the potential of a consortium comprising four Streptomyces species, specifically Streptomyces chilikensis RC1830, Streptomyces barkulensis RC1831, Streptomyces chitinovorans RC1832, and Streptomyces griseoincarnatus RB7AG, isolated from the Chilika Lake estuary, against Fusarium oxysporum and Rhizoctonia solani. The consortia exhibited significant antagonistic activity against both soil-borne pathogens. In addition, inoculation with the consortium increased the shoot length by 40%, root length by 47.2%, total chlorophyll by 47.5%, and shoot dry weight 45.3% compared to the control. The disease severity index (DSI) was also reduced by approximately 50% following application of the Streptomyces consortium. Formulation trials revealed that formulations prepared using vermiculite, activated charcoal, and biochar were most effective in maintaining viable propagule counts, greatest shelf life when stored at 4°C (∼40-60 × 106 CFU mL⁻¹ after 3 months). These findings indicate that Streptomyces-based consortia can be efficiently utilized for sustainable disease management and growth promotion in rice.
Bacteria represent a vast and underexplored reservoir of bioactive compounds with significant anticancer potential. Numerous bacterial taxa, particularly actinomycetes, Bacillus, Pseudomonas, and marine-derived species, produce structurally diverse metabolites exhibiting cytotoxic, cytostatic, pro-apoptotic, immunomodulatory, and anti-angiogenic activities against cancer cells. Clinically established agents such as actinomycin D and bleomycin highlight the therapeutic relevance of bacterial natural products, while recent discoveries continue to expand the repertoire of bioactive polyketides, peptides, alkaloids, and proteins. These compounds act through multiple mechanisms, including DNA intercalation, induction of apoptosis, cell cycle arrest, metabolic disruption, and modulation of the tumor microenvironment. Advances in metagenomics, genome mining, and synthetic biology have enabled the identification and activation of previously silent biosynthetic gene clusters, significantly enhancing drug discovery potential. In addition to metabolite-based anticancer agents, advances in synthetic biology have enabled the development of engineered bacterial platforms capable of selectively colonizing tumors, delivering therapeutic molecules, and activating prodrug therapies within the tumor microenvironment. Despite ongoing challenges related to toxicity, limited yield, selectivity, and clinical translation, bacterial-derived compounds remain a promising frontier in oncology. This review summarizes bacterial sources, bioactive metabolites, molecular mechanisms, preclinical and clinical applications, and future prospects for developing effective and safe anticancer strategies.
Biofilms formed by foodborne pathogens pose a significant threat to food safety, as they enhance microbial resistance to disinfectants and environmental stress. Understanding the relationship between their intricate structure and macroscopic mechanical properties is crucial for developing effective control strategies. This review systematically investigates how the combined application of confocal laser scanning microscopy (CLSM) and rheology provides a powerful solution to this problem. CLSM technology reveals biofilm structure and extracellular polymeric substance (EPS) distribution through high-resolution 3D imaging and multi-component fluorescence labeling. Rheology quantitatively analyzes key mechanical parameters such as viscoelasticity and yield stress. Their integration establishes a multidimensional "structure-function-mechanics" research framework, enabling cross-scale correlations from microscopic features to macroscopic mechanical responses. Additionally, this review analyzes current biofilm removal strategies, fluorescent labeling methods for EPS components, and the composition and functions of EPS in typical bacterial biofilms. We further explore optimized technical approaches to overcome food matrix interference, applications in cleaning process optimization, and future standardization directions integrated with artificial intelligence. This comprehensive methodology provides a systematic and efficient paradigm for understanding biofilm mechanisms and developing control strategies, holding significant implications for biofilm prevention in the food industry.
