This study investigated antibacterial effects of Cinnamomum loureirii essential oil (CLEO) against three foodborne microorganisms: Escherichia coli, Staphylococcus aureus, Pseudomonas putida. GC-MS showed trans-cinnamaldehyde was CLEO's major constituent (91.18%). CLEO's minimum inhibition concentrations (MICs) were 1, 0.25, 2 mg mL-1 for E.coli, S.aureus, P.putida, respectively. With rising CLEO concentration, nucleic acid and alkaline phosphatase (AKR) leakage of three bacteria increased gradually; S. aureus had higher leakage at 1/2MIC. This demonstrated CLEO disrupts cell wall/membrane integrity, affects cell structure, changes membrane permeability of bacterial proteins. Scanning/transmission electron microscopy (SEM/TEM) showed obvious bacterial morphological/ultrastructural changes, affecting normal cell division, further confirming CLEO-induced cell wall/membrane damage. Additionally, CLEO exhibited antibiofilm activity at sub-MIC levels, inhibiting three bacteria's biofilm formation. Under 2 × MIC, inhibition rates of E. coli, S. aureus, P. putida were 85.21%, 31.29%, 72.36%, respectively. Overall, CLEO is a promising natural antibacterial agent/multifunctional food additive for food and pharmaceutical industries.

Bacillus spp. can colonize various plant tissues, promoting growth and providing biocontrol. Bacillus amyloliquefaciens GZY63 exhibits strong antagonistic activity against anthracnose pathogens in Camellia oleifera. However, its potential as a biocontrol agent in Ca. oleifera remains underexplored, mainly because knowledge regarding its colonization behavior in this host plant is limited. In this study, we used a chloramphenicol-resistant GFP-tagged GZY63 strain to assess the colonization patterns of the bacterium in Ca. oleifera through quantitative Polymerase Chain Reaction (qPCR). In particular, we investigated the bacterium's colonization dynamics across three Ca. oleifera varieties, with a focusing on differences between the root and foliar inoculation methods. The results revealed that root inoculation of B. amyloliquefaciens GZY63 resulted in significantly higher colonization efficiency and more rapid bacterial translocation to leaf tissues than its foliar inoculation. Moreover, the colonization efficiency varied among the three Ca. oleifera varieties, indicating that host genotype influences the bacterium's endophytic compatibility. Although the underlying biochemical factors and regulatory mechanisms remain unclear, our results provide valuable insights into host-microbe interactions in Ca. oleifera. These findings provide a theoretical basis for optimizing the application of B. amyloliquefaciens GZY63 as biocontrol agent in Ca. oleifera and underscore the importance of selecting compatible host genotypes for effective microbial inoculation.

Studies have demonstrated the potential for utilizing agro-industrial byproducts for Pleurotus spp. Extracts containing polysaccharides from mushrooms obtained through solid culture, as well as from mycelial biomass obtained through liquid culture (endopolysaccharides), can regulate the immune response and increase resistance to disease. Additionally, exopolysaccharide extracts obtained from culture broth can regulate the immune response. This study aimed to utilize food industry waste to produce mycelial biomass and exopolysaccharides from Pleurotus ostreatus grown in media containing apple, grape, lemon, and orange peels. P. ostreatus produced mycelial biomass in all four residues, with the highest concentration (4.91 g L-1) in grape peels. Together with lemon and orange peels, it also produced higher concentrations of exopolysaccharides (3.22 to 3.95 g L-1). The exo- and endo-polysaccharides obtained from the culture broth and the mycelial biomass cultivated with grape peels, respectively, had α-D-glucose, α-D-galactose, α-D-xylose, and galacturonic acid as their main constituents.

Group B Streptococcus (GBS) is a major cause of maternal and neonatal infections, complicated by increasing antimicrobial resistance and high virulence. In this study, from 235 vaginal swabs, 45 GBS isolates were identified and screened; 12 representative isolates (strong biofilm producers with the complete virulence gene profile) were selected for downstream analyses. Molecular analysis showed high prevalence of virulence genes (fbsA 95.5%, lmb 91.1%, pavA 88.8%, fbsB 86.6%) and biofilm-related genes (pil-1 88.8%, pil-2a 91.1%, pil-2b 84.4%). Subsequently, a Bifidobacterium bifidum (designated B. bifidum BB-6; GenBank accession number PX474696) isolated from human breast milk was used to prepare a cell-free supernatant (CFS). GC-MS analysis of the CFS of B. bifidum identified several bioactive compounds, including acetic acid, propionic acid and lactic acid. Checkerboard assays indicated synergism between CFS and penicillin/vancomycin, with a fractional inhibitory concentration index (FICI) of ≤0.5 in most cases. Sub-minimum inhibitory concentration (sub-MIC) of CFS significantly inhibited biofilm formation (P < 0.01), and quantitative real-time PCR (qRT-PCR) revealed downregulation of virulence (fbsB down -4.40-fold) and biofilm genes (pil-2b down -5.49-fold). These results highlight the therapeutic potential of B. bifidum CFS against GBS, warranting further studies to isolate active compounds and evaluate safety and efficacy in vivo.

Salmonella enterica serotype Typhimurium is a leading cause of foodborne outbreaks linked to chickens in the United States. To investigate the genomic landscape of this population, 1048 chicken-associated S. Typhimurium genomes collected nationwide were analyzed. Overall, 84.5% of isolates carried at least one antimicrobial resistance (AMR) determinant, and 34.3% displayed multidrug resistance (MDR). Sulfonamide and tetracycline resistance were most prevalent (1.03 and 1.01 AMR determinant per isolate, respectively). AMR gene (ARG) co-occurrence analysis identified strong associations among tet(A), sul1, sul2, aac(3)-VIa, and aadA1, with blaCMY-2 emerging within an expanding co-resistance cluster. ARG-plasmid replicon association analysis indicated significant positive links between sul2 and tet(A) with IncC and ColRNAI. Machine learning-driven association rule mining demonstrated that integron carriage perfectly predicted MDR, and their absence in both non-AMR and AMR but non-MDR isolates underscores their pivotal role as the defining marker of MDR. We observed a pan-genome pattern with 3855 core and 9549 accessory genes, reflecting both genomic conservation and diversity. Whole-genome phylogenetic analysis revealed two major clades, with sequence type 19 comprising 99.0% of the isolates. This research provides a comprehensive genomic characterization of chicken-associated S. Typhimurium in the United States, highlighting key AMR mechanisms relevant for food safety and public health.

Fish sauce, a seasoning commonly utilized in East Asian cuisine, is produced from fish combined with a substantial quantity of salt. However, biogenic amines (BAs) accumulation poses safety concerns in fermented fish sauce during fermentation. This study characterized Tetragenococcus halophilus A003, isolated from fish sauce, which exhibited the weakest decarboxylase gene activation and lowest BA production among the tested strains. Starter inoculation with A003 yielded minimal chemical alteration compared to natural fermentation. Cadaverine levels were substantially lower (19.1 ± 1.49 mg/l) than those in sauce fermented without a starter or with T. halophilus BCRC12250. Histamine and tyramine were undetectable in isolate A003-inoculated samples. Metagenomic analysis revealed an enrichment of low BA-producing taxa, notably Tetragenococcus and Staphylococcus, comprising 97.91% of the community. These findings suggest T. halophilus A003 confers a selective advantage for low BA microbiota during fish sauce fermentation.

Fungal pathogens continue to pose a significant challenge to global agriculture, food security, and public health, resulting in substantial crop losses, food spoilage, and the accumulation of harmful mycotoxins. The extensive reliance on synthetic chemical fungicides raises serious concerns, including the emergence of resistant fungal strains, ecotoxicity, and adverse health effects. This review highlights the potential of Bacillus spp. and Lactobacillus (LAB) in the development of natural antifungal agents. These bacterial genera are known for the production of a wide range of bioactive metabolites, such as cell-free supernatants (CFS) with potent antifungal properties, volatile organic compounds (VOCs), phenolic acids, and lipopeptides. Lactobacillus-derived compounds, particularly phenyl lactic acid, play a dual role in inhibiting fungal growth while also improving food quality and safety. Similarly, Bacillus spp. produce lipopeptides and volatile organic compounds (VOCs) that are effective against a wide range of airborne and soilborne plant pathogens. Recent studies also highlight Bacillus-mediated green synthesis of nanoparticles (e.g. silver, zinc oxide, and selenium) with potent antifungal activity. These eco-friendly nanomaterials inhibit fungal growth, disrupt biofilms, and enhance plant defenses, offering a targeted and sustainable alternative to chemical fungicides. This review advocates the integration of bacterial antifungal strategies into sustainable agriculture and post-harvest management systems.

Forage from four locations (blocks) was inoculated with 150 000 CFU/g Lactiplantibacillus plantarum DSM26571, Lactococcus lactis NCIMB30117, and Enterococcus lactis 22502 (LPEL); 150 000 CFU/g Lactococcus lactis DSM22501 and Lentilactobacillus buchneri DSM11037 (LB); or distilled water (CON), packed full (D100; 264 kg DM/m3) or half (D50; 132 kg DM/m3) density, and ensiled for 7, 14, 60, or 90 d. Data were analyzed as a randomized complete block with fixed (packing density, bacterial inoculation, storage length, and their interactions) and random (block) effects. An interaction between inoculation and storage length was observed for pH. At early storage lengths, LPEL had the lowest pH, without differences afterwards. A three-way interaction was observed for lactic acid concentration; D100 was greater than D50. An interaction between packing density and bacterial inoculation was observed for acetic acid concentration; D50-LPEL and D50-LB had the lowest acetic acid concentrations. Bacterial inoculants had minor effects, although the homofermentative inoculant decreased pH. Greater packing density increased organic acids. Thus, a combination of greater packing density and homofermentative microbial inoculants may improve fermentation and minimize storage losses. Further research is warranted to better characterize changes in the microbial community of alfalfa silage with different microbial inoculants and ensiled at different packing densities.

Salmonella has caused widespread foodborne disease risks in China. Studies suggest that plasmid-mediated quinolone resistance (PMQR) is the main route for the spread of Salmonella's drug resistance. To establish a method for rapid detection of the genus gene invA carried by foodborne Salmonella and four PMQR genes carried by drug-resistance strains based on multiplex PCR combined with liquid chip technology. The detection limits, sensitivity, specificity, and repeatability of the method were evaluated. Our findings revealed that in terms of detection sensitivity, this method can detect the invA and qnrS with a limit as low as 5 CFU/mL. The detection limits for aac(6')-Ib-cr, oqxA, and oqxB genes were 25 CFU/mL, 10 CFU/mL, and 10 CFU/mL, respectively. In terms of specificity, no positive signals were detected for the nontarget bacteria strains and the negative control. In the repeatability experiments, the coefficient of variation (CV) for all target gene detections was <5%. In the simulation sample verification, the concordance rate with the results of conventional PCR reached 100%. Therefore, this method can provide technical support for the detection of foodborne Salmonella and PMQR genes, as well as the monitoring of drug resistance.

Continuous cultivation can lead to soil nutrient imbalances and have adverse effects on soil rhizosphere microorganisms. This study investigated the impact of continuous cultivation of Salvia miltiorrhiza on rhizosphere soil microbial communities by comparing four planting durations: CK (no cultivation), D1 (annual planting), D2 (two consecutive years), and D3 (three consecutive years). High-throughput sequencing technology was used to analyze changes in rhizosphere soil microbial communities. The results revealed that the planted soil of Salvia miltiorrhiza exhibited lower abundances of beneficial bacteria such as Bacillus and Acidothermus compared to CK. The pathogenic fungus Fusarium was found to found to have the highest abundance in D2 soil. Linear discriminant analysis effect size analysis at the genus level identified several biomarkers, including g_Candidatus_Solibacter, g_Sphingomonas, g_RB41, and g_Chitinophaga as bacterial markers, while g_Talaromyces, g_Thermomyces, g_Trichothecium, g_Solicoccozyma, and g_Pseudopithomyces as fungal markers. Correlation analysis between these microbial markers and environmental factors showed that total nitrogen had a significant positive correlation with bacteria g_Candidatus_Solibacter and fungi g_Solicoccozyma, but a significant negative correlation with bacteria g_Sphingomonas and g_RB41. Soil organic matter showed a negative correlation with g_Sphingomonas; g_Candidatus Solibacter was negatively correlated with g_Sphingomonas.

The presence of multidrug-resistant Escherichia coli strains in healthy cattle represents a major threat to public health because they can act as a reservoir of resistance genes. Little is known about the antimicrobial susceptibility profile and genetic characteristics of E. coli strains isolated from healthy calves in Panama and Central America. We conducted a cross-sectional study evaluating isolates from healthy calves in Panama. Antimicrobial susceptibility was determined against 15 antibiotics, and strains were classified as resistant, intermediate, or susceptible. 120 E. coli isolates were obtained, of which 61% were resistant to at least one antibiotics analyzed, while only 39% were susceptible. The most frequent resistance was to tetracycline (36%) and ampicillin (34%). Fifteen percent of the isolates (18/120) were multidrug-resistant, and 2% presented an extended spectrum β-lactamase phenotype; blaTEM (22%) and blaCTX-M (10%) genes were detected, with the Extended-Spectrum β-lactamases phenotype observed in isolates carrying blaCTX-M. Plasmid-mediated quinolone resistance gene qnrA (10%), qnrB (13%), and qnrS (23%) were detected among the isolates. Our results suggest that a significant proportion of antimicrobial resistance occurs in intestinal strains of E. coli isolated from healthy calves in Panama.

Avian pathogenic E. coli (APEC) is a leading cause of colibacillosis in poultry worldwide. Molecular genotyping using polymerase chain reaction (PCR) is a rapid and effective way to separate Avian Pathogenic E. coli (APEC) from commensal E. coli in poultry. This study assessed the accuracy of four different molecular genotyping panels in a head-to-head comparison, using 250 APEC isolates from diseased birds and 106 fecal E. coli (AFEC) isolates from healthy birds. The cut-off value of all four panels was observed and optimal cut-off values with sensitivity and specificity were suggested in this study. The results found that the APECtyper panel had the highest sensitivity (81.2%) being significantly different to the other 3 panels, while the 9-gene panel demonstrated the highest specificity (89.6%). The cut-off of the 13-gene panel was optimized with a predicted probability of 12%, balancing sensitivity (57.2%) and specificity (75.5%), which was used in this study. Overall, three of the four panels except the 13-gene panel showed good performance with distinct strengths. This research provides valuable insights, but further studies are needed to confirm the reliability and reproducibility of these assays across different geographical regions, various poultry populations, and diverse field conditions.

Mango (Mangifera indica) is an economically significant fruit crop in Sri Lanka. Recently, a new disease, stem bark blackening, emerged on mango in the country, causing concerns among growers. This study aimed to identify the pathogen and explore effective in vitro strategies for its management. Field observations revealed characteristic black, irregularly shaped patches encircling the mango stems and branches. Morphological studies and phylogenetic analysis of the ITS region confirmed the pathogen as Aspergillus spp. In vitro assays demonstrated 92.4% fungal inhibition by Trichoderma viride. Among the tested plant extracts, Azadirachta indica showed the highest antifungal activity (72.3% colony size reduction compared to control). Commercial fungicides tested were less effective. In vitro biocontrol activity of T. viride and A. indica extract is more effective than conventional fungicides, highlighting their potential in integrated disease management of mango stem bark blackening disease.

Faecal microbiota transplantation (FMT) is an effective treatment for Clostridioides difficile infection, but current FMT product manufacturing protocols are time-sensitive, labour-intensive, and require specialized staff. Delays can compromise microbial viability and product quality. Efficient storage methods for partially processed FMT material could therefore improve flexibility and streamline production. Therefore, this feasibility study evaluated the impact of storage conditions on live bacteria in partially processed donations containing glycerol as a cryoprotectant. Using culturomics, we assessed the concentration and composition of live bacteria immediately after half-way processing (reference, 0 h) or after storage at 5°C (refrigerator) or -80°C (freezer) for up to 72 h. Each FMT product was initially processed from one faecal donation provided by one of four screened donors. Storage at -80°C preserved the amount and composition of live bacteria in partially processed products. In contrast, storage at 5°C led to reduced bacterial concentrations and compositional shifts, particularly affecting the abundance of Bacteroides spp. These results highlight the potential adverse effects of refrigerator storage on bacterial viability, suggesting it may not be suitable for maintaining the quality of partially processed encapsulated FMT products. Freezer storage, however, emerged as a reliable method to preserve the content of live bacteria for at least 72 h.

Bacillus thuringiensis Berliner is a well-known biocontrol agent that produces insecticidal crystal proteins encoded by cry genes, which are effective against various insect orders. However, only a limited number of B. thuringiensis strains are known to be toxic to white grubs, a major pest of coconut, groundnut, and sugarcane that can cause up to 70%-80% yield losses. In this study, five indigenous B. thuringiensis strains, isolated from soil and Anomala elata cadavers, were screened for toxicity against second-instar Holotrichia serrata larvae, a highly destructive white grub species. Among them, the strain NBAIR BtAe exhibited the highest toxicity with an LC₅₀ of 115.36 μg mL-1. Whole-genome sequencing of NBAIR BtAe revealed a 5.67 Mb circular chromosome with 35.64% GC content. BtToxin_Digger analysis identified a novel cry gene with 39.57% similarity to cry21Aa2, along with other virulence genes including zwa6, zwa5A, chitinase C, inhA1, inhA2, bmp1, spp1Aa1, enhancin, and tpp80Ab1-like. These genes were validated through PCR. Additionally, genes encoding secondary metabolites such as lanthipeptides, paenilamicin, petrobactin, bacillibactin, and fengycin were detected. The presence of diverse pesticidal and antimicrobial genes highlights the potential of NBAIR BtAe as a promising candidate for bioinsecticide development targeting H. serrata in integrated pest management programs.

This study aimed to evaluate the antimicrobial susceptibility and clonal relatedness of Klebsiella pneumoniae isolates recovered from the milk of cows with mastitis in a large-scale study that clinical severity was scored. A total of 48 K. pneumoniae complex isolates were subjected to in vitro antimicrobial susceptibility tests (AST), PCR for carbapenemase-encoding genes, and molecular typing by pulsed-field gel electrophoresis (PFGE). Thirteen isolates, selected by PFGE type and AST results were subjected to whole-genome sequencing (WGS) for the identification of sequence types (STs) and the detection of antimicrobial resistance genes and virulence-encoding genes. A total of 39 different PFGE restriction profiles were identified. Thirteen different STs, including two novel STs, were identified among the 13 sequenced strains. The blaCTX-M-8, qnrE1, aadA2, cmlA4, dfrA15, sul1, tetA, and tetB genes were identified. Two isolates presented the yersiniabactin-encoding gene ybtAEPQSTUX. Klebsiella pneumoniae isolates from the milk of cows with mastitis clinically scored revealed high genetic diversity, according to both PFGE and MLST analysis, as well as harboring resistance genes commonly found in human clinical isolates.

This study aimed to explore the cultivable actinobacterial diversity in Krem Dam cave, Meghalaya, India, and to evaluate their biotechnological potential through antimicrobial activity, plant growth-promoting traits, and metabolic pathway prediction. Sediment samples were collected from five locations within the cave, pretreated, and cultured on selective media to isolate actinobacteria. Isolates were characterized morphologically, physiologically, and chemotaxonomically, followed by 16S rDNA sequencing for molecular identification. The PAPRICA pipeline was used to predict metabolic pathways from 16S rDNA sequences. Antimicrobial activity was assessed against Gram-positive, Gram-negative bacteria, and Candida species using cross-streak and agar-well diffusion methods, while biosynthetic gene clusters (PKS-I, PKS-II, and NRPS) were screened via polymerase chain reaction (PCR). Plant growth-promoting (PGP) traits, including IAA production, phosphate solubilization, siderophore activity, and nitrogen fixation were evaluated, along with antagonism against phytopathogens and seedling vigor in tomato. Forty-eight isolates were identified, predominantly Streptomyces thermocarboxydus-related strains, with one Amycolatopsis species. Seventy-seven percent harbored at least one biosynthetic gene cluster, and significant antimicrobial activity was observed, particularly against Gram-positive bacteria. Several isolates exhibited multiple PGP traits, and two (KD-21, KD-29) enhanced tomato seedling vigor. The study concludes that Krem Dam cave harbors diverse, bioactive actinobacteria with promising applications in pharmaceuticals and sustainable agriculture, warranting further metabolomic and genomic investigations.

Mycoplasma pneumonia (MP) is a common pathogen of human respiratory infections and one of the leading causes of community-acquired pneumonia. Early and rapid diagnosis of its infection is crucial for clinical treatment decisions. In this study, we innovatively combined recombinase polymerase amplification with Pyrococcus furiosus Argonaute protein (PfAgo) to establish a novel molecular diagnostic method for MP. This assay demonstrated high specificity with no cross-reactivity with other common respiratory pathogens. The limit of detection was 2 × 104 copies μl-1. Furthermore, evaluation using 37 clinical samples showed 100% specificity and 86.36% sensitivity compared to quantitative real-time polymerase chain reaction (qPCR). The entire workflow, including sample preparation, can be completed within 2.5 h and requires only basic instrumentation. This method holds great potential for application in primary healthcare settings and resource-limited regions.

Mycoplasma genitalium is challenging to work with and new methods are needed to study this bacterium directly in clinical samples. This study designed and validated a proof-of-concept polymerase chain reaction (PCR)-based 'tiling' methodology to sequence M. genitalium genomes. Primers were designed to produce 2.5 kb amplicons covering the 580 kb genome with a minimum overlap of 100 bp. Analysis was performed using the laboratory strain G37 and a clinical isolate. Amplicons were sequenced on the Oxford Nanopore MinION using ligation sequencing. Reads were mapped to a reference to produce a consensus genome. A total of 262 primer pairs were designed and amplification was successful for 99.5% (261/262) of 2.5 kb amplicons, with G37 genome coverage of 99.5% (mean read depth, 1973X). Using larger 5kb amplicons, amplification was successful for 92.4% (121/131) of primer pairs, with a coverage of 92.2% (mean read depth, 223X). When validated on a clinical isolate, 98.3% coverage was achieved (read depth, 443X). In conclusion, this study developed a PCR-based tiling approach to whole genome sequencing of M. genitalium by designing and validating a set of 262 primer pairs.

Enzymatic activity potency in Candida albicans is depends on different pH levels. This study investigates the importance of matching in vitro pH conditions to those encountered at infection sites. A total of 20 vulvovaginitis C. albicans isolates were investigated for phospholipase, proteinase, and esterase activities. The activities were measured at three pH levels (4, 5, and 7) representing for the healthy vaginal range, potential infection condition, and a standard in vitro reference point, respectively. Specific media were used to assess enzyme activity. Phospholipase and proteinase activity were significantly higher at acidic pH compared to neutral pH. In contrast, esterase activity showed a slight increase at neutral pH. Analysis revealed significant differences in enzyme activity between pH 5 and 7, highlighting the importance of using pH-relevant conditions for studying Candida virulence. This study reveals that the acidic vaginal pH characteristic of C. albicans infections significantly enhances the activity of its key damaging enzymes. This highlights the limitations of using a standard pH protocol for enzyme activity analysis. By employing pH-mimicking conditions, future research can unlock a deeper understanding the actual C. albicans virulence that might occur in site of infection. Data analysis should consider beyond this factor.

The global increase in fungal infections and antifungal resistance has drawn attention to environmental sources of potentially pathogenic yeasts. This study investigated the fungal load and antifungal susceptibility of yeasts isolated from fresh fruits (strawberries, bananas, guavas, and apples) grown under organic and conventional farming systems. Samples were analyzed for total yeast counts (CFU g-1) and for quantification of yeasts capable of growing on medium supplemented with fluconazole (FLU) (8 and 64 µg ml-1), followed by phenotypic identification and antifungal susceptibility testing via broth microdilution. The highest fungal loads were observed in strawberries across both cultivation systems. While no differences were found between organic and conventional fruits, variation in total and FLU-tolerant yeast counts was observed between fruit types. Among 29 presumptively FLU-resistant isolates, 23 were confirmed as resistant by minimum inhibitory concentration testing. All isolates were resistant to itraconazole, and most showed cross-resistance to voriconazole. Identified species included Meyerozyma guilliermondii, Pichia kudriavzevii, Trichomonascus ciferrii, Papiliotrema laurentii, and Rhodotorula mucilaginosa, many of which are associated with invasive fungal infections. These findings highlight the potential of fresh fruits to act as reservoirs for opportunistic and multidrug-resistant yeasts. Enhanced microbiological surveillance of plant-derived foods is recommended as part of integrated public health strategies, particularly within the One Health framework.