Drug-loaded lipid nanoparticles improve the removal rates of the Staphylococcus aureus biofilm.

Biofilms of the foodborne pathogen Staphylococcus aureus show improved resistance to antibiotics and are difficult to eliminate. To enhance antibacteria and biofilm dispersion via extracellular matrix diffusion, a new lipid nanoparticle was prepared, which employed a mixture of phospholipids and a 0.8% surfactin shell. In the lipid nanoparticle, 31.56 µg mL-1 of erythromycin was encapsulated. The lipid nanoparticle size was approximately 52 nm and the zeta-potential was -67 mV, which was measured using a Marvin laser particle size analyzer. In addition, lipid nanoparticles significantly dispersed the biofilms of S. aureus W1, CICC22942, and CICC 10788 on the surface of stainless steel, reducing the total viable count of bacteria in the biofilms by 103 CFU mL-1 . In addition, the lipid nanoparticle can remove polysaccharides and protein components from the biofilm matrix. The results of laser confocal microscopy showed that the lipid nanoparticles effectively killed residual bacteria in the biofilms. Thus, to thoroughly eliminate biofilms on material surfaces in food factories to avoid repeated contamination, drug-lipid nanoparticles present a suitable method to achieve this.

Prevention of high-fat-diet-induced obesity in mice by soluble dietary fiber from fermented and unfermented millet bran.

Obesity-related diabetes, cardiovascular disease, and hypertension pose many risks to human health. Thus, mice on a high-fat diet were gavaged with millet bran (unfermented/fermented) soluble dietary fiber (RSDF/FSDF, 500 mg·kg-1) for 10 weeks in current research, and then evaluated the various biological indicators. These findings revealed that RSDF and FSDF supplements could prevent fat synthesis by inhibiting sterol regulatory element-binding protein-1c gene expression. The RSDF supplements can also accelerate fat catabolism through enhanced the mRNA expression levels of adipose triglyceride lipase and peroxisome proliferator-activated receptor α. FSDF supplements can prevent obesity by decreasing 3-hydroxy-3-methyl-glutaryl-CoA reductase expression and increasing cholesterol 7α-hydroxylase expression. Moreover, FSDF also controls obesity development by lowering total cholesterol and low-density lipoprotein cholesterol levels in the blood, triglyceride, total cholesterol, and bile acid levels in the liver. Notably, FSDF supplements can promote Bacteroides and Prevotella propagation; excretive propionic acid binds to free fatty acid receptor 2/3 and then stimulates intestinal epithelial cells to generate glucagon-like-peptide-1 and peptide YY, which can reduce food and energy intake and ultimately prevent obesity. All evidence suggests that FSDF supplements play a crucial role in preventing obesity.

Establishment of real-time fluorescence and visual LAMP for rapid detection of Escherichia coli O157:H7 and kits construction.

Escherichia coli O157:H7 is a common pathogenic bacterium in food and water that can pose a threat to human health. The aim of this study was to develop loop-mediated isothermal amplification (LAMP) method for the detection of E. coli O157:H7 in food based on the specific gene Ecs_2840 and to construct rapid detection kits based on the established methods. Specifically, we established two methods of real-time fluorescent LAMP (RT-LAMP) and visual LAMP with calcein as an indicator. In pure bacterial culture, the cell sensitivity and genomic sensitivity of the RT-LAMP kit were 8.8 × 100 CFU ml-1 and 4.61 fg µl-1, respectively. The sensitivity of the visual LAMP kit was 2.35 × 100 CFU ml-1 and 4.61 fg µl-1. Both kits had excellent specificity and anti-interference performance. In addition, milk inoculated with 2.26 × 100 CFU ml-1E. coli O157:H7 could be detected within the reaction time after enrichment for 3 h. The results showed that the LAMP kits were rapid, sensitive, and specific for the detection of E. coli O157:H7 in food and had good application prospects in food safety surveillance.

Effects of the deletion and substitution of thioesterase on bacillomycin D synthesis.

OBJECTIVES: The importance of thioesterase domains on bacillomycin D synthesis and the ability of different thioesterase domains to selectively recognize and catalyze peptide chain hydrolysis and cyclization were studied by deleting and substituting thioesterase domains. RESULTS: No bacillomycin D analogs were found in the thioesterase-deleted strain fmbJ-ΔTE, indicating that the TE domain was essential for bacillomycin D synthesis. Then the thioesterase in bacillomycin D synthetases was replaced by the thioesterase in bacillomycin F, iturin A, mycosubtilin, plipastatin and surfactin synthetases. Except for fmbJ-S-TE, all others were able to synthesize bacillomycin D homologs because a suitable recombination site was selected, which maintained the integrity of NRPSs. In particular, the yield of bacillomycin D in fmbJ-IA-TE, fmbJ-M-TE and fmbJ-P-TE was significantly increased. CONCLUSION: This study expands our understanding of the TE domain in bacillomycin D synthetases and shows that thioesterase has excellent potential in the chemical-enzymatic synthesis of natural products or their analogs.

The global regulator SpoVG regulates Listeria monocytogenes biofilm formation.

Biofilms provide a suitable environment for L. monocytogenes and are the cause of enormous risks in the food industry. SpoVG is a global regulatory factor that plays a vital role in physiological activity of L. monocytogenes. We constructed spoVG mutant strains to investigate the effects of these mutants on L. monocytogenes biofilms. The results show that L. monocytogenes biofilm formation was decreased by 40%. Furthermore, we measured biofilm related phenotypes to study the regulation of SpoVG. The motility capacity of L. monocytogenes was found to decrease after the deletion of spoVG. The cell surface properties changed in the spoVG mutant strains, with an increase in both the cell surface hydrophobicity and the auto-aggregation capacity after spoVG deletion. SpoVG mutant strains were found to be more sensitive to antibiotics, and had a reduced tolerance to inappropriate pH, salt stress and low temperature. The RT-qPCR results showed that SpoVG effectively regulated the expression of genes related to quorum sensing, flagella, virulence and stress factors. These findings suggest that spoVG has potential as a target to decrease biofilm formation and control L. monocytogenes contamination in the food industry.

Transcriptomic analysis reveals that Bacillomycin D-C16 induces multiple pathways of disease resistance in cherry tomato.

BACKGROUND: Bacillomycin D-C16 can induce resistance in cherry tomato against pathogens; however, the underlying molecular mechanism is poorly understood. Here, the effect of Bacillomycin D-C16 on induction of disease resistance in cherry tomato was investigated using a transcriptomic analysis. RESULTS: Transcriptomic analysis revealed a series of obvious enrichment pathways. Bacillomycin D-C16 induced phenylpropanoid biosynthesis pathways and activated the synthesis of defense-related metabolites including phenolic acids and lignin. Moreover, Bacillomycin D-C16 triggered a defense response through both hormone signal transduction and plant-pathogen interactions pathways, and increased the transcription of several transcription factors (e.g., AP2/ERF, WRKY and MYB). These transcription factors might contribute to the further activated the expression of defense-related genes (PR1, PR10 and CHI) and stimulated the accumulation of H2O2. CONCLUSION: Bacillomycin D-C16 can induce resistance in cherry tomato by activating the phenylpropanoid biosynthesis pathway, hormone signal transduction pathway and plant-pathogen interactions pathway, thus activating comprehensive defense reaction against pathogen invasion. These results provided a new insight into the bio-preservation of cherry tomato by the Bacillomycin D-C16.

Effect and regulation of fatty acids on bacillomycin D synthesis.

Bacillomycin D is a cyclic antimicrobial lipopeptide that has excellent antifungal effects, but its application is limited due to its low yield. At present, it is not clear whether fatty acids regulate the synthesis of bacillomycin D. Therefore, the effects of nine fatty acids on the yield of bacillomycin D produced by Bacillus amyloliquefaciens fmbJ were studied. The results showed that sodium propionate, propionic acid, and butyric acid could increase the yield of bacillomycin D by 44, 40, and 10%, respectively. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expression levels of bacillomycin D synthesis gene, signaling factors and genes related to fatty acid metabolism, so as to explore the mechanism of sodium propionate regulating bacillomycin D synthesis. In conclusion, sodium propionate could accelerate the tricarboxylic acid cycle and promoted spore formation, cell movement, the secretion of extracellular protease and the transcription of bacillomycin D synthesis gene by upregulating the expression of signal factors degU, degQ, sigH, sigM and spo0A and ultimately promoted the synthesis of bacillomycin D. In this study, the mechanism of sodium propionate increasing bacillomycin D production was explored from multiple perspectives, which provided theoretical support for the large-scale production of bacillomycin D and was expected to promote its wide application in food, agriculture and medicine fields.

Construction and optimization of a multiplex PMAxx-qPCR assay for viable Bacillus cereus and development of a detection kit.

In this study, a PMAxx-qPCR method for the detection and quantification of viable Bacillus cereus (B. cereus) was established based on the cesA gene that is involved in cereulide synthesis, enterotoxin gene bceT and hemolytic enterotoxin gene hblD combined with modified propidium monoazide (PMAxx). The sensitivity detection limit of the method was as follows: the DNA extracted by the kit reached 140 fg/µL, and the bacterial suspension without enrichment reached 2.24 × 101 CFU/mL; 14 nonB. cereus strains of the 17 tested strains all tested as negative, whereas the 2 strains of B. cereus carrying the target virulence gene(s) could be accurately detected. In terms of application, we assembled the constructed PMAxx-qPCR reaction into a detection kit and evaluated its application performance. The results showed that the detection kit has high sensitivity, strong anti-interference capability, and has good application potential. The purpose of this study is to provide a reliable detection method for the prevention and traceability of B. cereus infections.

Biochemical and molecular regulatory mechanism of the pgpH gene on biofilm formation in Listeria monocytogenes.

AIMS: PgpH gene has an important regulatory role on bacterial physiological activity, but studies on its regulation mechanism on biofilm formation of Listeria monocytogenes are lacking. Our aim was to investigate the effect of pgpH gene deletion on biofilm formation in L. monocytogenes. METHODS AND RESULTS: The ΔpgpH deletion strain of L. monocytogenes LMB 33 426 was constructed by homologous recombination. Deletion of the pgpH gene resulted in a significant reduction in biofilm formation. The swimming ability of the ΔpgpH strain on semisolid plates was unchanged compared to the wild-type strain (WT), and the auto-aggregation capacity of L. monocytogenes was decreased. RNA-seq showed that ΔpgpH resulted in the differential expression of 2357 genes compared to WT. pgpH inactivation resulted in the significant downregulation of the cell wall formation-related genes dltC, dltD, walK, and walR and the flagellar assembly related genes fliG and motB. CONCLUSIONS: This study shows that the deletion of pgpH gene regulates biofilm formation and auto-aggregation ability of L. monocytogenes by affecting the expression of flagellar assembly and cell wall related genes. pgpH has a global regulatory effect on biofilm formation in L. monocytogenes.

Separation, characterization and anti-inflammatory activities of galactoglycerolipids from Perilla frutescens (L.) Britton.

The study was to optimize the separation procedures, characterize the galactoglycerolipids and explore their anti-inflammatory activities. Two monogalactosyldiacylglycerols (MGDGs) and three digalactosyldiacylglycerols (DGDGs) from Perilla frutescens (L.) Britton were obtained through one-step silica gel column chromatography and preparative high-performance liquid chromatography with evaporative light scattering detection (HPLC-ELSD). The presence of additional MGDG (1-O-9Z,12Z,15Z-octadecatrienoyl-2-O-7Z,10Z,13Z-hexadecatrienoyl-3-O-(ß-D-galactopyranosyl)-sn-glycerol) and DGDG (1-O-9Z,12Z-octadecadienoyl-2-O-9Z,12Z,15Z-octadecatrienoyl-3-O-(ß-D-galactopyranosyl-(1'→6'')-α-D-galactopyranosyl)-sn-glycerol) was concluded for the first time in perilla, by liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR). In lipopolysaccharide (LPS)-induced RAW264.7 cells, five galactoglycerolipids exhibited good inhibitory activities against nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) gene expression in a dose-dependent manner, suggesting that fatty acid chain length and unsaturation degree affected their anti-inflammatory activities.

LsrR-like protein responds to stress tolerance by regulating polysaccharide biosynthesis in Lactiplantibacillus plantarum.

In addition to their biological functions, polysaccharides assist Lactiplantibacillus plantarum in resisting harsh conditions. To enhance the polysaccharide biosynthesis and increase the survival of L. plantarum in gut environment. We analyzed the transcriptional regulators that regulated the polysaccharide biosynthesis. A new transcriptional inhibitor, LsrR (UniProtKB: Q88YH7), had been identified, which repressed polysaccharide synthesis by binding to the polysaccharide synthesis promoter cps4A-J (Pcps4A-J). The EPSs and CPSs production of L. plantarum 163 was reduced by 42 % and 36 % (p < 0.05), respectively, when lsrR was overexpressed. Furthermore, alkaline shock proteins Asp2 and Asp1, heat shock protein Hsp3, and an autoinducer-2 (AI-2) related quorum-sensing regulator Rrp6 recovered the synthesis of polysaccharides to 50, 33, 55, and 60 %, respectively, by inhibiting the LsrR activity. This suggested that LsrR regulates polysaccharide synthesis in response to external stress signals such as pH, temperature, and AI-2 concentration. Finally, we showed that polysaccharides increased the survival rate of L. plantarum (Lp163-ΔlsrR) by 2.1 times during lyophilization and enhanced its tolerance to pH 2.0 and 0.2 % bile salts by 15.3 and 60 times due to increased capsular thickness and enhanced the autoaggregation. We provide critical data regarding Lactobacillus survival during preservative lyophilization and under gastrointestinal conditions.

Prevalence, antimicrobial resistance and genetic characterization of Vibrio parahaemolyticus isolated from retail aquatic products in Nanjing, China.

Vibrio parahaemolyticus, is one of the most frequently reported pathogenic microorganisms that causes foodborne illnesses worldwide. The aims of the current study were to determine the prevalence, virulence genes, antimicrobial resistance, biofilm formation ability (BFA) and genetic characterization of V. parahaemolyticus recovered from retail aquatic products in Nanjing, China. There were 131 samples (71.6%) that tested positive for V. parahaemolyticus. The thermostable direct hemolysin-related hemolysin (trh) gene was found in two isolates (1.5%). Antimicrobial susceptibility tests showed that 46.6% of isolates were multidrug resistant. High resistance was observed to ampicillin (100%), cephalosporin (99.2%), trimethoprim/sulfamethoxazole (38.2%) and tetracycline (16.0%). Ten resistance patterns were found. The crystal violet staining assay showed that 35.1% had strong BFA, and 52.7% had intermediate BFA; notably, five (3.8%) extremely strong BFA strains were obtained from wet markets. According to whole genome sequencing analysis of 59 randomly selected isolates, 46 sequence types (STs) were identified, including 22 novel STs, and ST1042 was the dominant sequence type. It is clear that the V. parahaemolyticus population exhibits a high level of genetic variation. Our findings provide comprehensive insight into the prevalence and phylogenetic relationship of V. parahaemolyticus in aquatic products, suggesting potential hazards to consumers in Nanjing.

Translocation of subunit PPSE in plipastatin synthase and synthesis of novel lipopeptides.

Nonribosomal peptide synthase (NRPS) is a unique molecular assembly mechanism with high hybridity. Its recombination is conducive to the development of novel lipopeptides. However, there are few reports on NRPS subunit recombination of plipastatin at present. In this paper, plipastatin synthase was modified by the forward movement of subunit PPSE and the replacement of the communication-mediating (COM) domain. The results showed that ppsABE, a new assembly line, could synthesize novel lipopeptides such as cycle pentapeptide (C16-18ß-OHFA-E-O-cyclo(Y-T-I), and its antimicrobial activity against Rhizopus stolonifer and Staphylococcus aureus was better than that of plipastatin. However, the reactivity of ppsABCE disappeared, but the substitution of COMD ppsC/COMA ppsD or COMD ppsD/COMA ppsE for COMD ppsC/COMA ppsE could restore its activity and conduct the biosynthesis of linear hexapeptide (C16-17ß-OHFA-E-O-Y-T-E-A/V) and heptapeptide (C17-18ß-OHFA-E-O-Y-T-E-A-I). Collectively, these findings indicated that the COM donor domain at the C-terminus of PPSB could communicate with the COM acceptor domain at the N-terminus of PPSE and that the compatible COM domain is an important tool for communication between nonpartner subunits. Moreover, the integrity and selective compatibility of the COM acceptor domain of subunit PPSE are essential to promote the interaction between PPSE and other subunits. This work further complemented the rules of NRPS subunit recombination and provided a theoretical basis for the development of novel high-efficiency lipopeptides.

Thermostability Improvement of L-Asparaginase from Acinetobacter soli via Consensus-Designed Cysteine Residue Substitution.

To extend the application range of L-asparaginase in food pre-processing, the thermostability improvement of the enzyme is essential. Herein, two non-conserved cysteine residues with easily oxidized free sulfhydryl groups, Cys8 and Cys283, of Acinetobacter soli L-asparaginase (AsA) were screened out via consensus design. After saturation mutagenesis and combinatorial mutation, the mutant C8Y/C283Q with highly improved thermostability was obtained with a half-life of 361.6 min at 40 °C, an over 34-fold increase compared with that of the wild-type. Its melting temperature (Tm) value reaches 62.3 °C, which is 7.1 °C higher than that of the wild-type. Molecular dynamics simulation and structure analysis revealed the formation of new hydrogen bonds of Gln283 and the aromatic interaction of Tyr8 formed with adjacent residues, resulting in enhanced thermostability. The improvement in the thermostability of L-asparaginase could efficiently enhance its effect on acrylamide inhibition; the contents of acrylamide in potato chips were efficiently reduced by 86.50% after a mutant C8Y/C283Q treatment, which was significantly higher than the 59.05% reduction after the AsA wild-type treatment. In addition, the investigation of the mechanism behind the enhanced thermostability of AsA could further direct the modification of L-asparaginases for expanding their clinical and industrial applications.

Comparison of osteogenesis of bovine bone xenografts between true bone ceramics and decalcified bone matrix.

Xenograft bone scaffolds have certain advantages such as mechanical strength, osteoinductive properties, sufficient source and safety. This study aimed to compare osteogenesis of the two main bovine bone xenografts namely true bone ceramics (TBC) and decalcified bone matrix (DBM), and TBC or DBM combined with bone morphogenetic protein (BMP)-2 (TBC&BMP-2 and DBM&BMP-2). The characteristics of TBC and DBM were investigated by observing the appearance and scanning electron microscopic images, examining mechanical strength, evaluating cytotoxicity and detecting BMP-2 release after being combined with BMP-2 in vitro. The femoral condyle defect and radial defect models were successively established to evaluate the performance of the proposed scaffolds in repairing cortical and cancellous bone defects. General observation, hematoxylin and eosin (HE) staining, mirco-CT scanning, calcein double labeling, X-ray film observation, three-point bending test in vivo were then performed. It indicated that the repair with xenograft bone scaffolds of 8 weeks were needed and the repair results were better than those of 4 weeks whatever the type of defects. To femoral condyle defect, TBC and TBC&BMP-2 were better than DBM and DBM&BMP-2, and TBC&BMP-2 was better than TBC alone; to radial defect, DBM and DBM&BMP-2 were better than TBC and TBC&BMP-2, and DBM&BMP-2 was better than DBM alone. This study has shown that TBC and DBM xenograft scaffolds can be more suitable for the repair of cancellous bone and cortical bone defects for 8 weeks in rats, respectively. We also have exhibited the use of BMP-2 in combination with DBM or TBC provides the possibility to treat bone defects more effectively. We thus believe that we probably need to select the more suitable scaffold according to bone defect types, and both TBC and DBM are promising xenograft materials for bone tissue engineering and regenerative medicine. Graphical abstract.

Transcriptome sequencing reveals the difference in the expression of biofilm and planktonic cells between two strains of Salmonella Typhimurium.

Salmonela enterica serovar Typhimurium (S. Typhimurium) is a food-borne pathogen that can form biofilms to increase its resistance to the external environment. Through the detection of biofilm of several S. Typhimurium strains in this study, strain CDC3 with strong biofilm forming capacity and strain CVCC3384 with weak biofilm forming capacity were identified. The genes expressed in planktonic and biofilm cells of two S. Typhimurium strains were analysed by transcriptome sequencing. Results showed that the genes related to the signal transduction pathway were upregulated and genes related to motility were downregulated in strain CDC3. By comparing biofilms and planktonic cells of the two strains, we found that CDC3 regulates biofilm formation mainly through the two-component system kdpABC, while strain CVCC3384 does so mainly through motility and quorum sensing. This study revealed regulation mechanism of biofilms formation between different biofilm forming capacity strains, and provided a theoretical basis for subsequent research.

Plantaricin A reverses resistance to ciprofloxacin of multidrug-resistant Staphylococcus aureus by inhibiting efflux pumps.

Overexpression of Staphylococcus aureus efflux pumps is commonly associated with antibiotic resistance, causing conventional antibiotics to be unsuccessful in combating multidrug-resistant bacterial infections. Reducing the activity of the efflux pump is an urgently required to tackle this problem. Here, we found that plantaricin A (PlnA), an antimicrobial peptide derived from Lactobacillus plantarum, had a synergistic effect with ciprofloxacin (CIP), reducing the IC90 of CIP by eight times. Subsequently, changes in membrane permeability, membrane potential, and reactive oxygen species (ROS) were determined; changes that did not explain the synergistic effect were previously observed. Ethidium bromide intake and efflux experiments showed that PlnA inhibited the function of the efflux pump by binding it and altering the structure of MepA, NorA, and LmrS. Then, a series of PlnA mutants were designed to explore the underlying mechanism; they showed that the charge and foaming of PlnA were the predominant factors affecting the structure of NorA. In a skin wound infection model, PlnA significantly reduced the dose of CIP, relieved inflammation, and promoted wound healing, indicating that PlnA and CIP synergy persisted in vivo. Overall, PlnA reduced the use of CIP for combination therapy, and allowing the continued used of CIP to kill MDR S. aureus. Multidrug-resistant Staphylococcus aureus threatens our life as a tenacious pathogen, which causes infections in hospitals, communities and animal husbandry. Various studies have showed that efflux pump inhibitors (EPIs) have been considered potential therapeutic agents for rejuvenating the activity of antibiotics. Unfortunately, small molecule EPIs exhibit several side effects that limit their use for clinical application. The present study showed a new EPI (plantaricin A) produced by Lactobacillus plantarum, which has low cytotoxicity and haemolysis and powerful inhibitory activity on efflux pumps. Therefore, it helps the design of new EPIs and controls the infection of MDR S. aureus.

Designed symmetrical ß-hairpin peptides for treating multidrug-resistant salmonella typhimurium infections.

The rapid emergence and prevalence of multidrug-resistant salmonellosis lack effective therapies, which causes epidemic health problems and stimulates the development of antimicrobials with novel modes of action. In this research, 10 short symmetrical ß-hairpin peptides are synthesized by combining the ß-turn of Leucocin-A with recurring hydrophobic and cationic amino acid sequences. Those designed peptides exhibited potent antibacterial activities against drug-susceptible and drug-resistant Salmonella. One of the 10 peptides, WK2 ((WK)2CTKSGC(KW)2), displayed best cell selectivity towards Salmonella cells over macrophages and erythrocytes in a co-culture model. Fluorescent measurements and microscopic observations reflected that WK2 exerted its antimicrobial activity through a membrane-lytic mechanism. Moreover, the ß-hairpin peptides can bind to endotoxin (LPS) and suppress the production of LPS-induced proinflammatory cytokines in RAW264.7 cells, indicating as a potent anti-inflammatory activity. The preliminary in vivo studies can also demonstrate that WK2 decreased loads of Salmonella in the liver and spleen, mitigated Salmonella-caused inflammation and maintained the integrity of intestinal mucosal surfaces. Ultimately, the results highlight that WK2 is a promising therapeutic agent to prevent multidrug-resistant S. Typhimurium infections in humans and animals.

Anti-toxicogenic fungi and toxin-reducing effects of bacillomycin D in combination with fungicides.

Mycotoxins are toxic secondary metabolites produced by fungus including Aspergillus and Fusarium. They can contaminate food and cause major health issues. Bacillomycin D (BD) is a natural antimicrobial lipopeptide generated by Bacillus that has excellent antifungal capabilities, but its high price prevents it from being widely used. Chemically produced and essential oil-based fungicides are also currently the most frequent types. In the study, the effects of combining BD with two types of fungicides on the growth of toxicogenic fungi as well as the generation of deoxynivalenol (DON) and fumonisin B1 (FB1) were examined. It was discovered that BD was more effective in suppressing molds than the other two types of fungicides, and it could be combined with synthetic or essential oil-based fungicides to provide a synergistic or additive effect. BD 31.25 µg/mL + Thymol (Thy) 7.81 µg/mL and BD 11.45 µg/mL + Cinnamon oil (Cin) 3.90 µg/mL inhibited F. graminearum, respectively. The combination of BD+Thy and BD+Cin at this concentration considerably reduced 60%-80% spore germination, when DON dropped below 300 ng/L. Furthermore, both combinations suppressed F. moniliforme growth and FB1 synthesis in a dose-dependent manner at lower concentrations. At an action dose of 2 MIC, FB1 production might be reduced to less than 100 ng/L. Our findings indicated that BD might interact synergistically with various fungicides, suggesting that it could be useful in the field of antifungal and toxicity reduction in food.