Listeria monocytogenes is a common foodborne pathogen, and its biofilm is an important cause of food contamination and disease transmission.In this study, the cell free supernatant (CFS) of Bacillus subtilis was selected as an inhibitor to explore the role and potential of B. subtilis CFS in inhibiting L.monocytogenes 118 (No.118 for short) biofilm.Firstly, the minimum inhibitory concentration (MIC) of B. subtilis CFS was determined, and then the biofilm inhibition rate of B. subtilis CFS to No.118 was determined by using crystal violet staining method.Results showed that CFS at different sub-MICs had a significant inhibitory effect on No.118 biofilm, and the biofilm inhibition rate could still reach 47% at 1/64 MIC.Meanwhile, B. subtilis CFS also significantly inhibited the metabolic activity, self-aggregation rate, surface hydrophobicity, swaming and extramembrane polymer secretion of No.118 biofilm cells.Finally, the changes of No.118 biofilm structure and cell distribution under the action of CFS were observed by confocal laser scanning microscope.After CFS treatment, the structure of No.118 biofilm was loose, and the number of bacteria was significantly reduced.Consequently, this study found that B. subtilis CFS can inhibit the biofilm formation of No.118 biofilm cells by affecting their metabolic activity, self-aggregation ability and swaming, as well as the secretion of extramembrane polymers.This has important reference value for exploring the application of probiotic metabolites as biofilm inhibitors.
[1] LIU Y T, SUN W X, SUN T M, et al.The prevalence of Listeria monocytogenes in meat products in China:A systematic literature review and novel meta-analysis approach[J].International Journal of Food Microbiology, 2020, 312:108358.
[2] ADETUNJI C O, OLANIYAN O T, ANANI O A, et al.Role of biosurfactant in the destruction of pores and destabilization of the biological membrane of pathogenic microorganisms[M]//Green Sustainable Process for Chemical and Environmental Engineering and Science.Amsterdam:Elsevier, 2022:175-188.
[3] KASPAR F, NEUBAUER P, GIMPEL M.Bioactive secondary metabolites from Bacillus subtilis:A comprehensive review[J].Journal of Natural Products, 2019, 82(7):2038-2053.
[4] LIU Y G, DONG P C, ZHU L X, et al.Effect of four kinds of natural antimicrobial compounds on the biofilm formation ability of Listeria monocytogenes isolated from beef processing plants in China[J].LWT, 2020, 133:110020.
[5] VERMA P, ANAND S.Antimicrobial activity as a potential factor influencing the predominance of Bacillus subtilis within the constitutive microflora of a whey reverse osmosis membrane biofilm[J].Journal of Dairy Science, 2020, 103(11):9992-10000.
[6] YU X X, LI J Y, MU D S, et al.Green synthesis and characterizations of silver nanoparticles with enhanced antibacterial properties by secondary metabolites of Bacillus subtilis (SDUM301120)[J].Green Chemistry Letters and Reviews, 2021, 14(2):190-203.
[7] MELO T A, DOS SANTOS T F, DE ALMEIDA M E, et al.Inhibition of Staphylococcus aureus biofilm by Lactobacillus isolated from fine cocoa[J].BMC Microbiology, 2016, 16(1):250.
[8] 尹清干, 程俊茗, 刘腾飞, 等.环境因子对鳗弧菌生物膜形成的影响[J].微生物学通报, 2018, 45(1):138-145.
YIN Q G, CHENG J M, LIU T F, et al.Characteristics of biofilm formation of pathogenic Vibrio anguillarum[J].Microbiology China, 2018, 45(1):138-145.
[9] KIM Y J, YU H H, SONG Y J, et al.Anti-biofilm effect of the cell-free supernatant of probiotic Saccharomyces cerevisiae against Listeria monocytogenes[J].Food Control, 2021, 121:107667.
[10] WANG H H, CAI L L, LI Y H, et al.Biofilm formation by meat-borne Pseudomonas fluorescens on stainless steel and its resistance to disinfectants[J].Food Control, 2018, 91:397-403.
[11] YANG X, PEI Z J, HU R J, et al.Study on the inhibitory activity and possible mechanism of myriocin on clinically relevant drug-resistant Candida albicans and its biofilms[J].Biological & Pharmaceutical Bulletin, 2021, 44(3):305-315.
[12] KARYGIANNI L, ATTIN T, THURNHEER T.Combined DNase and proteinase treatment interferes with composition and structural integrity of multispecies oral biofilms[J].Journal of Clinical Medicine, 2020, 9(4):983.
[13] ZHANG J, GU S B, ZHANG T R, et al.Characterization and antibacterial modes of action of bacteriocins from Bacillus coagulans CGMCC 9951 against Listeria monocytogenes[J].LWT, 2022, 160:113272.
[14] MELIAN C, BENTENCOURT E, CASTELLANO P, et al.Biofilm genes expression of Listeria monocytogenes exposed to Latilactobacillus curvatus bacteriocins at 10 ℃[J].International Journal of Food Microbiology, 2022, 370:109648.
[15] CHIANG W C, PAMP S J, NILSSON M, et al.The metabolically active subpopulation in Pseudomonas aeruginosa biofilms survives exposure to membrane-targeting antimicrobials via distinct molecular mechanisms[J].FEMS Immunology and Medical Microbiology, 2012, 65(2):245-256.
[16] WANG D, LIU Y F, LI X P, et al.Unraveling the antibacterial mechanism of Lactiplantibacillus plantarum MY2 cell-free supernatants against Aeromonas hydrophila ST3 and potential application in raw tuna[J].Food Control, 2023, 145:109512.
[17] ZHOU Y H, XU C G, YANG Y B, et al.Histidine metabolism and IGPD play a key role in cefquinome inhibiting biofilm formation of Staphylococcus xylosus[J].Frontiers in Microbiology, 2018, 9:665.
[18] YANG S J, LEE J E, LIM S M, et al.Antioxidant and immune-enhancing effects of probiotic Lactobacillus plantarum 200655 isolated from kimchi[J].Food Science and Biotechnology, 2018, 28(2):491-499.
[19] TRAVIER L, GUADAGNINI S, GOUIN E, et al.ActA promotes Listeria monocytogenes aggregation, intestinal colonization and carriage[J].PLoS Pathogens, 2013, 9(1):e1003131.
[20] ALIZADEH BEHBAHANI B, NOSHAD M, FALAH F.Inhibition of Escherichia coli adhesion to human intestinal Caco-2 cells by probiotic candidate Lactobacillus plantarum strain L15[J].Microbial Pathogenesis, 2019, 136:103677.
[21] WEI Z H, SHAN C J, ZHANG L X, et al.A novel subtilin-like lantibiotics subtilin JS-4 produced by Bacillus subtilis JS-4, and its antibacterial mechanism against Listeria monocytogenes[J].LWT, 2021, 142:110993.
[22] SATPUTEA S K, BANPURKAR A G, BANAT I M, et al.Multiple roles of biosurfactants in biofilms[J].Current Pharmaceutical Design, 2016, 22(11):1429-1448.
[23] RUHAL R, KATARIA R.Biofilm patterns in gram-positive and gram-negative bacteria[J].Microbiological Research, 2021, 251:126829.
[24] SOUSA C, TEIXEIRA P, OLIVEIRA R.The role of extracellular polymers on Staphylococcus epidermidis biofilm biomass and metabolic activity[J].Journal of Basic Microbiology, 2009, 49(4):363-370.
[25] RODRIGUES L R, TEIXEIRA J A, VAN DER MEI H C, et al.Isolation and partial characterization of a biosurfactant produced by Streptococcus thermophilus A[J].Colloids and Surfaces.B, Biointerfaces, 2006, 53(1):105-112.
[26] CHANDLER J R, DUERKOP B A, HINZ A, et al.Mutational analysis of Burkholderia thailandensis quorum sensing and self-aggregation[J].Journal of Bacteriology, 2009, 191(19):5901-5909.
[27] MELIAN C, SEGLI F, GONZALEZ R, et al.Lactocin AL705 as quorum sensing inhibitor to control Listeria monocytogenes biofilm formation[J].Journal of Applied Microbiology, 2019, 127(3):911-920.
