铁基纳米酶对鼠伤寒沙门菌生物被膜的影响
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摘要
运用结晶紫染色定量法、生物被膜形态观察、生物被膜干重称量法、活菌定量计数法和细菌内活性氧检测法,评估氧化铁纳米酶和硫化铁纳米酶对鼠伤寒沙门菌生物被膜的影响及其机制.结果显示:鼠伤寒沙门菌S025株与这两类铁基纳米酶共孵育48h后,其生物被膜结晶紫染色吸光度值(A)、生物被膜厚度、生物被膜干重和活菌数量与未处理组相比均显著下降,活性氧水平显著上升,其中硫化铁纳米酶效果优于四氧化三铁纳米酶;在生物被膜形成后,加入铁基纳米酶处理0.5h、2h和12h,生物被膜结晶紫染色A值、生物被膜厚度、生物被膜干重和活菌数量与未处理组相比均显著下降,活性氧水平显著上升,硫化铁纳米酶效果同样优于四氧化三铁纳米酶.以上结果表明,铁基纳米酶通过调控鼠伤寒沙门菌胞内活性氧水平,不仅可以预防该菌的生物被膜形成,而且可以破坏已形成的生物被膜,本研究将有助于预防和治疗鼠伤寒沙门菌生物被膜引起的相关疾病.
To evaluate the effect of iron-based nanozymes on the biofilm of S. typhimurium, the biofilms were detected by crystal violet staining method, biofilm integrity, biofilm biomass, bacterial activity, and the intracellular ROS levels of S.typhimurium S025. Our results showed that, after treatment with iron oxide(Fe_3O_4)nanozyme and iron sulfide(nFeS) nanozyme during biofilm formation for 48 h, the A_(550) value, biofilm height,biofilm biomass, and bacteria viability were significantly decreased compared to the untreated group, but the intracellular ROS levels remarkably increased. Moreover, to determine whether there is a similar inhibitory effect on pre-formed biofilms, Fe_3O_4 nanozyme and nFeS nanozyme were incubated from the apical side of the biofilms for different time. As expected, both nanozymes appreciably destroyed the biofilms that had formed. Importantly,nFeS nanozyme showed better efficacy than Fe_3O_4 nanozyme in above biofilm treatment. Taken together, these findings clearly demonstrated that iron-based nanozymes, as a suitable agent, regulated intracellular ROS levels to prevent the biofilm formation of S. typhimurium and destroyed the matured biofilm, which is helpful to preventing and treating relevant diseases caused by biofilm of S.typhimurium.
To evaluate the effect of iron-based nanozymes on the biofilm of S. typhimurium, the biofilms were detected by crystal violet staining method, biofilm integrity, biofilm biomass, bacterial activity, and the intracellular ROS levels of S.typhimurium S025. Our results showed that, after treatment with iron oxide(Fe_3O_4)nanozyme and iron sulfide(nFeS) nanozyme during biofilm formation for 48 h, the A_(550) value, biofilm height,biofilm biomass, and bacteria viability were significantly decreased compared to the untreated group, but the intracellular ROS levels remarkably increased. Moreover, to determine whether there is a similar inhibitory effect on pre-formed biofilms, Fe_3O_4 nanozyme and nFeS nanozyme were incubated from the apical side of the biofilms for different time. As expected, both nanozymes appreciably destroyed the biofilms that had formed. Importantly,nFeS nanozyme showed better efficacy than Fe_3O_4 nanozyme in above biofilm treatment. Taken together, these findings clearly demonstrated that iron-based nanozymes, as a suitable agent, regulated intracellular ROS levels to prevent the biofilm formation of S. typhimurium and destroyed the matured biofilm, which is helpful to preventing and treating relevant diseases caused by biofilm of S.typhimurium.
引文
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[2]Voetsch A C,Van Gilder T J,Angulo F J,et al.Food Net estimate of the burden of illness caused by nontyphoidal Salmonella infections in the United States.Clin Infect Dis,2004,38(Suppl 3):S127-134
[3]Romling U,Bokranz W,Rabsch W,et al.Occurrence and regulation of the multicellular morphotype in Salmonella serovars important in human disease.Int J Med Microbiol,2003,293(4):273-285
[4]Marin C,Hernandiz A,Lainez M.Biofilm development capacity of Salmonella strains isolated in poultry risk factors and their resistance against disinfectants.Poult Sci,2009,88(2):424-431
[5]Scher K,Romling U,Yaron S.Effect of heat,acidification,and chlorination on Salmonella enterica serovar typhimurium cells in a biofilm formed at the air-liquid interface.Appl Environ Microbiol,2005,71(3):1163-1168
[6]Joseph B,Otta S K,Karunasagar I,et al.Biofilm formation by Salmonella spp.on food contact surfaces and their sensitivity to sanitizers.Int J Food Microbiol,2001,64(3):367-372
[7]Flemming H C,Wingender J.The biofilm matrix.Nat Rev Microbiol,2010,8(9):623-633
[8]Shi W Y,Zhou X D.Preface for the microbial biofilm issue.Int JOral Sci,2011,3(2):47-48
[9]Duran N,Duran M,De Jesus M B,et al.Silver nanoparticles:a new view on mechanistic aspects on antimicrobial activity.Nanomedicine,2016,12(3):789-799
[10]Pelgrift R Y,Friedman A J.Nanotechnology as a therapeutic tool to combat microbial resistance.Adv Drug Deliv Rev,2013,65(13-14):1803-1815
[11]Gao L,Zhuang J,Nie L,et al.Intrinsic peroxidase-like activity of ferromagnetic nanoparticles.Nat Nanotechnol,2007,2(9):577-583
[12]Wei H,Wang E.Nanomaterials with enzyme-like characteristics(nanozymes):next-generation artificial enzymes.Chem Soc Rev,2013,42(14):6060-6093
[13]Wu J,Wang X,Wang Q,et al.Nanomaterials with enzyme-like characteristics(nanozymes):next-generation artificial enzymes(Ⅱ).Chem Soc Rev,2019,48(4):1004-1076
[14]Toumey C.Quick lessons on environmental nanotech.Nat Nanotechnol,2015,10(7):566-567
[15]Gao L,Yan X.Discovery and current application of nanozyme.Prog Biochem Biophys,2013,40(10):892-902
[16]Xu Z,Qiu Z,Liu Q,et al.Converting organosulfur compounds to inorganic polysulfides against resistant bacterial infections.Nat Commun,2018,9(1):3713
[17]Pratt L A,Kolter R.Genetic analysis of Escherichia coli biofilm formation:roles of flagella,motility,chemotaxis and typeⅠpili.Mol Microbiol,1998,30(2):285-293
[18]El Hag M,Feng Z,Su Y,et al.Contribution of the csg A and bcs Agenes to Salmonella enterica serovar Pullorum biofilm formation and virulence.Avian Pathol,2017,46(5):541-547
[19]Lawrence J R,Wolfaardt G M,Korber D R.Determination of diffusion coefficients in biofilms by confocal laser microscopy.Appl Environ Microbiol,1994,60(4):1166-1173
[20]Banin E,Brady K M,Greenberg E P.Chelator-induced dispersal and killing of Pseudomonas aeruginosa cells in a biofilm.Appl Environ Microbiol,2006,72(3):2064-2069
[21]Hengzhuang W,Wu H,Ciofu O,et al.Pharmacokinetics/pharmacodynamics of colistin and imipenem on mucoid and nonmucoid Pseudomonas aeruginosa biofilms.Antimicrob Agents Chemother,2011,55(9):4469-4474
[22]Wang H Y,Hua X W,Wu F G,et al.Synthesis of ultrastable copper sulfide nanoclusters via trapping the reaction intermediate:potential anticancer and antibacterial applications.ACS Appl Mater Interfaces,2015,7(13):7082-7092
[23]Yang X,Li J,Liang T,et al.Antibacterial activity of twodimensional Mo S2 sheets.Nanoscale,2014,6(17):10126-10133
[24]Fang F C.Antimicrobial actions of reactive oxygen species.MBio,2011,2(5),pii:e00141-11
[25]Dunnill C,Patton T,Brennan J,et al.Reactive oxygen species(ROS)and wound healing:the functional role of ROS and emerging ROS-modulating technologies for augmentation of the healing process.Int Wound J,2017,14(1):89-96
[26]Tang Y,Qiu Z,Xu Z,et al.Antibacterial mechanism and applications of nanozymes.Prog Biochem Biophys,2018,45(2):118-128