Novel Tetracycline-loaded Nano-antibacterial Agents To Combat Biofilms Of Food-borne Pathogens

Foodborne pathogens has always been one of the important sources of food safety problems.In food processing,the food-borne pathogens left on the contact surface of food or factory equipments will form biofilm,which cause serious biological pollution and equipment corrosion and further cause great food safety hazard.In addition,since the formation of biofilms increases the resistance of bacteria,the curing of related diseases requires higher doses of traditional antibacterial agents and the infections are easy to relapse.The trend of the failure of antibiotics and other traditional antibacterial agents is increasingly obvious.This paper designed novel nano-antibacterial agents based on the combination of two-dimensional（2D）or three-dimensional（3D）nanomaterials and tetracycline（Tet）,a broad-spectrum antibiotic,to restore the drug efficacy of low-dose antibiotics to overcome biofilm-related bacterial resistance.In addition,specific modification is used to extende applications of the nano antibacterial agents.The contents and results of our research are as follows:1.Synthesis and study on anti-biofilm activity of chitosan functionalized molybdenum disulfide nanosheets antibacterial agents loaded with tetracycline.A new 2D antibacterial nanocomposite namely chitosan-functionalized MoS₂nanosheet loaded with tetracycline（CM-Tet）was designed by using ionic liquid assisted grinding method.Scanning electron microscopy（SEM）,UV-vis absorption spectroscopy（UV-vis）and Fourier transform infrared spectroscopy（FTIR）demonstrated the successful synthesis of CM-Tet.The better antibacterial effect of CM-Tet than MoS₂ nanosheets and antibiotics was confirmed by the Oxford Cup method and growth curves.Conventional crystal violet staining and MTT assay confirmed that CM-Tet could effectively inhibit the formation and destroy biofilms of Gram-positive Staphylococcus aureus and Gram-negative Salmonella.In addition,the minimum inhibitory concentration（MIC）of CM-Tet was several tens of times lower than that of Tet antibiotic alone,indicating that CM-Tet can overcome biofilm-associated bacterial resistance by restoring the efficacy of traditional inefficient antibiotics.Therefore,CM-Tet could destroy biofilms and overcome bacterial resistance to treat related medical diseases and has the potential to become a substitute for traditional antibiotics.2.Synthesis and study on the anti-biofilm effect of Tet@ZIF-8 nano-antibacterial agents.A novel 3D antibacterial nanocomposite namely Tet-loaded zeolitic imidazolate frameworks（ZIF-8）（abbreviated to Tet@ZIF-8）was synthesized by self-assembly of one-pot method based on the combination of metal-organic framework（MOFs）and antibiotics.It was confirmed that Tet@ZIF-8 was successfully prepared and Tet was encapsulated inside the ZIF-8 structure by in situ according to the characterization results and the PVP experiment.The fractional inhibitory concentration（FIC）index calculated according to the checkerboard method indicates that the ZIF-8 and Tet components of Tet@ZIF-8 play synergistic roles in the antibacterial behavior.The antibacterial effect of Tet@ZIF-8 was better than both Tet and ZIF-8,and significantly reduce the dose of antibiotics.Crystal violet staining,MTT assay and SEM confirmed that Tet@ZIF-8 could effectively inhibit the formation and destroy biofilms of gram-positive/negative bacteria.Therefore,Tet@ZIF-8 provides a new approach to restore the efficacy of traditional antibiotics,that is,destroy the biofilm structure by metal ions through similar drug delivery systems and release of antibiotics in specific infection areas to overcome biofilm-associated bacterial resistance.3.Synthesis and study on effects of eliminating intracellular bacteria of Tet@ZIF-8@HA nano-antibacterial agents.A novel 3D nanoparticle was designed for stubborn intracellular bacteria through hyaluronic acid（HA）specifically modified on Tet@ZIF-8（Tet@ZIF-8@HA,TZH）to extend the application of the antibacterial agent.The characterization results demonstrated the successful preparation of TZH and its stability at room temperature.The release curves confirmed that antibiotics could targeted delivery and controlled release at intracellular bacterial infection area and improve the therapeutic effects due to pH-responsive of ZIF-8structural basis of the system.Both in vitro and in vivo results showed that ZIF-8 and Tet in TZH have synergistic and effective antibacterial effects.Fluorescence localization images confirmed that TZH can targetedly eliminate intracellular bacteria through the"barrier"of the cell membrane through the HA-mediated pathway.TZH significantly reduces the dose of antibiotics and can eliminate more than 98%of intracellular bacteria within the safe dose range.Therefore,it provided a new platform to restore the efficacy of traditional failed antibiotics and overcome bacterial resistance.