Exposure To Diode Laser Affects The Bacterial Adherence And Biofilm Formation In A E.faecalis Biofilm Model

Objective:A large number of studies have shown that the diode laser can effectively kill Enterococcus faecalis(E.faecalis)present in the root canal.However,as far as we know,there is little research on the deep-level killing mechanism of diode laser on E.faecalis.In this experiment,bactericidal activity of diode laser was evaluated by using plate count method and scanning electron microscopy(SEM).The distribution of extracellular polysaccharide was assessed by Confocal laser scanning microscopy(CLSM).Anthrone method was used to quantify the amount of water-soluble exopolysaccharide(WSE)and water-insoluble exopolysaccharides(WIE)in E.faecalis.Real-time quantitative polymerase chain reaction(RT-q PCR)was employed to assess transcription of genes(gel E,ace and esp)related to formation and adherence of E.faecalis biofilm.The study aiming to examine the diode Whether the laser will affect the formation and adhesion of biofilm.Methods:1.Plate colony count:An E.faecalis biofilm model was established and divided into 3 groups.After 48hours of continuous cultivation,the biofilm was treated twice a day,including:(1)using a diode laser(810nm)with an energy density of 28.7J/cm~2;(2)1%sodium hypochlorite(Na OCl)as a positive control;(3)0.9%physiological Saline(Na Cl)was used as a negative control.After the treatment of each group was completed,it was diluted step by step,and the plate colony count was performed to determine the number of viable bacteria in each group.2.Scanning electron microscopy(SEM)The samples of each group were dehydrated step by step,sprayed with gold in a vacuum ion sputtering instrument,and the biofilm samples were observed under a scanning electron microscope.3.Determination of Hydrophobicity The obtained group of E.faecalis biofilm samples were tested for cell surface hydrophobicity using the microbial adhesion hydrocarbon method(MATH).4.Exopolysaccharide measurement results4.1 Anthrone measurement results:Compared with the Na Cl group,the amount of extracellular polysaccharide in E.faecalis biofilm decreased in the laser group(p<0.05);the amount of extracellular polysaccharide in E.faecalis biofilm in the laser groupthe difference was not statistically significant(p>0.05).4.2 Laser confocal image processing results:Large area of green fluorescence can be seen in the Na Cl group image,while the red fluorescence is less,indicating that there are more viable bacteria in this area,and extracellular polysaccharide synthesis is active.Large-scale red and green fluorescence can be seen in the images of the laser irradiation group and the Na OCl group,indicating that the number of dead bacteria in the sample increased,while the amount of extracellular polysaccharides decreased relatively.5.q PCR test resultsCompared with the Na Cl group,the relative expression of gel E,ace and esp genes in the laser group decreased,and the difference was statistically significant(p<0.05).Compared with the Na OCl group,the relative expression of gel E,ace and esp genes in the laser group decreased,and the difference was statistically significant(p<0.05).Conclusion:The diode laser(810 nm)can reduce the bacterial viability of E.faecalis biofilm,which has better laser sterilization effect than Na OCl.The diode laser(810 nm)laser can reduce the surface hydrophobicity of E.faecalis and the synthesis of E.faecalis biofilm WIE and WSE,thereby inhibiting the adherence of E.faecalis biofilm;The diode laser(810 nm)can reduce the formation of E.faecalis biofilm and inhibit its adherence by inhibiting the expression of gel E,ace and esp related genes.