Activity Of Dially Trisulfide Against Enterococcus Faecalis Biofilms In Vitro

Objective:This study is to observe dially trisulfide (DATS) in eradicating or inhibiting the biofilm of Enterococcus faecalis in vitro.Method:1.Times’dilution method was used to examine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of DATS and hydrochloric acid doxycycline to Enterococcus faecalis (ATCC29212).2. Bacterial growth curve drawing has been determined by MTT method, which was used to determine the activity of Enterococcus faecalis at different time intervals. Bacteria was cultivated in96-well cell culture plates, forming the bacterial biofilm of different growth periods. Bacteriostasis rate of the bacterial biofilm in different periods of DATS or hydrochloric acid doxycycline in different concentration is determined by MTT method.3. Morphological characteristics in the process of bacterial biofilm formation in the cultivated bacteria of the dentin slices were observed by the Concofocal Scanning Laser Machine (CLSM). CLSM was also used to observe the antibacterial effect of different concentrations of DATS or doxycycline hydrochloride to the different growth cycle of mature bacteria biofilm.Results:1.The minimum bacteriostatic concentration (MIC) of DATS is1280μg/ml, and the minimum bactericidal concentration (MBC) is2560μg/ml; while in the control group doxycycline, MIC is80μg/ml and MBC is160μg/ml.2.1280μg/ml of DATS has the bacteriostatic rate of about71%to logarithmic phase and hunger issue of the biofilm of Enterococcus faecalis, which is significantly higher than the biofilm in stability phase whose inhibition rate is about58%, the difference was statistically significance (P <0.05).The2560μg/ml of DATS and5120μg/ml of DATS anti-biofilm Enterococcus faecalis effect is significantly better than the DATS of1280μg/ml, the difference was statistically significant (P<0.05). DATS of the5120μg/ml has an antibacterial rate of97%to the Enterococcus faecalis in hunger issue, which is significantly higher than the antibacterial rates of the other concentrations of DATS in any other phases of biofilm, or this concentration to the other two phase of the biofilm of Enterococcus faecalis, the results were statistically significant(P<0.05).The difference of inhibition rate is not statistically significant (P>0.05) between80μg/ml of hydrochloride doxycycline and5120μg/ml of DATS to the stages logarithmic or hunger biofilm,but has statistically significant (P<0.05) to the stabilization of biofilm.3.Enterococcus faecalis biofilm has a three-dimensional structure, biofilm seen by the red dye District (dead bacteria) with green stained area (live bacteria). Logarithmic phase, the stabilization period and hunger period biofilm experienced three stages that are adhesion, development and mature.From the adhesion period to mature, the thickness of biofilm increases and becomes more solid and dense.The bacteria biofilm, in the starvation period has a lower density, looser structure and a surface roughness than that in logarithmic phase or stable period.Both DATS and Doxycycline make the red dye area increase when seen in CLSM, which means both can eradicate or inhibit the biofilm of Enterococcus faecalis. Furthermore when the concentration is increased, the red dye area increases, as well as the bacterial biofilms become more decentralized and loose.Conclusion:1. DATS can kill or inhibit significantly Enterococcus faecalis.2. DATS can destroy the biofilm structure of Enterococcus faecalis, hence kill the bacteria inside the biofilm.3. DATS significantly kill the bacteria of hunger issue of the biofilm of Enterococcus faecalis.