In Vitro Analyses Of Ethanol Against Trichosporon Asahii Biofilm

Background:Biofilm is resistant to the most common clinical antifungals, which plays an important role on the Trichosporon asahii infection. Some studies showed that the MICs of fungi cells in biofilms to antifungal drugs were1,600times higher than that of planktonic cells. Biofilm related infections are often difficult to be treated, eradicated and associate with high mortality. More and more physicians take attention to the biofilm related infection.It is necessary to find a new method to treat with biofilm infection.Ethanol (ETOH) is used for a variety of medical purposes. Some research show that ETOH can effectivly inhibite biofilm formation of Candida albicans and Staphylococcus aureus polymicrobial. The advantage of EtOH is low cost, universal availability and no severe side effects to EtOH treatment. But the effect of ETOH against Trichosporon asahii biofilm is not clear yet. In our research, we chose all the Trichosporon asahii strains preserved in our lab to test the EtOH effects.Objective:1. To build the in vitro model of T. ashii biofilm.2. To observe the MICs and SMICs of T. ashii to ETOH.3. To observe the effect of different concentration of ETOH on the different phages of T. ashii biofilm.Method:1. To build the in vitro model of T. ashii biofilm.Using the96-well plate method, we have built the in vitro model of T. ashii biofilm. 2. To test the MICs and SMICs of T. ashii to ETOH.According to the CLSI M27-A3method, the MICs of planktonic cells to ETOH were tested. According to the XTT-based96-well plate method, the SMICs of T. ashii to ETOH were detected.3. To observe the effect of different concentration of ETOH on the different phages of T. ashii biofilm.T. ashii cells were suspended in RPMI-1640to a final standardized cell suspensions at a density of106cells/ml counting by DENSIMAT, then pipping the suspensions to different column wells of96-well microtiter plates, and adding different concention of ETOH to different column at different stage of biofilm. At last we observed the influence.Result:1. To build the in vitro model of T. ashii biofilm.T. ashii CBS2479, T. ashii CBS8904, BZP07003, BZP07004, BZP09001, BZP09002, come from the Institute of Beijing military region general hospital skin damage repair and06108,06198,6674and6956come from Peking University first hospital fungal research center all existing T. ashii biofilms have been building a successful.2. To observe the MICs and SMICs of T. ashii to ETOH.According to XTT method we found that the MIC various on different strains. Our research shows that the range of MIC for T. ashii plantic cells was4to12.5g/ml, SMIC50was8g/ml, SMIC80was12.5g/ml; Our research shows that the range of MIC for T. ashii biofilm was25-50g/ml, SMIC50was25%, SMIC80was50%.3. To observe the effect of different concentration of ETOH in different time on the different phages of T. ashii biofilm.1) The phase of biofilm adhesion:2%ETOH forl h can obviously inhibit the adhesion rate.2) The germ tube induction and yeast-hypha transition:4%ETOH for8h can significantly inhibit the germ tube induction and yeast-hypha transitionthe of T. ashii.3) The early phase of biofilm formation:25%ETOH for8h can obviously inhibit the formation of T.ashii biofilm.4) The mature phase of biofilm:50%ETOH can eradicate the T. ashii mature biofilm.Conclusion:1. According to the96-well plates method we successfully build the in vitro model of biofilm.2. Different concentrations of ethanol can effectively inhibit the T. ashii biofilm at different times.3. T. ashii biofilm sensitivity to ethanol is higher than other antifungal drugs.4. Our study may provide a new method to treat with biofilm related infection, and throw the light on T. ashii biofilm related infection.