Methylene Blue Mediated Photodynamic Therapy Inactivation Of Pseudomonas Aeruginosa

Part1 Study on different concentrations of methylene blue mediated photodynamic therapy against pseudomonas aeruginosaObjective To evaluate the efficacy of different concentrations of methylene blue(MB)mediated photodynamic therapy(PDT)on pseudomonas aeruginosa in vitro.Methods Pseudomonas aeruginosa was used to prepare bacteria suspension at a concentration of 10~8 colony-forming units/mL.The study was divided into the PDT group,the photosensitizer group,the light group and the blank control group.(1)The PDT group:the pseudomonas aeruginosa suspension was mixed with different concentrations of MB and incubated in dark at room temperation for 30 min.Then the mixed suspension was exposed to the illuminant at the same energy and wavelength for 13 min according to the experiment design;(2)The photosensitizer group:the pseudomonas aeruginosa suspension was mixed with different concentrations of MB and incubated in dark at room temperation for 30 min without illumination;(3)The light group:the pseudomonas aeruginosa suspension without photosensitizer was exposed to the same energy of illumination as the PDT group;(4)The blank control group:the pseudomonas aeruginosa suspension was without photosensitizer and illumination.The survival rate of pseudomonas aeruginosa was calculated by counting colony-forming units after above treatments.Results The survival rate was 73.6559%,9.8896%,1.2565%,0.1148%,0.0645%,0.0112%,0.0016%,respectively,at different concentration of MB(5?mol/L,7.5?mol/L,10?mol/L,12.5?mol/L,15?mol/L,17.5?mol/L,20?mol/L).At the concentration of MB from 5?mol/L to 20?mol/L,the PDT effect showed concentration-dependent killing(P<0.05).at the concentration of2.5?mol/L,The survivaI rate of pseudomonas aeruginosa was 89.5262%(P>0.05).Conclusion Under the same light energy condition,the PDT effect of MB showed concentration-dependent killing in a certain range of concentration.Part2 Study on different energies and MB-mediated PDT of pseudomonas aeruginosaObjective To evaluate the efficacy of different energies and MB-mediated PDT on pseudomonas aeruginosa in vitro.Methods Pseudomonas aeruginosa was used to prepare bacteria suspension at a concentration of 10~8 colony-forming units/mL.The study was divided into the PDT group,the blank control group.(1)The PDT group:pseudomonas aeruginosa cell suspension was mixed with MB(the final concentration of MB was 20?mol/L),incubated in dark for 30 min,and then irradiated with a red light source with a wavelength of 660 nm for 0 min,2.6 min,5.2 min,10.4 min and 20.8 min,respectively(corresponding energies were 0,2,4,8 and 16 J/cm~2,respectively).(2)The blank control group:the pseudomonas aeruginosa suspension was without MB and illumination.The survival rate of pseudomonas aeruginosa was calculated by counting colony-forming units after above treatments?Results Under the same concentration of MB(20?mol/L)parameter,the survival rate of pseudomonas aeruginosa was 79.25%,38.94%,3.562%,0.03156%and 0.0001523%,respectively,when the red light source with different energies(0,2,4,8,16 J/cm~2)were irradiation.At the energy of 16J/cm~2,MB-mediated PDT reduced the colony-forming units of pseudomonas aeruginosa by about 6 log at the concentration of 20?mol/L.Conclusion Under the same concentration of MB.the PDT effect of MB showed energy-dependent killing in a certain range of energy.Part3 Study on potassium iodide combined with MB-mediated PDT of pseudomonas aeruginosaObjective To evaluate whether potassium iodide(KI)potentials the efficacy of MB-mediated PDT on pseudomonas aeruginosa in vitro.Methods Pseudomonas aeruginosa was used to prepare bacteria suspension at a concentration of 10~8 colony-forming units/mL.The study was divided into the MB-PDT group,KI+MB-PDT group and the blank control group.(1)The MB-PDT group:the pseudomonas aeruginosa suspension was mixed with MB(final concentration was 7.5?mol/L)and incubated in dark at room temperation for 30 min.Then the mixed suspension was exposed to the illuminant at the same energy and wavelength according to the study design;(2)The KI+MB-PDT group:the pseudomonas aeruginosa suspension was mixed with different concentrations of KI(1 mmol/L,2 mmol/L,5 mmol/L,10 mmol/L)and MB(7.5?mol/L)and incubated in dark at room temperation for 30 min without illumination;(3)The blank control group:the pseudomonas aeruginosa suspension was without KI,MB and illumination.The survival rate of pseudomonas aeruginosa was calculated by counting colony-forming units after above treatments.Results At 10 J/cm~2 of light energy,the survival rate was 9.589%,6.452%,0.2258%,0.0006452%respectively,at different concentrations of KI(1mmol/L,2 mmol/L,5 mmol/L,10 mmol/L)combined with MB(7.5?mol/L).Otherwise MB-mediated PDT reduced the colony-forming units of pseudomonas aeruginosa by about 5 log at the concentration of 7.5?mol/L combined with KI at the concentration of 10 mmol/L(P<0.05).Conclusion KI enhanced the killing effect of MB-mediated PDT against pseudomonas aeruginosa.