| Objective To explore the antimicrobial photodynamic therapy(aPDT)mediated by Methylene blue(MB)combined with potassium iodide(KI)for different sources of Meheicillin-resistant Staphylococcus aureus(MRSA)and Carbapenem-resistant Eschenrichia coli(CR-E.coli)under different parameters.Methods 14 strains of MRSA and 11 strains of CR-E.coli identified and isolated from the laboratory microbiology of the First Affiliated Hospital of Guangxi Medical University were used as the research objects,MRSA and CR-E.coli bacterial suspensions were prepared with 108 Colony forming units(CFU)/mL.(1)Antibacterial effects of different concentrations of MB mediated photodynamic therapy for different sources of MRSA and CR-E.coli.The experiment was divided into four groups,a.Photodynamic group 1:108 CFU/mL of MRSA suspension was mixed with different concentrations of MB(the final concentration of MB was 0.1 μmol/L,0.5 μmol/L,1 μmol/L,3μmol/L,5 pmol/L,10 μmol/L).Photodynamic group 2:The bacterial suspension of 108 CFU/mL CR-E.coli was mixed with different concentrations of MB(the final concentration was 1 μmol/L,5 μmol/L,10 μmol/L,15 μmol/L,20 μmol/L,30 μmol/L).Both groups were incubated in the dark for 30min,and then illuminated with 660 nm red light at 10 J/cm2.Simple MB group:108 CFU/mL of MRSA or CR-E.coli suspension was mixed with MB(the concentration of MB was same as the photodynamic group)and then incubated for 30min without illumination.Pure light group:108 CFU/mL of MRSA or CR-E.coli bacterial suspensions were illuminated with 660 nm red light at 10 J/cm2.Blank control group:Only 108 CFU/mL of MRSA or CR-E.coli suspension,no MB,no illumination.Each group was continuously diluted in gradient and inoculated in the medium.The survival fraction of MRSA or CR-E.coli was evaluated by counting CFU.(2)Study on antibacterial effect of different concentration KI combined with MB mediated photodynamic therapy for different sources of MRSA and CR-E.coli.The experiment was divided into four groups.a.Photodynamic group 1:108 CFU/mL of MRSA suspension was mixed with 0.5 μmol/L MB,different concentrations of KI were added(the final concentration was 10 mmol/L,30 mmol/L,50 mmol/L,100 mmol/L)after 30 minutes of dark incubation.Photodynamic group 2:the bacterial suspension of 108 CFU/mL CR-E.coli was mixed with the 1 μmol/L MB,After 30 minutes of dark incubation,different concentrations of KI were added(the final concentration of KI was 1 mmol/L,5 mmol/L,10 mmol/L,15 mmol/L,20 mmol/L,50 mmol/L).Both groups were illuminated with 660 nm red light at 10 J/cm2.b.Dark control group 1:108 CFU/mL of MRSA or CR-E.coli suspension was mixed with MB and incubated for 30min before KI was added(the concentrations of MB and KI were same as the photodynamic group),no light.Dark control group 2:108 CFU/mL of MRSA or CR-E.coli suspension was mixed with KI(the concentration of KI were same as the photodynamic group),no light.Light control group:108 CFU/mL of MRSA or CR-E.coli bacterial suspensions were illuminated with 660nm red light at 10 J/cm2.Blank control group:Only 108 CFU/mL MRS A or CR-E.coli suspension,no MB,no KI,no illumination.Each group was continuously diluted in gradient and inoculated in the medium.The survival fraction of MRSA or CR-E.coli was evaluated by counting CFU.(3)Study on the antibacterial effect of KI combined with MB mediated photodynamic therapy for different sources of MRSA and CR-E.coli by different light energy densities.The experiment was divided into four groups.a.Photodynamic group 1:108 CFU/mL of MRSA bacterial suspension was mixed with 0.5 μmol/L of MB,and 50 mmol/L of KI was added after 30 min of dark incubation.Photodynamic group 2:108 CFU/mL of bacterial suspension of CR-E.coli was mixed with the final concentration of 1 μmol/L MB,and 20 mmol/L KI was added after 30 min of incubation in the dark.Both groups were illuminated at different light energy densities(2 J/cm2,4 J/cm2,6 J/cm2,8 J/cm2,10 J/cm2).b.Dark control group:108 CFU/mL of MRSA or CR-E.coli suspension was mixed with MB and incubated for 30min,then KI was added(The concentrations of MB and KI were same as those in the photodynamic group),no illumination.c.Light control group:108 CFU/mL of MRSA or CR-E.coli bacterial suspensions were illuminated under different light energy densities(The light energy density was same as the photodynamic group),d.Blank control group:Only 108 CFU/mL of MRSA or CR-E.coli suspension,no MB,no KI,no illumination.Each group was continuously diluted in gradient and inoculated in the medium.The survival fraction of MRSA or CR-E.coli was evaluated by counting CFU.Results(1)Antibacterial effects of different concentrations of MB mediated photodynamic therapy for different sources of MRS A and CR-E.coli.Under photodynamic therapy mediated by different concentration of MB(the final concentration of MB were 0.1 μmol/L,0.5 μmol/L,1 μmol/L,3μmol/L,5 μmol/L,10 μmol/L),The survival rates of MRSA were 59.75%,9.22%,0.24%,0.0040%,0.000051%,0.000020%,and when MB were 1μmol/L,5 μmol/L,10μmol/L,15 μmol/L,20 μmol/L,30 μmol/L,the survival rates of CR-E.coli were 9.99%,0.16%,0.0031%,0.00023%,0.000016%,0.000011%.Compared with blank control group,the differences were statistically significant(P<0.05).There was no significant difference among the MB group,the light group and the blank control group(P>0.05).(2)Antibacterial effect of different concentration KI combined with MB mediated photodynamic therapy for different sources of MRSA and CR-E.coli.With 0.5 μmol/L MB combined with different concentrations of KI(the final concentration of KI were 10 mmol/L,30 mmol/L,50 mmol/L,100 mmol/L)mediated photodynamic therapy,The survival rates of MRSA were 80.65%,0.93%,0.000048%,0.0000087%.Compared with blank control group,the differences were statistically significant(P<0.05).1μmol/L MB combined with different concentrations of KI(the final concentration of KI were 1 mmol/L,5 mmol/L,10 mmol/L,15 mmol/L,20 mmol/L,50 mmol/L)mediated photodynamic effect,The survival rates of CR-E.coli were 9.18%,4.42%,0.55%,0.0087%,0.000040%,0.0000080%.compared with blank control group the differences were statistically significant(P<0.05).There was no statistical significance in MB combined with KI mediated photodynamic therapy on MRSA and CR-E.coli that from different sources(P>0.05).There was no significant difference between dark control group,light control group and blank control group(P>0.05).(3)Antibacterial effect of KI combined with MB mediated photodynamic therapy for different sources of MRS A and CR-E.coli with different light energy densities.Under different light energy densities(2 J/cm2,4 J/cm2,6 J/cm2,8 J/cm2,10 J/cm2),0.5 μmol/L MB combined with 50 mmol/L KI mediated photodynamic therapy,MRSA survival rates were 0.027-0.89%,0.000013%,0.000013%,0.000013%,0.000013%.1 μmol/L MB combined with 20 mmol/L KI mediated photodynamic therapy,CR-E.coli survival rates were 25.0-89.17%,3.4212.29%,0.20-1.64%,0.00054-0.0095%,0.000017%.Compared with blank control group,the differences were statistically significant(P<0.05).There was no significant difference between dark control group,light control group and blank control group(P>0.05).Conclusion(1)The origin and drug resistance situation of MRSA and CR-E.coli strains did not affect the bactericidal effect of MB-mediated APDT on MRSA and CR-E.coli.Under the radiant exposure of 10 J/cm2,the photodynamic effect of MB anti-MRSA and CR-E.coli were enhanced by increasing the concentration of MB.(2)The photodynamic antibacterial effection of MB combined with KI did not change with the origin and drug resistance situation of MRSA and CR-E.coli.Photodynamic effects of MB on MRSA and CR-E.coli was enhanced after adding KI.(3)Under the fixed concentration of MB and KI,The antimicrobial effect of MB combined with KI mediated photodynamic therapy on MRSA and CR-E.coli can be improved by adjusting different parameters. 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