研究目的
Investigating the synergistic effects of indocyanine green and ethylenediamine tetraacetate-mediated photodynamic antimicrobial chemotherapy combined with antibiotics for resistant bacterial biofilms in diabetic foot infection.
研究成果
ICG and EDTA-mediated PACT combined with antibiotics significantly enhances bactericidal effects on planktonic bacteria and resistant biofilms, disrupts biofilm structure, and improves antibiotic susceptibility, offering a promising approach for treating diabetic foot infections and other superficial infectious diseases with biofilms, though clinical trials are required.
研究不足
The study is in vitro and may not fully replicate in vivo conditions of diabetic foot infections. Clinical validation is needed. Photothermal effects from laser irradiation could pose risks in patients with compromised temperature regulation.
1:Experimental Design and Method Selection:
The study used in vitro models to test the effects of PACT with ICG and EDTA on planktonic bacteria and biofilms of S. aureus and P. aeruginosa, including resistant strains (MRSA and MRPA). Methods included bacterial viability assays, biofilm growth assessment, photothermal effect measurement, PACT treatments, antimicrobial susceptibility tests, confocal microscopy, and resazurin assays.
2:Sample Selection and Data Sources:
Bacterial strains included S. aureus (ATCC 25923), MRSA (10485), P. aeruginosa (ATCC 27853), and MRPA (10911), obtained from the State Key Laboratory of Infectious Diseases and Parasites and the First Affiliated Hospital of Chongqing Medical University.
3:List of Experimental Equipment and Materials:
Equipment included a diode laser (808 nm, Mid-River Ltd.), microplate reader (Varioskan Flash, Thermo Fisher Scientific), infrared camera (Fluke Ti32, Fluke Company), confocal microscope (Leica TCS.SP8), and various reagents such as ICG (Aladdin Industrial Corporation), EDTA, antibiotics, and staining kits.
4:Experimental Procedures and Operational Workflow:
Procedures involved preparing bacterial suspensions, incubating with ICG and EDTA, laser irradiation at 808 nm with
5:5 W/cm2 power intensity and 135 J/cm2 energy dose, colony counting, disk diffusion tests, microdilution assays, biofilm staining, and metabolic activity measurements. Data Analysis Methods:
Statistical analysis was performed using GraphPad Prism 6.0 with one-way ANOVA and Tukey's test for multiple comparisons.
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microplate reader
Varioskan Flash
Thermo Fisher Scientific
Used to measure optical density (OD600) of bacterial suspensions and absorbance in assays.
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infrared camera
Ti32
Fluke Company
Used to measure temperature during laser irradiation to assess photothermal effects.
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confocal microscope
TCS.SP8
Leica
Used for confocal laser scanning microscopy to detect biofilm morphology and bacterial viability with staining.
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diode laser
excitation wavelength 808 nm
Mid-River Ltd.
Used for photodynamic antimicrobial chemotherapy (PACT) experiments to irradiate bacterial samples.
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photosensitizer
indocyanine green (ICG)
Aladdin Industrial Corporation
Used as a photosensitizer in PACT to generate reactive oxygen species for bacterial inactivation.
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LIVE/DEAD Baclight Bacterial Viability Kit
Molecular Probes
Used to stain biofilms for viability assessment with SYTO9 and propidium iodide stains.
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Alamar Blue
Yeasen
Used in resazurin assay to quantify metabolic activity of bacteria in biofilms.
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