DETERMINATION OF ADHERENCE KINETICS OF P. ACNES AND EFFICACY OF TIO2-PDMS SILVER COATING

dgarcia

Background

Propionibacterium acnes

  • Slow-growing, anaerobic-aerotolerant, gram positive rod.
  • Commonly found as part of the natural flora of the large intestine, conjunctiva, oral cavity, and in the pilosebaceous follicles of the skin.
    • Most common organism colonizing shoulder area.
  • Commonly recognized as the causative agent of acne vulgaris.
  • Recognized as an opportunistic pathogen colonizing implants on the shoulder region.
    • Most frequently isolated pathogen in prosthetic shoulder joint infections.
  • Current pre-surgical dressings like chlorheximide gluconate appear to not effective against it.

Methodology

  • Cultured in a humidified anaerobic environment in Reduced Clostridial Media (BD) for 48hrs at 37oC.
  • Implants inoculated with 1×107 cfu/ml for variable times of adherence and proliferation.
  • Dehydrated, Fixed, and Labeled.
  • Visualized via SEM and CLSM.
  • Coated implants dip-coated in 95% 10X (95% TiO2: 5% PDMS) and 100% Ag.

Experimental Conditions

4-20 8-16 12-12 16-8 20-4
Table 1.  Adherence and Proliferation times for PA.  The first number in each set corresponds to the number of hours allowed for adherence, while the second number corresponds to the number of hours allowed for proliferation. Each of these conditions was examined for each coating condition tested.
Control Experimental Control
Uncoated 95% 10X 100% Ag
Table 2. Coating conditions for PA experiments. Concentration ratios of TiO2 to PDMS doped with silver. 95% refers to TiO2 to PDMS ratio, while 0x and 10x correspond to silver neodecanoate concentrations. 100% Ag refers to coating with no TiO2/PDMS. 

Results

Spinal Implant Materials

Fig 1.  100x magnification of implant materials after machining, non-abrasive cleaning, and autoclave sterilization. (A) Ti, (B) SS, (C) CC, (D) PEEK, (E) TiA.

Fig 2. Contact Angle of Spinal Implant Materials

 

 

 

 

 

 

 

 

Fig 3. Representative SEM images at 5,000x magnification (Left) and Confocal Laser Scanning Microscopy at 120x (right) of P. acnes on native material surfaces.  (A) PEEK, (B) CC, (C) SS, (4) Ti, (5) TiA. (4-20)

Fig 4. Representative SEM images at 5,000x magnification (Left) and Confocal Laser Scanning Microscopy at 120x (right) of P. acnes on native material surfaces.  (A) PEEK, (B) CC, (C) SS, (4) Ti, (5) TiA. (8-16)

 

 

 

 

 

 

 

Fig 5. Average number of bacteria on SEM images of uncoated materials at 8 hours adherence and 16 hours proliferation.

Fig 6. Percentage area coverage on confocal microscopy of uncoated materials at 8 hours adherence and 16 hours proliferation.

 

 

 

 

 

 

 

 

Fig 7. Representative SEM images at 5,000x magnification (Left) and Confocal Laser Scanning Microscopy at 120x (right) of P. acnes on native material surfaces.  (A) PEEK, (B) CC, (C) SS, (4) Ti, (5) TiA. (16-8)

Fig 8. Average number of bacteria on SEM images of uncoated materials at 16 hours adherence and 8 hours proliferation.

Fig 9. Percentage area coverage on confocal microscopy of uncoated materials at 16 hours adherence and 8 hours proliferation.

Fig 10. Representative SEM images at 5,000x magnification (Left) and CLSM at 120x (Right) of P. acnes on 100% Ag coated surfaces.  (A) PEEK, (B) CC, (C) SS, (4) Ti, (5) TiA (8-16).

Fig 11. Average number of bacteria on SEM images of 100% Ag coated materials at 8 hours adherence and 16 hours proliferation.

Fig 12. Percentage area coverage on confocal microscopy of 100% Ag coated materials at 8 hours adherence and 16 hours proliferation.

Fig 13. Representative SEM images at 5,000x magnification (Left) and CLSM at 120x (Right)

Fig 14. Average number of bacteria on SEM images of 95 10x coated materials at 8 hours adherence and 16 hours proliferation.

Fig 15. Percentage area coverage on confocal microscopy of 95 10x coated materials at 8 hours adherence and 16 hours proliferation.

Fig 16. Representative samples of uncoated pigskin after H&E staining and histopathology at 20x magnification.

Fig 17. Representative samples of pigskin coated in chlorhexidine gluconate after H&E staining and histopathology at 20x magnification.
Fig 18. Representative samples of pigskin coated in silver doped TiO2-PDMS after H&E staining and histopathology at 20x magnification.

Conclusion

  • P. acnes is able to form a biofilm within 4hrs of adherence and 20hrs of propagation on PEEK implants.
  • Ag-Doped Coating is able to prevent biofilm formation and reduce overall bacterial adhesion.
  • Ag-Doped Coating can penetrate deeper into pilosebaceous follicles than chlorheximide gluconate.