Novel murine model for delayed wound healing using a biological wound dressing with Pseudomonas aeruginosa biofilms.

Bacterial biofilms impair healing in 60% of chronic skin wounds. Various animal models (mice, rats, rabbits, and pigs) have been developed to replicate biofilm infected wounds in vivo. We developed a sustained wound infection model by applying preformed Pseudomonas aeruginosa biofilms on a wound dressing to full-thickness murine skin wounds. We bathed a commercially available wound dressing in P. aeruginosa for 48 h, allowing a biofilm to establish on the dressing prior to application to the wound. Dressings were removed from the wounds after 3 days at which time the wound beds contained ∼108 bacterial cells per gram tissue. Significant numbers of P. aeruginosa persisted within the skin wounds for up to 21 days. Un-inoculated wounds reached closure between 9 and 12 days. In contrast, biofilm-inoculated wounds achieved closure between 18 and 21 days. Histologic analysis confirmed decreased re-epithelialization and collagen deposition, coupled with increased inflammation, in the biofilm-inoculated wounds compared to un-inoculated controls. This novel model of delayed healing and persistent infection of full-thickness murine skin wounds may provide a robust in vivo system in which to test novel treatments to prevent wound infection by bacterial biofilms.

Inhibition of Pseudomonas aeruginosa biofilm formation on wound dressings.

Chronic nonhealing skin wounds often contain bacterial biofilms that prevent normal wound healing and closure and present challenges to the use of conventional wound dressings. We investigated inhibition of Pseudomonas aeruginosa biofilm formation, a common pathogen of chronic skin wounds, on a commercially available biological wound dressing. Building on prior reports, we examined whether the amino acid tryptophan would inhibit P. aeruginosa biofilm formation on the three-dimensional surface of the biological dressing. Bacterial biomass and biofilm polysaccharides were quantified using crystal violet staining or an enzyme linked lectin, respectively. Bacterial cells and biofilm matrix adherent to the wound dressing were visualized through scanning electron microscopy. D-/L-tryptophan inhibited P. aeruginosa biofilm formation on the wound dressing in a dose dependent manner and was not directly cytotoxic to immortalized human keratinocytes although there was some reduction in cellular metabolism or enzymatic activity. More importantly, D-/L-tryptophan did not impair wound healing in a splinted skin wound murine model. Furthermore, wound closure was improved when D-/L-tryptophan treated wound dressing with P. aeruginosa biofilms were compared with untreated dressings. These findings indicate that tryptophan may prove useful for integration into wound dressings to inhibit biofilm formation and promote wound healing.

Reduction in wound bioburden using a silver-loaded dissolvable microfilm construct.

Silver is a widely used antimicrobial agent, yet, when impregnated in macroscopic dressings, it stains wounds, can lead to tissue toxicity, and can inhibit healing. Recently, polymeric nanofilms containing silver nanoparticles were reported to exhibit antimicrobial activity at loadings and release rates of silver that are 100× lower than conventional dressings. Here, fabrication of composite microfilm constructs that provide a facile way to transfer the silver-loaded polymeric nanofilms onto wounds in vivo is reported. The construct is fabricated from a silver nanoparticle-loaded polymeric nanofilm that is laminated with a micrometer-thick-soluble film of polyvinylalcohol (PVA). When placed on a moist wound, the PVA dissolves, leaving the silver-loaded nanofilm immobilized on the wound-bed. In vitro, the immobilized nanofilms release <1 µg cm(-2) d(-1) of silver over 30 d from skin dermis and they kill 5 log10 CFUs of Staphylococcus aureus in 24 h. In mice, wounds inoculated with 10(5) CFU S. aureus presented up to 3 log10 less bacterial burden when treated with silver/nanofilms for 3 d, as compared to unmodified wounds. In uncontaminated wounds, silver/nanofilms allow normal and complete wound closure by re-epithelialization. Dissolvable microfilm constructs may overcome key limitations associated with current uses of silver in wound healing.