Abstract
A biofilm is a formation of surface‐associated microbial cells that are enclosed in a self produced extracellular polymeric substance matrix. This definition is acceptable for in‐vitro research but a clear definition for biofilm‐associated diseases has yet to be elucidated. A structural/morphological definition is currently used to define biofilms; additionally, physiological or molecular criteria will allow us to define biofilms in disease infection accurately. Our objective over the past several years has been to characterize and observe bacterial biofilms in wound infections using different wound models. To broaden our current understanding of biofilm morphology in wounds for this study we used epifluorescent and light microscopy to visualize wound pathogenic pseudomonas aeruginosa bacteria in a porcine partial thickness infection model.
Three experimental animals were used for this study. After animal preparation, partial thickness wounds were created on the backs of 3 animals. Wounds were then inoculated with 106 colony forming units/ml (CFU/ml) of Pseudomonas aeruginosa and covered for 48 hours to allow bacteria to colonize and infect the wound. Biopsies were obtained from normal skin, wounds before inoculation, wounds at 48 hours and wounds 48 hours after inoculation. Biopsies were processed and stained with Hematoxylin and Eosin for light microscopy and Calcofluor White and Ethidium Bromide for epifluorescence microscopy.
Images obtained using epifluorescent and light microscopy demonstrate that bacteria form aggregates of microcolonies. These structures are representative of bacterial biofilms and support the hypothesis that bacteria live as biofilms in wound infection. Although currently there is no established and accepted definition for biofilm associated diseases, we anticipate that more studies looking into physiological changes of these structures will clarify our current understanding of wound infection and treatment.