Bacterial Fluorescence Imaging Detects Planktonic Bacteria and Biofilm in Vitro

Background: Chronic wounds commonly harbor polymicrobial biofilms, and certain combinations of microbes may result in more aggressive infections. Culture-based diagnostics identify dominant microorganisms as well as their antimicrobial susceptibility, however the lag time to obtain those results (3 days – 4 weeks) significantly impacts wound care and treatment. Bacterial fluorescence imaging uses safe violet light to detect fluorescent properties of bacteria. Many species of bacteria utilize aminolevulinic acid (ALA) to produce porphyrins, which fluoresce red under specific wavelengths of light. This imaging device allows physicians to detect bacterial bioburden in a wound in real time, and can direct specimen sampling to the area with the heaviest bioburden, improving diagnostic capabilities. Prior work demonstrated its detection of bacteria from in vivo and in vitro monomicrobial planktonic cultures.

Purpose: We have investigated its capability to detect biofilm using our polymicrobial in vitro biofilm model, consisting of Bolton’s broth and bovine plasma, which is representative of the chronic wound environment.

Methods and Results: Staphylococcus aureus, Escherichia coli, and Enterobacter cloacae were selected as representative wound pathogens. When grown in the in vitro wound-like model for seven days, followed by the induction of porphyrin production by the addition of ALA for 24 hours, we demonstrated that the device can readily detect bacterial fluorescence from both monomicrobial and polymicrobial biofilms.

Conclusion: These data demonstrate that bacterial fluorescence imaging detects porphyrin-positive species of bacteria growing both planktonically and as a biofilm, as well as monomicrobial and polymicrobial communities, which further validates the clinical capability and relevance of the device for use in wound care.