Antibiotic penetration and bacterial killing in a Pseudomonas aeruginosa biofilm model.

Objectives: Treating biofilm infections successfully is a challenge. We hypothesized that biofilms may be considered as independent compartments with particular pharmacokinetics. We therefore studied the pharmacokinetics and pharmacodynamics of tobramycin in a seaweed alginate-embedded biofilm model.

Methods: Seaweed alginate beads containing Pseudomonas aeruginosa were cultured in LB medium, sampled at day 1, 3, 5 or 7 and examined for the effect of treatment with tobramycin for 30 min. Treated beads were homogenized and the number of cfu was determined. The antibiotic concentration in the solution of homogenized beads was measured. Finally, beads were examined for live cells by Syto9 staining and for dead cells by propidium iodide staining using a confocal laser scanning microscope.

Results: The antibiotic level in each bead was relatively stable (range 30-42 mg/L; MIC = 1.5 mg/L). There were fewer cfu in the tobramycin-treated beads than the non-treated beads (P < 0.016) and bacterial killing was reduced as the culture period increased from 1 to 7 days. Throughout the study period, increasing size and more superficial positioning of the microcolonies within the beads were demonstrated by confocal laser scanning microscopy. More dead cells (measured by propidium iodide staining) were observed in the treated group of beads, which supports the results obtained by culture.

Conclusions: The present study, simulating the clinical pharmacokinetics of tobramycin, demonstrates fast absorption of tobramycin in an in vitro biofilm model. In addition, this model system enables parallel investigation of pharmacokinetics and pharmacodynamics, providing a model for testing new treatment strategies.