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2009 OMIG, Abstract 16

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Inhibition of Staphylococcal Biofilm Formation in vitro by DuraSite and AzaSite.
E. Wu, F.S. Mah, E.G. Romanowski, R.P. Kowalski, Y.J. Gordon, and R.M.Q. Shanks
The Charles T. Campbell Laboratory, UPMC Eye Center, Ophthalmology and Visual Sciences Research Center, Eye and Ear Institute, Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA

Purpose. To test the hypothesis that topical AzaSite® and its components (DuraSite®, azithromycin (AZ), benzalkonium chloride (BAK)) would inhibit biofilm formation by clinical blepharitis isolates of Staphylococcus aureus and coagulase negative Staphylococcus (CNS). Bacterial biofilms are associated with chronic and antibiotic resistant infections, and may contribute to infectious blepharitis. 
Four AZ resistant, and 4 susceptible de-identified clinical isolates were used in all experiments.  Serial dilutions of AzaSite, DuraSite, AZ, and BAK were added to biofilm cultures in polystyrene microtiter plates, with bacterial inocula normalized to A600 nm = 0.01.  Biofilms were allowed to form for 20 hours at which time gross bacterial growth was determined by reading absorbance at 600 nm, non-biofilm cells were removed by washing with water, and biofilms were stained with crystal violet (CV) (0.1% w/v).  To quantitate biofilms, CV was solubilized with glacial acetic acid (30% v/v) and measured by absorbance at 590 nm.  Experiments were repeated at least two times on different days in triplicate. 
Results.  All AZ susceptible strains demonstrated a statistically significant reduction in bacterial growth (p<0.001) and biofilm formation (p<0.05) at all studied dilution strengths of Azasite® and AZ.  Resistant strains treated with Azasite® at concentrations of 0.5%, 0.25%, and 0.125% demonstrated a statistically significant reduction in biofilm formation compared to controls (p<0.005), whereas planktonic growth showed no statistically significant difference. AZ did not affect planktonic growth in AZ resistant strains but did have a small but statistically significant effect on biofilm formation at concentrations of 1%, 0.5%, 0.25%, and 0.125% in most tested strains.  DuraSite® was noted to have no significant impact on planktonic growth at any dilution but conferred a statistically significant reduction in biofilm formation at concentrations of 1%, 0.5%, and 0.25% in all studied bacterial strains (p<0.05). BAK inhibited both planktonic growth and biofilm formation between concentrations of 0.0375% and 0.0042% (p<0.005). 
Conclusions. When bacterial growth was inhibited by AZ, Azasite®, or BAK, biofilm formation was also prevented as expected. AZ and Azasite® inhibited biofilm formation at subinhibitory doses, suggesting that they inhibit the ability of staphylococci to attach to surfaces.  Unexpectedly, DuraSite®, while having no effect on planktonic bacterial growth, significantly inhibited biofilm formation at concentrations from 0.25-1.00% on all tested strains.  Supported by Inspire, Inc., RPB
Disclosure Code: F,C

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