2001 Ocular Microbiology and Immunology Group, Abstract 12

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Antibiotic Resistance of Ciprofloxacin is Associated with Biofilm Formation in Ocular Isolates of Pseudomonas Aeruginosa
M. E. Zagans¹, F. Romen² and G. O=Toole²¹Section of Ophthalmology and ²Department of Microbiology, Dartmouth Medical School

Purpose: Bacteria can grow in a free-living, planktonic state or in a biofilm. Recent data suggests that growth in biofilms may predominate in most clinical and environmental settings. Growth in a biofilm has been associated with increased resistance to antibiotics, but this has not been studied extensively in ocular isolates. In this study we compare the susceptibility of ocular clinical isolates of Pseudomonas aeruginosa to ciprofloxacin when growing in a biofilm and under planktonic conditions.

Methods: Ocular isolates of P.aeruginosa used in this study were collected at the Campbell Ophthalmic Laboratory at the University of Pittsburgh. The minimal bacteriocidal concentration of planktonically grown bacteria to ciprofloxacin was determined for each isolate using a high throughput microtiter plate assay. Biofilm-specific antibiotic resistance was measured using two different methods. In the first method, isolates were grown in biofilms on a microtiter dish, followed by exposure to a range of ciprofloxacin concentrations. Bacterial cell viability was determined after 24 hours of exposure to ciprofloxacin. The second method utilizes the colony biofilm technique. The colony biofilm forms on a polycarbonate filter placed on an agar plate and cell viability is determined after the colony is exposed to ciprofloxacin for 24 and 48 hours.

Results: All P.aeruginosa isolates showed a statistically significant resistance to ciprofloxacin when grown in a biofilm as compared with planktonic conditions. This increase in resistance was 10-fold for most strains tested. Even isolates that exhibited a relatively poor biofilm formation phenotype in vitro demonstrated an increased resistance to ciprofloxacin when growing in a biofilm.

Conclusions: All ocular P.aeruginosa isolates tested demonstrated increased resistance to ciprofloxacin when grown in a biofilm rather than under planktonic conditions. Two independent methods for determining resistance were employed. A detailed understanding of conditions that promote bacterial growth in a biofilm and mechanisms for biofilm mediated antibiotic resistance may be important in developing novel strategies for preventing and treating bacterial eye infections.

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