ABSTRACT
BACKGROUND: The minimum inhibitory concentration (MIC) does not provide information on the efficacy of antimicrobial agents against infections involving biofilms, which are many times more resistant than planktonic forms of bacteria. This report is on the design and initial trial of a device for growing standard biofilms and testing antimicrobial agents. METHODS: We constructed a durable, autoclaveable laboratory model biofilm fermenter (LMBF) that holds hydroxyapatite discs 300 microm below a surface onto which an artificial saliva medium drips at a rate comparable to human salivary flow. Inoculated with Streptococcus sanguinis, the device formed biofilms that were swept with a Teflon wiper under aerobic conditions. Five-day-old biofilm-coated discs were aseptically removed and placed in 3 ml of sterile saline, 0.12% chlorhexidine gluconate, or 0.1% phosphate-buffered chlorine dioxide mouthwash for 1 minute. The discs and test agent were immediately diluted with saline to 10 ml, vortexed for 30 seconds, serially diluted, plated on blood agar, and incubated anaerobically 2 days. Bacterial counts were done, and the MIC of each mouthwash was determined. RESULTS: In tests with sterile water and sterile medium, the device maintained a closed system. After inoculation with S. sanguinis, a steady state was reached at day 5. Chlorhexidine at stock concentration achieved about a 2 log10 reduction (P = 0.002), but never achieved complete killing. Chlorine dioxide had no significant effect. The MIC against planktonic S. sanguinis was 112.8 microg/ml for chlorhexidine and 9.0 microg/ml for chlorine dioxide. CONCLUSIONS: The LMBF generates and maintains a single-species oral model biofilm to a steady state and enables in vitro tests of disinfectant mouthwashes in simulated clinical use. It should be usable for more advanced tests of multiple species biofilms.
