Ultrasonic destruction of microbial biofilms revealed by confocal microscopy

Biofilms are communities of microbes that adhere to a surface by means of a secreted polymeric matrix. The formation of antibiotic resistant biofilms on internal biomedical devices has contributed to a recent growth in the number of pernicious nosocomial infections (Ikeno, 2007). Ultrasound has been previously demonstrated to kill planktonic bacteria via oxidation, DNA damage and membrane disruption (Vollmer, 1998). In order to study the destructive effect of insonification on biofilms, we have developed three biofilm growth systems that incorporate ultrasonic transducers for direct delivery of ultrasonic energy to the biofilm. E. coli and P. aeruginosa biofilms were grown for 18-24 hours under a continuous flow of growth medium. Subsequently, biofilm destruction by insonification was observed in real time by confocal microscopy. Insonification at 810 kHz for 10 minutes at a O.2V power level is sufficient to remove approximately 50% of P. aeruginosa biofilm density from a glass microscope slide, and up to 90% removal is possible using a 0.6 V power level. Pre-treatment with low levels of the antibiotics chloramphenicol or tetracycline results in greater effectiveness of ultrasound in destroying the biofilm. We conclude that insonification in combination with other antimicrobial agents may be a useful tool for removing biofilms from medical and environmental systems