The aim of this research was to explore the interaction between ultrasound-activated microbubbles (MBs) and Pseudomonas aeruginosa biofilms, specifically the effects of MB concentration, ultrasound exposure and substrate properties on bactericidal efficacy. Biofilms were grown using a Centre for Disease Control (CDC) bioreactor on polypropylene or stainless-steel coupons as acoustic analogues for soft and hard tissue, respectively. Biofilms were treated with different concentrations of phospholipid-shelled MBs (107-108 MB/mL), a sub-inhibitory concentration of gentamicin (4 µg/mL) and 1-MHz ultrasound with a continuous or pulsed (100-kHz pulse repetition frequency, 25% duty cycle, 0.5-MPa peak-to-peak pressure) wave. The effect of repeated ultrasound exposure with intervals of either 15- or 60-min was also investigated. With polypropylene coupons, the greatest bactericidal effect was achieved with 2 × 5 min of pulsed ultrasound separated by 60 min and a microbubble concentration of 5 × 107 MBs/mL. A 0.76 log (83%) additional reduction in the number of bacteria was achieved compared with the use of an antibiotic alone. With stainless-steel coupons, a 67% (0.46 log) reduction was obtained under the same exposure conditions, possibly due to enhancement of a standing wave field which inhibited MB penetration in the biofilm. These findings demonstrate the importance of treatment parameter selection in antimicrobial applications of MBs and ultrasound in different tissue environments.
Keywords: Antibiotic; Antimicrobial resistance; Bacteria; Biofilm; Chronic wound; Microbubble; Substrate; Ultrasound.
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