Mechanical destruction of pseudomonas aeruginosa biofilms by ultrasound exposure

Abstract

Medical implants are prone to colonization by bacterial biofilms, which are highly resistant to antibiotics. Normally, surgery is required to replace the infected implant. One promising non-invasive treatment option is to destroy the biofilm with high-intensity focused ultrasound (HIFU) exposure. In our study, Pseudomonas aeruginosa bacterial biofilms were grown on graphite disks in a flow chamber for three days prior to exposing them to ultrasound pulses of varying duration or burst period. The pulses were 20 cycles in duration at a frequency of 1.1 MHz from a spherically focused transducer (f/1, 63 mm focal length), creating peak compressional and rarefactional pressures at the disk surface of 30 and 13 MPa, respectively. P. aeruginosa were tagged with GFP and cells killed by HIFU were visualized using propidium iodide, which permeates membranes of dead cells, to aid determining the extent of biofilm destruction and whether cells are alive or dead. Our results indicate that a 30-s exposure and 6-ms pulse period or those combinations with the same number of pulses, were sufficient to destroy the biofilm and to kill the remaining cells. Reducing the number of pulses decreased biofilm destruction, leaving more dead and live bacteria on the surface. Abstract. Medical implants are prone to colonization by bacterial biofilms, which are highly resistant to antibiotics. Normally, surgery is required to replace the infected implant. One promising non-invasive treatment option is to destroy the biofilm with high-intensity focused ultrasound (HIFU) exposure. In our study, Pseudomonas aeruginosa bacterial biofilms were grown on graphite disks in a flow chamber for three days prior to exposing them to ultrasound pulses of varying duration or burst period. The pulses were 20 cycles in duration at a frequency of 1.1 MHz from a spherically focused transducer (f/1, 63 mm focal length), creating peak compressional and rarefactional pressures at the disk surface of 30 and 13 MPa, respectively. P. aeruginosa were tagged with GFP and cells killed by HIFU were visualized using propidium iodide, which permeates membranes of dead cells, to aid determining the extent of biofilm destruction and whether cells are alive or dead. Our results indicate that a 30-s exposure and 6-ms pulse period or those combinations with the same number of pulses, were sufficient to destroy the biofilm and to kill the remaining cells. Reducing the number of pulses decreased biofilm destruction, leaving more dead and live bacteria on the surface.

Comparison of techniques for counting prokaryotes in marine planktonic and biofilm samples

“…The efficiency of the ultrasound procedure for detaching prokaryote cells from steel, glass, ceramic and plastic surfaces has been reported (Xu et al 2012, Sgier et al 2016, but not from plywood, which shows a more heterogeneous surface. Moreover, cell damage has not been evaluated.…”

“…However, FC as well as some EFM techniques require pre-treatments to detach the individual bacterial cells from surfaces without rupturing cells. In the literature, one of the main procedures for removing bacterial biofilms from surfaces is ultrasound treatment (sonication) , Xu et al 2012, Kerstens et al 2015. However, this technique should be used with care, because ultrasound waves have the capacity to kill bacterial cells, depending on the frequency applied (Xu et al 2012, Kerstens et al 2015.…”

“…In the literature, one of the main procedures for removing bacterial biofilms from surfaces is ultrasound treatment (sonication) , Xu et al 2012, Kerstens et al 2015. However, this technique should be used with care, because ultrasound waves have the capacity to kill bacterial cells, depending on the frequency applied (Xu et al 2012, Kerstens et al 2015. Furthermore, the age of the biofilm could interfere in the composition and quantity of EPS and cell abundance (Flemming and Wingender 2010).…”