Adhesion of biofilms on titanium measured by laser-induced spallation

Eradication of established implant-associated and bacterial biofilm-forming infections remains difficult in part because these biofilms remain well-adhered to the implant surface. Few experimental techniques are available to measure macro-scale strength of bacterial biofilm-implant adhesion. We have adapted the laser spallation technique to compare the macro-scale adhesion strength of biofilms formed on titanium. By using a rapid pressure wave (35 ns) to load the interface, we prevent disturbance of the biofilm surface prior to measurement, and preclude the time necessary for the biofilm to respond to and adapt under loading. Biofilms of Streptococcus mutans, a Gram-positive bacterium associated with human dental caries (cavities) were cultured directly on commercially pure titanium within our custom substrate assembly. Growth conditions were varied by adding sucrose to the Todd Hewitt Yeast (THY) broth: THY control, 37.5 mM, 75 mM, 375 mM, and 750 mM sucrose. Multiple locations on each biofilm were loaded using the laser spallation technique. Loading pressure wave amplitude was controlled by adjusting laser fluence, energy per area. Initially, addition of sucrose to the media increased biofilm adhesion to titanium. However, once a saturation concentration of 75 mM sucrose was reached, increasing the sucrose concentration further resulted in a decrease in biofilm adhesion. This study is the first demonstration of the adaptation of the laser spallation technique to measure bacterial biofilm adhesion. Establishment of this macro-scale biofilm adhesion measurement technique opens the door for many biofilm-surface adhesion studies. We anticipate further work in this area towards understanding the complex relationships among bacteria species, environmental factors, surface characteristics, and biofilm adhesion strength.