Biofilm Cohesiveness Measurement Using a Novel Atomic Force Microscopy Methodology

Abstract

ABSTRACT Biofilms can be undesirable, as in those covering medical implants, and beneficial, such as when they are used for waste treatment. Because cohesive strength is a primary factor affecting the balance between growth and detachment, its quantification is essential in understanding, predicting, and modeling biofilm development. In this study, we developed a novel atomic force microscopy (AFM) method for reproducibly measuring, in situ, the cohesive energy levels of moist 1-day biofilms. The biofilm was grown from an undefined mixed culture taken from activated sludge. The volume of biofilm displaced and the corresponding frictional energy dissipated were determined as a function of biofilm depth, resulting in the calculation of the cohesive energy. Our results showed that cohesive energy increased with biofilm depth, from 0.10 ± 0.07 nJ/μm 3 to 2.05 ± 0.62 nJ/μm 3 . This observation was reproducible, with four different biofilms showing the same behavior. Cohesive energy also increased from 0.10 ± 0.07 nJ/μm 3 to 1.98 ± 0.34 nJ/μm 3 when calcium (10 mM) was added to the reactor during biofilm cultivation. These results agree with previous reports on calcium increasing the cohesiveness of biofilms. This AFM-based technique can be performed with available off-the-shelf instrumentation. It could therefore be widely used to examine biofilm cohesion under a variety of conditions.