Employing the xCELLigence® technology from ACEA Biosciences, a team of Spanish scientists have developed a real-time assay for screening anti-biofilm treatments and quantifying their efficacy
Biofilms are surface-attached communities of bacterial or fungal cells that are enmeshed in an extensive extracellular matrix which makes them more resistant to both antibiotics and the immune system. With biofilms estimated to be responsible for >60% of microbial infections, and 80% of chronic infections in humans, finding ways to disrupt established biofilms is of critical importance. In recent years there has been a growing interest in exploiting bacteriophages, or the lytic proteins that they encode, to treat biofilms.
Citing that the techniques currently used to identify anti-biofilm activities in phage-derived proteins have the “important shortcomings” of being laborious endpoint assays that suffer from poor reproducibility, in the recent issue of Frontiers in Microbiology a team of scientists lead by Diana Gutierrez report a proof of concept study using an xCELLigence Real-Time Cell Analysis instrument to monitor the disruption of clinically important Staphylococcus aureus biofilms. Within the proprietary xCELLigence microtiter plates that contain gold biosensors, biofilms of S. aureus were established and then exposed to different bacteriophage-derived proteins that catalyze degradation of the key biofilm extracellular polymers peptidoglycan or exopolysaccharide. The authors demonstrated that the degradation of these polymers, and the concomitant dissipation of the biofilm, causes a decrease in the xCELLigence biosensor signal that is both time- and dose-dependent. Importantly, this real-time biofilm disruption data correlated well with the data from traditional end point assays such as the labor intensive crystal violet staining technique.
Because each well of a xCELLigence microtiter plate can acquire thousands of data points, each individual well yields a complete time course for biofilm deposition or dissipation, greatly reducing the number of wells/plates needed and the total workload. Because of this, the authors were able to easily screen multiple S. aureus strains and four different lytic enzymes over a range of concentrations. The dose-response curves generated by this approach made it possible to accurately calculate key parameters such as the MBEC50 (the minimum biofilm eradicating concentration that removes 50% of the biofilm) for each treatment. In the ongoing effort to develop better biofilm therapeutics, Gutierrez and colleagues suggested that the synergistic effects of combining phage enzymes with different substrate specificities should be a key area of focus. With this in mind, they concluded that the reduced workload, improved efficiency, and higher reproducibility of the xCELLigence assay will make it possible to “quickly assess and compare by standardized parameters the disaggregating activity of phage anti-biofilm proteins.”