Quantifying Staphylococcus Aureus Biofilm Formation on Stainless Steel and Titanium Alloy Used in Orthopedic Fracture Fixation

In order to assess and compare the antibacterial property of implants surfaces, a standard method is needed to quantify bacterial load. This study evaluated the effectiveness of three quantifying methods, namely, (I) crystal violet staining analysis, (II) ultrasound detachment with viable cell counts, and (III) confocal laser scanning microscopy for characterizing S. aureus Seattle 1945 (ATCC 25923) biofilm on metallic coupons. The accuracy of the results, time for completion, and ease of use of methods were compared. The crystal violet method is relatively faster and more straightforward for analyzing biofilm formation. However, the accuracy of the confocal laser scanning microscopy method is found to be considerably higher than that of the other methods. Confocal laser scanning microscopy method is considered to be more time-consuming for data collection and analysis and costlier. The ultrasound detachment followed by viable cell count of recovered cells is recommended for biofilm quantification analysis on orthopedic materials when there is a large number of samples (more than ten samples). This info could provide guidelines that would facilitate the selection of suitable method for quantifying biofilm formation on orthopedic implants based on investigators’ consideration on method accessibility, assay cost, assay time, and complexity of method.