Extended antimicrobial susceptibility assay for Staphylococcus aureus isolates from bovine mastitis growing in biofilms

Staphylococcus aureus is one of the most prevalent causes of bovine mastitis. The antimicrobial treatment of this disease is currently based on antimicrobial susceptibility tests according to CLSI standards. However, various studies have shown that there is a discrepancy between the results of this standard susceptibility test and the actual cure rate of the applied antimicrobial treatment. Increasing evidence suggests that biofilm formation by S. aureus is associated with this problem. The currently available antimicrobial susceptibility assays for bacteria growing in biofilms, are not considered reliable enough for routine application. Therefore, the objective of this study was to further develop a susceptibility test for bacteria growing in biofilm, suitable for routine testing of the antimicrobial susceptibility of S. aureus. With the expansion of the available MBEC assay to an extended biofilm susceptibility test, that comprises 2 and 4 consecutive days of antimicrobial challenge, the antimicrobial susceptibility for S. aureus growing in biofilm was further analysed. The results showed clear differences between strains and various antimicrobial agents with respect to the effect of longer duration of the antimicrobial challenge on the eradication of S. aureus growing in biofilm. The extended biofilm susceptibility test also indicates that each bacterial strain requires a specific duration of antimicrobial therapy, which cannot be derived from a standard susceptibility test or from a 24-h biofilm susceptibility test.