Correcting a Fundamental Flaw in the Paradigm for Antimicrobial Susceptibility Testing

The emergence and prevalence of antibiotic-resistant bacteria are an increasing cause of death worldwide, resulting in a global 'call to action' to avoid receding into an era lacking effective antibiotics. Despite the urgency, the healthcare industry still relies on a single in vitro bioassay to determine antibiotic efficacy. This assay fails to incorporate environmental factors normally present during host-pathogen interactions in vivo that significantly impact antibiotic efficacy. Here we report that standard antimicrobial susceptibility testing (AST) failed to detect antibiotics that are in fact effective in vivo; and frequently identified antibiotics that were instead ineffective as further confirmed in mouse models of infection and sepsis. Notably, AST performed in media mimicking host environments succeeded in identifying specific antibiotics that were effective in bacterial clearance and host survival, even though these same antibiotics failed in results using standard test media. Similarly, our revised media further identified antibiotics that were ineffective in vivo despite passing the AST standard for clinical use. Supplementation of AST medium with sodium bicarbonate, an abundant in vivo molecule that stimulates global changes in bacterial structure and gene expression, was found to be an important factor improving the predictive value of AST in the assignment of appropriate therapy. These findings have the potential to improve the means by which antibiotics are developed, tested, and prescribed.