Metabolism‐Based Biomarkers for Rapid Phenotypic Antibiotic Susceptibility Testing

The accelerating global crisis of antimicrobial resistance (AMR) demands rapid and accurate methods for antibiotic susceptibility testing (AST). Conventional phenotypic assays remain the gold standard but are hindered by long culture times, while genotypic tests cannot reliably predict phenotypic resistance. In recent years, metabolism-based AST has emerged as a promising alternative, enabling the rapid detection of bacterial responses to antibiotics through shifts in metabolic activity. These approaches bridge molecular speed with phenotypic precision, allowing susceptibility determination within hours, or even minutes, without requiring cell proliferation. In this review, we summarize the latest advances in metabolism-based biomarkers for rapid AST. First, we discuss how antibiotics influence bacterial metabolism, linking resistance mechanisms to metabolic activities. We then summarize emerging metabolic biomarkers, categorized by their physiological underpinnings: nutrient uptake, respiratory activity, metabolic reprogramming, and enzymatic function. Finally, we list key challenges and future directions toward deployable metabolism-based AST platforms.