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Antimicrobial resistance (AMR) is a global health crisis driven significantly by the overuse of antibiotics. Most of this systemic misuse in human healthcare is a direct consequence of a diagnostic void created by the temporal gap between patient presentation and microbiological results. Traditional Antibiotic Susceptibility Testing (AST) relies on macroscopic biomass accumulation, a process constrained by bacterial generation times, and requires 48 to 72 hours. This delay forces clinicians to rely on empirical guidelines, treating patients based on probability rather than diagnostic certainty. Dependence on empirical therapy drives antibiotic overuse, fueling AMR. However, as resistance rates rise, the utility of empirical guidelines diminishes, creating a paradox where increasing resistance demands immediate diagnostic answers that current technology cannot provide.

This thesis demonstrates that the temporal constraints of AST are not immutable biological constants, but technological artifacts resulting from limited detection sensitivity. It establishes that bacterial cells respond to effective antibiotics within minutes, and that detecting these immediate physiological changes requires only a sufficiently sensitive method. This detection is enabled by a novel nanofluidic platform that actively captures, maintains, and images individual bacterial cells. By utilizing phase-contrast microscopy, the method quantifies instantaneous single-cell growth rates in real-time. Paper I establishes the "Fast Antibiotic Susceptibility Testing" (FASTest) method, demonstrating that susceptibility profiles for Escherichia coli against nine antibiotics can be determined in 30 minutes, and that it correctly distinguishes resistant bacteria from susceptible ones across 49 uropathogenic E. coli isolates. Paper II validates the platform's robustness through a complex "lab-on-a-chip" application involving a multi-step molecular biology protocol for in situ genotyping. Finally, Paper III shows clinical feasibility of the method for 30-minute AST for sepsis using either positive blood cultures or isolated colonies.

The research presented in this thesis has been successfully translated from an academic concept into a fully automated medical device, the PA-100 AST System, which was recently awarded the Longitude Prize on Antimicrobial Resistance. This technology demonstrates that shifting observation from the population to the single cell bridges the diagnostic void, enabling the transition from empirical prescribing to evidence-based precision medicine for urinary tract infections at the Point-of-Care.