Chemiluminescent Protease Probe for Rapid, Sensitive, and Inexpensive Detection of Live .

Tuberculosis (TB) is a top-ten cause of death worldwide. Successful treatment is often limited by insufficient diagnostic capabilities, especially at the point of care in low-resource settings. The ideal diagnostic must be fast, be cheap, and require minimal clinical resources while providing high sensitivity, selectivity, and the ability to differentiate live from dead bacteria. We describe here the development of a fast, luminescent, and affordable sensor of Hip1 (FLASH) for detecting and monitoring drug susceptibility of (). FLASH is a selective chemiluminescent substrate for the protease Hip1 that, when processed, produces visible light that can be measured with a high signal-to-noise ratio using inexpensive sensors. FLASH is sensitive to fmol of recombinant Hip1 enzyme and can detect as few as thousands of cells in culture or in human sputum samples within minutes. The probe is highly selective for compared to other nontuberculous mycobacteria and can distinguish live from dead cells. Importantly, FLASH can be used to measure antibiotic killing of in culture with greatly accelerated timelines compared to traditional protocols. Overall, FLASH has the potential to enhance both TB diagnostics and drug resistance monitoring in resource-limited settings.