A highly multiplexed convection flow qPCR assay and platform delivering rapid, comprehensive identification and predictive AST of pathogens causing UTIs.

Project Summary Urinary tract infections (UTIs) are among the most prevalent community- and hospital-acquired infections (~10M annual medical visits), accounting for considerable morbidity and annual US healthcare spend of greater than $6B. While UTI pathogens are well understood, patients are frequently treated empirically. The lack of rapid, point of care molecular diagnostics that can identify UTI pathogens results in poor antimicrobial stewardship and high rates of UTI drug resistance. We propose to begin development of a rapid diagnostic assay and workflow that will concurrently detect all UTI pathogens and their associated resistance genes, suitable for point of care clinical applications. This assay is anticipated to allow for more effective personalized treatment of UTIs, reducing rates of antimicrobial resistance, and improving patient outcomes. As described in an upcoming Nature Biomedical Engineering paper, the Torus Biosystems’s qPCR system, Synestia™, has demonstrated reproducible 30-plex capability, accurate quantitation and SNP detection, sensitivity down to 10 genomic copies, and turnaround times of less than 30 minutes. The consumable is equipped with an embedded pre-quenched microarray that allows for real-time detection of amplification products without opening the reaction chamber. The sensitivity and dynamic range of the Synestia system has been evaluated across a range of input concentrations of DNA (101 to 105 copies) and the threshold value was found to be dependent on the log of the input DNA concentration. The platform has also successfully demonstrated rapid bacteria identification using a multiplex panel specific for 15 bacteria species. This Phase 1 application proposes to develop a 60-plex UTI panel for comprehensive assessment of uropathogen DNA within 30 minutes on the Synestia platform. In the subsequent Phase 2, we will develop, optimize, and validate a fully integrated consumable including urine sample prep (lysis and extraction) on the Synestia platform and validate clinical performance in collaboration with select clinical sites. The team anticipates that the demonstrated assay will constitute a solid basis for further development of a commercial point of care diagnostic test for urinary tract infections.Project Narrative Rapid and definitive detection of uropathogens is critical for effective, personalized treatment of urinary tract infections (UTIs) and the control of infection-associated antimicrobial resistance. We have developed a rapid molecular diagnostic system suitable for near patient, point-of-care use to diagnose infections and provide therapeutic selection decision support insights. This Phase I SBIR application outlines the design, optimization and validation of a rapid multiplex qPCR assay for the concurrent detection of 32 uropathogens and 18 drug resistance genes associated with UTIs.