We have established a CRISPR-Cas12a-based detection system that can ultra-sensitively detect trace amount of SARS-CoV-2 RNA in blood to diagnose COVID-19, predict disease severity, and evaluate infection resolution. We have adapted this approach to develop a blood-based multiplexed CRISPR-mediated TB diagnosis (CRISPR-TBD) assay that can detect circulating Mtb cfDNA, including SNPs responsible for drug-resistant TB. Our preliminary data from longitudinal serum samples of patients undergoing TB treatment provide strong proof-of-principle evidence for the clinical utility of this platform. We have adapted this approach to a paper strip-based point of care (POC) CRISPR-TBD m>detectionm> platform suitable for use in resource-limited regions with high TB burden, without decreasing assay sensitivity. We now propose to: 1) systematically optimize all CRISPR-TBD paper strip assay steps to improve quantitative m>detectionm> of Mtb-cfDNA in POC settings; 2) evaluate the performance of this POC assay to diagnose pulmonary and extrapulmonary TB and to identify drug-sensitive and -resistant TB cases; 3) quantify Mtb-cfDNA changes in serum during TB treatment as a measure of treatment efficacy or failure and for early m>detectionm> of nascent drug resistance; and 4) in-field validate the diagnostic performance of this POC assay in an independent TB patient cohort when performed in a clinical laboratory in a high endemic TB region and evaluate in parallel the performance our predictive Mtb-cfDNA model for TB treatment.