A streamlined CRISPR-based test for tuberculosis detection directly from sputum.

() is a major threat to global health, and there is an urgent need for affordable, simple tuberculosis (TB) diagnosis in underresourced areas. Here, we combine recombinase polymerase amplification with Cas13a and Cas12a detection to create two parallelized one-pot assays that detect two conserved elements of ( and ) and a human DNA internal control. These assays are compatible with lateral flow and can be readily lyophilized. Our final assay showed a limit of detection of 69.0 CFU per milliliter for H37Rv and 80.5 CFU per milliliter for BCG in spiked sputum, with no cross-reactivity to diverse bacterial or fungal isolates. Clinical tests on 13 blinded sputum samples revealed 100% (six of six) sensitivity and 100% (seven of seven) specificity compared to culture. SHINE-TB streamlines TB diagnosis from sample to answer by combining amplification and detection while being compatible with lateral flow and lyophilization.