CRISPR-Based TB Saliva Test Created
7 August 2025
A point-of-care, CRISPR-based test can rapidly diagnose tuberculosis (TB) from saliva without the need for expensive equipment, researchers report.
SHINE-TB combines an efficient sample processing method together with an optimized CRISPR Cas13a/Cas12a assay to streamline detection of two conserved elements of the pathogen Mycobacterium tuberculosis (Mtb).
The U.S.-based researchers say theirs is the first test to combine recombinase polymerase amplification and detection while being compatible with lateral flow and lyophilization.
It could be particularly useful in under-resourced regions, where there is an urgent need for a simple, affordable means to diagnose tuberculosis.
“Our simple one-pot assay provides robust Mtb detection, taking us one step closer to accessible and reliable TB diagnostics,” reported Cameron Myhrvold, PhD, from Princeton University, and colleagues in Science Advances.
Current methods of diagnosing tuberculosis—such as culture-based approaches, sputum smear microscopy, or polymerase-chain-reaction techniques—are limited by tradeoffs in sensitivity, speed, and the need for specialist equipment.
CRISPR-based diagnostics offer a potential alternative, leveraging the high specificity and collateral cleavage activities of CRISPR-associated proteins such as Cas12 and Cas13 and pairing them with isothermal amplification techniques to offer both accessibility and accuracy.
The team developed the new test from their general-purpose diagnostic platform, SHINE (Streamlined Highlighting of Infections to Navigate Epidemics).
SHINE-TB consists of two parallelized one-pot reactions that combine isothermal amplification by recombinase polymerase amplification (RPA), in vitro transcription, and detection, including a Cas13a assay for detecting two conserved elements in the Mtb genome (IS6110 and IS1081) and a Cas12a assay for detecting human DNA as an internal control.
The team notes that high-burden TB countries often face substantial resource limitations and isothermal techniques provide equipment-free ways to amplify nucleic acids.
The test requires just a small sputum sample volume of 100 μl per extraction and around 2 μl per reaction.
The assay had a limit of detection of 69.0 CFU/ml for Mtb H37Rv and 80.5 CFU/ml for Mycobacterium bovis BCG in spiked sputum, with no cross-reactivity to diverse bacterial or fungal isolates.
In a small set of 13 smear-positive clinical sputum samples, it detected Mtb with 100% specificity (six of six) and 100% sensitivity (seven of seven) compared to culture.
The authors noted that the test meets the World Health Organization’s (WHO) target product profile benchmarks of at least 85% sensitivity for near point-of-care and at least 90% sensitivity for low-complexity TB tests.
They maintained: “SHINE-TB has comparable performance to other gold-standard methods for detecting TB DNA.”
Source: Inside Precision Medicine
