CRISPR-Based Saliva Test Detects Tuberculosis Directly from Sputum

Tuberculosis remains a leading infectious killer worldwide, and diagnosing it quickly and accurately is still a major challenge. Current methods such as culture, sputum smear microscopy, and PCR have trade-offs in speed, sensitivity, and accessibility, often requiring costly equipment and specialized expertise. In low-resource regions, there is a pressing need for a simple, affordable tool to detect TB reliably at the point of care. Researchers have now developed a rapid, sputum-based saliva diagnostic test to meet this demand.

SHINE-TB, a point-of-care diagnostic test developed by researchers at Princeton University (Princeton, NJ, USA), rapidly detects tuberculosis from sputum without expensive equipment. The test integrates optimized CRISPR Cas13a/Cas12a assays with efficient sample processing to identify two conserved elements of the TB pathogen. Created from the SHINE diagnostic platform, it is the first test to merge recombinase polymerase amplification and detection while being compatible with lateral flow and lyophilization for field use.

Image: Researchers have developed a CRISPR-based tuberculosis saliva test (Photo courtesy of 123RF)

SHINE-TB integrates two parallel one-pot reactions: a Cas13a assay for detecting IS6110 and IS1081 sequences in the TB genome, and a Cas12a assay for detecting human DNA as an internal control. The isothermal amplification approach eliminates the need for expensive thermocyclers, and the assay requires only 100 μl of sputum per extraction and 2 μl per reaction. This design makes it highly suited to high-burden, low-resource settings.

In laboratory validation, the assay achieved limits 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 other bacteria or fungi. In clinical testing with 13 smear-positive sputum samples, it demonstrated 100% specificity and 100% sensitivity compared to culture, according to the study published in Science Advances.

The test meets World Health Organization target product profile benchmarks for TB diagnostics, including at least 85% sensitivity for near point-of-care use and 90% sensitivity for low-complexity settings. Its compatibility with lateral flow formats and stable lyophilized reagents positions it for deployment in remote or under-equipped healthcare facilities, bringing it one step closer to accessible and reliable TB diagnostics.

"Our simple one-pot assay provides robust Mtb detection, taking us one step closer to accessible and reliable TB diagnostics," stated the authors.