CRISPR-based LAMP assays detect tuberculosis with high accuracy in early study

Researchers have developed and clinically evaluated two LAMP-based CRISPR diagnostic pipelines that target the IS6110 sequence of Mycobacterium tuberculosis, the bacterium that causes tuberculosis. The study, published in Annals of Clinical Microbiology and Antimicrobials, reports that both pipelines achieved a limit of detection of 10 copies per microlitre and showed 100 percent specificity against non-tuberculous mycobacteria, while remaining compatible with lateral flow readouts that could support point-of-care use.

Tuberculosis remains a major global health challenge, and the authors underscore the need for rapid and accessible diagnostic tools suitable for resource-limited settings.

The two pipelines share a common front end but differ in their detection chemistry. Both are two-pot formats, meaning the amplification and detection steps are carried out in separate reactions. One pipeline relies on Cas12a-mediated DNA detection, in which the CRISPR enzyme recognises amplified DNA targets. The other uses Cas13a-mediated RNA detection, which requires an additional in vitro transcription, or IVT, step that converts amplified DNA into RNA before the Cas13a enzyme can act on it.

Analytical sensitivity was determined using serial dilutions of commercial M. tuberculosis DNA controls. Both pipelines reached the same limit of detection of 10 copies per microlitre in these controlled dilution experiments, indicating that the underlying detection chemistry can register very small quantities of target genetic material under laboratory conditions.

Clinical validation was performed using 76 culture-characterized clinical specimens, including both respiratory and extra-pulmonary samples. Assay performance was assessed using two readout formats: fluorescence measurements and lateral flow tests.

When tested on the 76 clinical samples, the LAMP-Cas12a assay demonstrated higher global diagnostic sensitivity than the LAMP-IVT-Cas13a pipeline, at 89 percent versus 76 percent. The authors suggest this difference is likely due to the additional IVT step required for Cas13a detection.

Despite the lower sensitivity in this head-to-head comparison, the researchers conclude that RNA-targeting Cas13a remains of interest for future applications aimed at identifying active disease.

Overall, the team concludes that LAMP/CRISPR diagnostics represent a promising alternative to conventional tuberculosis tests, with Cas12a currently offering superior performance in two-pot formats. The findings are based on a single clinical evaluation of 76 specimens characterised by culture, and the article is being shared early as a peer-reviewed, accepted version that is subject to further edits before the final version of record, so the reported performance figures should be read in that context.

The work adds to a growing body of research on CRISPR-Cas-based detection of M. tuberculosis, a field in which related studies have explored platforms such as Cas14a-enabled detection of rifampin resistance and one-pot biplex RPA-Cas assays using tongue swabs.

Subject of Research: Development and clinical evaluation of LAMP-CRISPR diagnostic pipelines using Cas12a and Cas13a for detecting Mycobacterium tuberculosis.

Article Title: Rapid accurate platform detection of Mycobacterium tuberculosis using Cas12a and Cas13a

Article References: Pablo-Marcos, D., Estévez-Manso, L., Fernández-Diego, L., García-García, S., Ortiz-Cartagena, C., Pacios, O., Blasco, L., Osorio-Chavez, J. S., Rodríguez-Grande, J., Miguez-Rey, E., Roiz-Mesones, M. P., Calvo-Montes, J., & Tomás, M. (2026). Rapid accurate platform detection of Mycobacterium tuberculosis using Cas12a and Cas13a. Annals of Clinical Microbiology and Antimicrobials. https://doi.org/10.1186/s12941-026-00894-z

Image Credits: AI Generated

DOI: 10.1186/s12941-026-00894-z

Keywords: tuberculosis, Mycobacterium tuberculosis, CRISPR, Cas12a, Cas13a, LAMP, diagnostics, lateral flow, point-of-care testing, IS6110, in vitro transcription, diagnostic sensitivity