Una estrategia de cascada DNAzyme-CRISPR para la detección sin preamplificación de Mycobacterium tuberculosis

Tuberculosis (TB) remains one of the most fatal infectious diseases worldwide, successful treatment is often limited by insufficient diagnostic capabilities. It creates a pressing need for diagnostic methods that combine high sensitivity, specificity, and operational robustness. In this work, we developed a DNAzyme-CRISPR cascade strategy (Dz-CRISPR) for the specific identification of the IS6110 sequence of Mycobacterium tuberculosis (MTB). This system integrated a thermodynamically stabilized hairpin probe, an Arch-shaped signal transduction switch, and an allosterically activated CRISPR-Cas12a cascade. The design enabled direct target recognition and subsequent signal amplification without a preamplification step, offering a simplified workflow with enhanced stability. The assay demonstrated a detection limit of 211.3 fM and exhibited high specificity by accurately discriminating the IS6110 from specific DNA sequence of non-tuberculous mycobacteria and other common respiratory pathogens. Validation using clinical bronchoalveolar lavage fluid samples further confirmed the method's reliable performance, reproducibility, and satisfactory recovery rates. Current Dz-CRISPR detection strategy provides a reliable and practical solution for tuberculosis diagnosis with high sensitivity, high specificity, and operational robustness, thus demonstrating potential for practical use in resource-constrained areas.