Light-controlled RAA-Cas12a platform enables one-pot, ultrasensitive detection of vibrio parahaemolyticus in seafood

Vibrio parahaemolyticus (Vp) is an important foodborne pathogen threatening human health and marine economy, requires ultrasensitive tools for its early and accurate identification. In this study, we developed a light-controlled, integrated RAA (recombinase-mediated isothermal nucleic acid amplification)-Cas12a one-pot system that enables ultrasensitive and highly specific detection of Vp. The light-controlled system utilizing acetylated crRNA enables a fully enclosed reaction, where light-triggered activation allows time-resolved control and mitigates the low sensitivity arising from feedback between RAA amplification and Cas12a trans-cleavage. Compared with the conventional method with unacetylated crRNA, the light-controlled system increased sensitivity by two orders of magnitude, achieving a detection limit of 1.36 copies/μL. We further designed a portable device, which integrated with the proposed sensing strategy, achieving detection performance comparable to qPCR, demonstrating strong potential for rapid and high-throughput Vp detection.