A lyophilized RPA-CRISPR/Cas13d one-pot platform for rapid detection of porcine circovirus type 3

Rapid molecular diagnostics are essential for effective surveillance of infectious diseases in swine production systems. Here, we report SHARP (Single-step Hybrid RPA-CRISPR/EsCas13d Platform), a one-pot CRISPR-based platform for detection of porcine circovirus type 3 (PCV3). By integrating rapid nucleic acid release with CRISPR/Cas13d detection, a single-step one-pot detection system was established, enabling detection within 30 min under simplified reaction conditions. In the visual readout mode, the detection limit was 50 copies/μL, with no cross-reactivity observed against common swine viruses. After lyophilization and rehydration, the SHARP system showed consistent detection performance across 50 clinical samples, in agreement with qPCR results. These results indicate that SHARP provides a simplified workflow for rapid PCV3 detection, with potential for decentralized molecular surveillance in livestock production systems.