CRISPR/Cas13a一步法系统用于基孔肯雅病毒快速检测

Abstract

Early and accurate diagnosis of Chikungunya virus (CHIKV) infection is critical for controlling its outbreaks. CRISPR/Cas-based detection offers promise for pathogen identification, yet one-step CRISPR/Cas systems are limited by suboptimal sensitivity, field-deployability, and adaptability to complex clinical samples, hindering their use in rapid outbreak response. Here, we developed a CRISPR/Cas13a-based One-Step System for Rapid Detection of Emerging Viruses (CRISPR-CORE) and applied it during the CHIKV outbreak in Guangdong Province, China. Multidimensional optimizations enabled the CRISPR-CORE system to achieve a limit of detection of 5 copies/μL within 40 min. An extraction-free RNA release protocol for CHIKV in blood samples and a premixed reagent approach were implemented. Furthermore, a portable fluorescence detector was used to enhance user-friendliness in point-of-care (POC) settings. The clinical CHIKV genomic information was identified through hybrid capture sequencing, informing the design of highly specific CRISPR RNA (crRNA). Clinical validation across three regions yielded 92.6% sensitivity and 100% specificity, underscoring the applicability and reliability of the CRISPR-CORE system. Our system demonstrates its suitability for CHIKV outbreak detection. It facilitates rapid and POC testing for emerging viruses in resource-limited settings. Furthermore, it provides a universal strategy for the prevention and control of infectious diseases.