Abstract
INTRODUCTION: Plant viral diseases threaten food security globally. Rapid, accurate and early detection of viral pathogens is critical for proactive surveillance and effective control. OBJECTIVES: This study aimed to develop a platform for the diagnosis of plant viruses in the field. METHODS: We developed an ALERT platform (AapCas12b coupled RT-LAMP Extraction-field RNA testing) that integrates RT-LAMP and CRISPR/Cas12b-based lateral flow system for the diagnosis of plant viruses. The portable metal incubator, coupled with the integrated microfluidic chip, enables parallel detection of six test samples in the field. Additionally, a raffinose additive solution effectively achieved the phase separation between the LAMP and CRISPR/Cas12b system through promoting dynamic diffusion of LAMP products amplicons. RESULTS: The ALERT platform exhibited comparable sensitivity to conventional RT-PCR, and field detection showed 100% diagnostic agreement with RT-PCR. CONCLUSIONS: This platform exhibits outstanding performance with high sensitivity and portability, and has great potential for rapid plant virus detection, especially in environments with resource-limited settings.