Rapid and sensitive on-site detection of PBoV G3 using an EsCas13d-based platform

Rapid, sensitive, and user-friendly nucleic acid diagnostic methods are crucial for controlling the spread of PBoV G3. In this study, we developed SENTRY (Single-step Enzymatic Nucleic acid Targeted Reporter Yield), a pathogen detection platform designed for on-site deployment in resource-limited settings. Combined with GENE (General Extraction-free Nucleic acid Exposure reagent), a room-temperature lysis protocol, SENTRY can release viral nucleic acids within 5 min. The addition of accessory proteins enhances the sensitivity of the one-step SENTRY platform and shortens the reaction time. SENTRY provides results within 30 min, with clinical sample detection rates matching those of qPCR, and shows no cross-reactivity with non-target viruses. The system operates efficiently at body temperature (∼37 °C) without requiring temperature control devices. Lyophilized reagents make SENTRY robust to transport conditions and extend shelf life. Overall, these features make SENTRY highly suitable for POCT outside conventional laboratory environments.