Rapid, Sensitive, and Low-Cost EsCas13d and RPA-Based Assay Platform for Rapid and Accurate Identification of SVA

Rapid, sensitive, and user-friendly nucleic acid diagnostics are critical for controlling outbreaks of Seneca Valley virus (SVA). Here, we present Field-deployable Isothermal Enzymatic Lysis Detection (FIELD), an RNA detection platform for resource-limited settings that integrates rapid detection, operational simplicity, and minimal equipment requirements. FIELD employs a room-temperature lysis protocol to release viral nucleic acids within 2 min, followed by a single-step RPA-CRISPR/Cas13d reaction completed in 30 min. FIELD detects SVA RNA at 8 copies/μL using a plate reader and at 50 copies/μL via visual readout within 35 min, with no cross-reactivity against nontarget viruses. FIELD remains functional at both body (∼37 °C) and room (∼25 °C) temperatures, eliminating the need for temperature-controlled devices. Lyophilized FIELD reagents retain full activity for 1 week at room temperature and remain functional after long-distance transport and storage at 4 °C for up to two months, supporting field deployment. In a clinical evaluation of 88 field samples, FIELD achieved 100% sensitivity and specificity relative to RT-qPCR. Collectively, these features render FIELD well suited for point-of-care testing (POCT) beyond conventional laboratory settings.