Athipanyasilp N.Saowpak S.Chaimayo C.Angkasekwinai N.Pattama A.Athipanyasilp A.Patchsung M.Aphicho K.Uttamapinant C.Horthongkham N.Mahidol University2026-02-142026-02-142026-02-03Microbiology Spectrum Vol.14 No.2 (2026) , e0229825https://repository.li.mahidol.ac.th/handle/123456789/115003Chikungunya virus (CHIKV), a major cause of acute febrile illness and joint pain, remains a significant public health threat in tropical regions. Rapid and accurate detection is essential for timely clinical management and outbreak control, particularly in resource-limited settings where real-time PCR (RT-qPCR) is often impractical. We developed and validated a SHERLOCK assay coupled with recombinase polymerase amplification for CHIKV RNA detection. Analytical performance was assessed by determining the limit of detection (LOD), cross-reactivity, clinical sensitivity and specificity, and predictive values. The assay achieved an LOD of 215 copies/reaction with no cross-reactivity against other alphaviruses or flaviviruses. Clinical testing of 146 plasma samples showed a sensitivity and specificity of 94.52% and 100% with lateral-flow readout and 97.26% and 100% with fluorescence readout, respectively. This study establishes a promising CRISPR-Cas13a-based SHERLOCK platform for CHIKV detection, demonstrating high analytical performance, rapid turnaround time, and potential for future adaptation to resource-limited settings.IMPORTANCEEarly and accurate detection of chikungunya virus (CHIKV) is critical for outbreak control, especially in resource-limited settings, where real-time PCR is not feasible. This study demonstrates that the CRISPR-Cas13a-based SHERLOCK platform, combined with RPA, achieves high diagnostic accuracy and a low detection limit, comparable to RT-qPCR. The assay's rapid turnaround time and simple lateral-flow readout make it a promising tool for point-of-care diagnostics during CHIKV outbreaks, potentially improving disease surveillance and clinical decision-making.Environmental ScienceBiochemistry, Genetics and Molecular BiologyMedicineImmunology and MicrobiologyArticleSCOPUS10.1128/spectrum.02298-252-s2.0-1050294817492165049741416796