CRISPR/Cas12a Technology Combined with Immunochromatographic Strips for the Portable Detection of SFTS Bunyavirus

Severe Fever with Thrombocytopenia Syndrome (SFTS), caused by SFTS virus (SFTSV), is a widely distributed infection with significant mortality. Diagnosis in resource-limited settings remains challenging. For rapid and convenient diagnosis, we developed a portable rapid diagnosis method that combines Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas12a technology with immunochromatographic test strips. The SFTSV RNA was amplified by reverse transcription loop-mediated isothermal amplification (RT-LAMP). The homologous target sequence and single stranded DNA (ssDNA) reporter gene were cleaved by CRISPR/Cas12a in parallel, and ssDNA probes labeled with FAM fluorescein and biotin were captured by an immunochromatographic strip. Finally, the signal on the immunochromatographic strips became visible to the naked eye. Based on CRISPR/Cas12a, a rapid SFTSV detection method was developed, featuring simplicity, rapidity, low cost, and ease of use. The method was applied for the nucleic acid detection of SFTSV in 40 clinical serum samples and compared with RT-polymerase chain reaction (PCR). The new method showed 100% sensitivity and 100% specificity with a detection agreement rate of 100%. The minimum detection limit of the method was 2.5 copies/μL, and no cross-reactivity with nucleic acids from other common pathogens was observed. Detection can be completed within 80 min, and results are observable with the naked eye. For the analysis of clinical samples, the method exhibits good detection performance and thus provides an attractive option for the nucleic acid detection of SFTSV in point-of-care and resource-limited medical settings.