Development of an RT-ERA-CRISPR/Cas12a assay for duck Tembusu virus.

Duck Tembusu virus (DTMUV) has continued to threaten the duck industry in China since 2010. Therefore, the establishment of a rapid, specific, and sensitive method for the field detection of DTMUV is urgently needed. Herein, a reverse transcription enzymatic recombinase amplification (RT-ERA) assay was combined with the CRISPR/Cas12a system to target the DTMUV C gene. This assay exhibited high specificity, effectively distinguishing DTMUV from other common avian viruses. Its limit of detection reached 1 copy/μL DTMUV RNA. Moreover, this assay can be completed at 42 °C within 15 min using a thermostatic water bath. Additionally, we tested 30 clinical samples from infected ducks using this assay, and the results showed 100% concordance with SYBR green quantitative PCR results. In summary, this rapid, specific, and sensitive assay shows promising potential for DTMUV detection in the field.