Development of a recombinase-aided amplification assay for detection of orf virus

Orf, caused by orf virus (ORFV), is an important zoonotic disease that infects goat and sheep, leading to huge economic losses. ORFV can also cause cutaneous lesions in people who come in close contact with the diseased animals. Although accurate diagnostic methods for ORFV infection exist, there is a need for a rapid, specific, and sensitive method for easy clinical application. Here, we successfully established a recombinase-aided amplification (RAA) assay for rapid detection of ORFV. The analytical sensitivity of the assay for ORFV detection is 1 × 10¹ copies per reaction. Moreover, no cross-reaction was observed with other common DNA viruses. A total of 45 archived suspected ORFV infected nasal scab skin samples were examined by RAA and SYBR Green-based real-time polymerase chain reaction (PCR). Compared with the real-time PCR assay, the kappa values of the RAA assay for ORFV detection was 0.845 (p <0.001), indicating that both assay results were fully in agreement. In conclusion, this detection assay provides a rapid, sensitive, and specific method for ORFV detection and is suitable for ORFV clinical testing.