Development of an isothermoal amplification-based assay for rapid visual detection of an Orf virus

Background: Orf virus (ORFV) is the causative agent of a severe infectious skin disease (also known as contagious ecthyma) in goats, sheep and other small ruminants. Importantly, ORFV also infect humans which causes a public health concern in the context of changing environment and increase in human populations. The rapid detection is critical in effective control of the disease and urgently needed. Results: A novel "point of care" molecular amplification assay for rapid visual detection of ORFV was developed based on isothermoal recombinase polymerase amplification (RPA) technology in combination with a simpler lateral flow immunoassay strip (ORFV RPA-LFD assay). The developed ORFV RPA-LFD assay was able to detect ORFV in less than 25 min. This assay was highly sensitive, with detection limit of as low as 80 copies/reaction, and highly specific, with no cross-reactions with capripox virus, foot-and-mouth disease virus and peste des petits ruminants virus. Furthermore, the ORFV RPA-LFD assay has good correlation with qPCR assay for detection of ORFV present in clinical samples. Conclusions: The developed ORFV RPA-LFD assay was a sensitive and specific method for rapid detection of ORFV, and has great potential as an onsite molecular diagnostic tool in control of Orf.