Multiplex Real-Time PCR Method for Simultaneous Detection and Differentiation of Goat pox Virus, Sheeppox Virus, and Lumpy Skin Disease Virus.

Abstract

The diseases caused by the Capripoxvirus species have very similar symptoms and are difficult to distinguish clinically. According to a recent report, Capripoxvirus are not strictly host specific. This study aimed to identify the viruses from ovine (include sheep and goat) or bovine, which will assist in selecting the appropriate vaccine and correct measures to control diseases. Universal primers for all Capripoxvirus and specific probes for lumpy skin disease virus, sheeppox virus, and goatpox virus were designed and analyzed to identify the viruses from ovine (including sheep and goats) or bovine species. The parameters of the system, such as the annealing temperatures and the quantities of primers and probes used, were optimized. The sensitivity, specificity, and reproducibility were tested. Each probe showed a specific fluorescent signal, with no cross reaction with other pathogens that cause symptoms similar to those of the poxviruses. The LOD was 102 copies of the target genome DNA. The 557 local clinical samples and samples from Ethiopia were successfully detected and the results were consistent with a restriction fragment length polymorphism PCR analysis of the P32 and RPO30 genes and gene sequencing. This optimized real-time PCR detection system has good diagnostic sensitivity and specificity and can be used for the rapid and effective differential diagnosis of these diseases in goats, sheep, and cattle. It is a rapid detection method to distinguish the viruses from ovine (include sheep and goat) or bovine.