Establishment of a duplex RT-PCR assay for simultaneous detection of Rift valley fever virus and peste des petits ruminants virus

Abstract

Rift valley fever(RVF) and peste des petits ruminants(PPR) are both highly contagious diseases of ruminants caused by RVF virus (RVFV) and PPR virus (PPRV), which are both the notifiable multiple species diseases in the OIE list. In order to establish a rapid method for detecting of RVF and PPR, two pairs of primers were designed according to the nucleotide sequence information of RVFV and PPRV published in GenBank. The duplex RT-PCR which contained two pairs of the primers and two templates in one PCR system was established and then the reaction conditions were optimized including temperature and primers concentration. The results showed that the target sequence-products of RVFV and PPRV were 318bp and 589bp, and the duplex RT-PCR assay could be used to detect the RVFV and PPRV with good sensitivity and the detection limit were approximately 53 copies of PPRV and 68 copies of RVFV. Besides, the specificity test proved that there were no cross-reaction with Foot and mouth disease virus, sheep pox virus, E.coli , Salmonella and Pasteurella multocida isolated from ruminants and blood sample of healthy sheep.