Abstract: To develop a fast, sensitive, specific TaqMan MGB probe real-time fluorescent quantitative PCR assay for detecting novel duck reovirus (NDRV), a pair of specific primers and a MGB probe were designed according to the conserved region of S3 segment of the NDRV genome. The results showed that recombinant plasmids were used to generate standard curve with correlation coefficient (R²) of 0.999, and the melting curve showed one specific peak. No cross-reaction was detected for classical muscovy duck reovirus (CDRV), avian influenza virus (AIV), duck Temubusu virus (DTMUV), duck hepatitis A virus (DHV-1), Newcastle diseases virus (NDV), duck plague virus (DPV) and muscovy duck parvovirus (MDPV). The limit detection of real-time RT-PCR was about 10 copies·μL^-1 for the target gene. The reproducibility tests in inter-assay and intra-assay indicated that the coefficients of variation were less than 2%. In addition, a collection of 239 clinical samples from Zhejiang during the year 2011-2015 were detected by the assay and conventional RT-PCR, respectively. The results showed that 75 samples were positive in the conventional RT-PCR test while 100 samples were positive in the one-step MGB probe real-time RT-PCR test. All the positive samples detected by conventional RT-PCR were positive by one-step MGB probe RT-PCR test, and the coincidence rate was 100%. The method established in this study could be useful for earlier rapid laboratory diagnosis and quantitative analysis of the infection of NDRV.

Key words: novel duck reovirus, MGB probe, fluorescent quantitative RT-PCR, diagnosis method