BLV-CoCoMo Dual qPCR Assay Targeting LTR Region for Quantifying Bovine Leukemia Virus: Comparison with Multiplex Real-Time qPCR Assay Targeting pol Region.

The proviral load (PVL) of the bovine leukemia virus (BLV) is a useful index for estimating disease progression and transmission risk. Real-time quantitative PCR techniques are widely used for PVL quantification. We previously developed a dual-target detection method, the "Liquid Dual-CoCoMo assay", that uses the coordination of common motif (CoCoMo) degenerate primers. This method can detect two genes simultaneously using a FAM-labeled minor groove binder (MGB) probe for the BLV long terminal repeat (LTR) region and a VIC-labeled MGB probe for the BoLA-DRA gene. In this study, we evaluated the diagnostic and analytical performance of the Dual-CoCoMo assay targeting the LTR region by comparing its performance against the commercially available Takara multiplex assay targeting the pol region. The diagnostic sensitivity and specificity of the Liquid Dual-CoCoMo assay based on the diagnostic results of the ELISA or original Single-CoCoMo qPCR were higher than those of the Takara multiplex assay. Furthermore, using a BLV molecular clone, the analytical sensitivity of our assay was higher than that of the Takara multiplex assay. Our results provide the first evidence that the diagnostic and analytical performances of the Liquid Dual-CoCoMo assay are better than those of commercially available multiplex assays that target the pol region.