Development of TaqMan-based real-time PCR assay based on the E1 genefor the quantitative detection of the Getah virus.

ABSTRACT

To develop a sensitive, specific, and rapid approach for the detection Getah virus (GETV), a set of primers targeting the conserved region of the E1 gene was created. The TaqMan-based real-time PCR method for GETV detection was developed by optimizing the reaction conditions. The method demonstrated excellent specificity, and amplification did not occur with the causative agents of all prevalent swine viral infections (CSFV, PRRSV, PRV, PEDV, PTV, and JEV), except GETV. Additionally, upon assessing the sensitivity of the method, the minimum detection limit for GETV was found to be 5.94 copies/µL, which is 10 times higher than that of the traditional PCR approach. Further, the intra- and inter-assay variation coefficients were less than 1%, demonstrating good repeatability. Moreover, GETV was found in 10 of the 20 field serum samples using real-time PCR but only in three of the samples using traditional PCR. Consequently, the first GETV TaqMan-based real-time PCR approach based on the E1 gene was developed for GETV pathogenic diagnoses, and this exhibited high specificity, sensitivity, and repeatability. This assay is practical for the pathogenic diagnosis and epidemiology of GETV.