Development of a droplet digital polymerase chain reaction for sensitive and simultaneous identification of porcine circovirus type 2 and 3.

In this study, a sensitive assay for simultaneous detection of porcine circovirus type 2 (PCV2) and type 3 (PCV3) was established using droplet digital PCR (ddPCR). Pairs of primers and probes were specifically designed to amplify PCV2 and PCV3. Specificity of the assay was confirmed by the failure of amplification of DNA of other relevant viruses. The detection limit for ddPCR was 2 copies/μL for PCV2 and 1 copy/μL for PCV3, a 3- and 10-fold greater sensitivity than TaqMan real-time PCR, respectively. Both methods showed a high degree of linearity, although TaqMan real-time PCR showed less sensitivity than ddPCR for clinical detection. In summary, the results demonstrated that the established ddPCR is an effective alternative method for the precise simultaneous quantification of PCV2 and PCV3 in clinical samples, especially in detecting lower concentrations of co-infected samples.