Multiplex quantitative PCR assay for porcine circoviruses 2 and 3 and its application for measuring infection rates at different stages of pig production.

Porcine circovirus 2 (PCV2) and porcine circovirus 3 (PCV3) are considered a threat to the pig industry due to their association with growth retardation and reproductive disorders. In this study, we developed a multiplex real-time PCR (qPCR) assay for simultaneous detection of PCV2 and PCV3 and used it to investigate the epidemiology of PCV2 and PCV3 at different stages of pig production. The sensitivity of the multiplex qPCR was 4.32 × 10¹copies/µL for PCV2 and 1.01 × 10¹copies/µL for PCV3, and the coefficient of variation was less than 1%. The correlation coefficients (R²) of the standard curves were all greater than 0.990. Out of 1241 samples tested, nine (0.73%) were positive for PCV2, 209 (16.84%) for PCV3, and three (0.24%) for both. PCV2 was detected in saliva from asymptomatic gilts on one farm, and PCV3 was detected in all sample types except semen at all production stages on all of the farms where samples were collected. The main sample types that tested positive were saliva (23.19%, 77/322), blood (17.80%, 102/573), saliva/blood mixture (18.75%, 9/48), and pigpen swabs (32.14%, 9/28). Viral loads ranged from 10^3.9 to 10^8.2 copies/mL. Gilts (37.85%, 81/214) and grow-finish pigs (42.25%, 30/71) were the main asymptomatic PCV3 carriers. Piglets (16.67%, 3/18) and nursery pigs (22.73%, 5/22) were the main symptomatic PCV3 carriers. The assay described here is a practical and sensitive diagnostic technique for identification and monitoring of PCV2 and PCV3 infections and can provide new information on the epidemiology of PCV2 and PCV3 that can be applied for developing effective prevention and control strategies.