Real-time PCR for quantitation of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2 in naturally-infected and challenged pigs.
Real-time PCR assays were developed for quantitative detection of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2). The established real-time PCR for the quantitation of PRRSV cDNA and PCV2 DNA were found to be in the 9-log(10) linear dynamic range with excellent linearity and reliable reproducibility. Using these techniques, the distribution and quantitation of PRRSV and PCV2 in naturally infected and challenged pigs were investigated. The viral concentrations were expressed as the mean log(10) viral DNA or cDNA copy numbers per mg or ml of tested samples. For pigs infected naturally with both viruses, the lung, spleen, tonsil and lymphoid organs had the highest viral burdens with ranges from 5.73 to 8.38 and 5.65 to 6.91 for PRRSV and PCV2, respectively. The injection of formalin-inactivated Salmonella choleraesuis emulsified in complete Freund's adjuvant 1 week before and after the inoculation of both viruses resulted in PRRSV replication enhancement 2 weeks post-challenge. However, this facilitated the clearance of PRRSV 4 weeks post-challenge. Results from this study show that the established quantitative PCR could be a useful tool when applied to vaccine development and pathogenesis studies in the future.
Chung
Wen-Bin
WB
Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung 912, ROC, Taiwan. wbchung@mail.npust.edu.tw
Chan
Wen-Hung
WH
Chaung
Hso-Chi
HC
Lien
Yi
Y
Wu
Chia-Chang
CC
Huang
Yu-Liang
YL
D000818:Animals; D018173:Circoviridae Infections / Q000662:veterinary* / Q000821:virology; D017925:Circovirus / Q000235:genetics / Q000302:isolation & purification*; D004279:DNA, Viral / Q000032:analysis; D016133:Polymerase Chain Reaction / Q000379:methods*; D019318:Porcine Reproductive and Respiratory Syndrome / Q000821:virology*; D019316:Porcine respiratory and reproductive syndrome virus / Q000235:genetics / Q000302:isolation & purification*; D015203:Reproducibility of Results; D013552:Swine / Q000821:virology; D013553:Swine Diseases / Q000821:virology*
D004279:DNA, Viral