Rapid detection of porcine reproductive and respiratory syndrome viral nucleic acid in blood using a fluorimeter based PCR method

Porcine reproductive and respiratory syndrome virus (PRRSV) is an Arterivirus recognised world wide as an important cause of reproductive failure and pneumonia in pigs. American and European strains of PRRSV, differentiated antigenically and genomically, have been reported. PRRSV infections are currently diagnosed using serology, virus isolation and/or immunocytochemistry. In order to overcome various drawbacks associated with these techniques, conventional, block-based RT-PCR methods for the detection of PRRSV nucleic acid in clinical samples have been described. These methods require gel electrophoresis for analysis of PCR products and present high risk of DNA carry-over contamination between the samples tested. We describe the detection of PRRSV RNA in serum samples and in blood impregnated filter disks (FDs), obtained from experimentally inoculated pigs, using a closed-tube, fluorimeter-based PCR assay. The assay eliminates the use of gel electrophoresis, and is as sensitive and specific as the conventional block-based PCR assay, detecting positive samples as early as 1 day post-inoculation. We also report a rapid fluorimeter based PCR method for differentiating American and European strains of PRRSV. (C) 2000 Elsevier Science B.V. All rights reserved.