COMPARISON OF 4 RNA EXTRACTION METHODS FOR THE DETECTION OF PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS BY RT-PCR

We compared four commercial RNA isolation methods for the extraction o f RNA from porcine reproductive and respiratory syndrome virus (PRRSV) . The sensitivity of the methods was determined by extraction of RNA f rom serial 10-fold dilutions of PRRSV diluted in PRRSV-negative porcin e serum or in Eagle's minimal essential medium (EMEM), followed by amp lification of extracted nucleic acids by RT-PCR. The PCR products were detected in ethidium bromide-stained agarose gels. The QIAamp viral k it, which is based on binding of RNA to silica particles, was the most sensitive, allowing the detection of an equivalent of 10 TCID50 of vi rus in 100 mu l of serum or EMEM. The QIAamp-tissue and the TRIzol LS kits detected 100 TCID50 per 100 mu l in both diluents whereas Ultra-s pec-3 detected 100 TCID50 of virus in EMEM and 1000 TCID50 in serum. T hese results indicate that QIAamp viral kit has a higher sensitivity f or RT-PCR analysis of PRRSV.