Reverse transcription and cDNA amplification by the polymerase chain reaction of equine arteritis virus (EAV)

A technique is described for the amplification and specific identification of equine arteritis virus (EAV) nucleotide sequences. The polymerase chain reaction (PCR) was evaluated initially by amplification of cloned virus specific cDNA sequences prior to amplification of single-stranded (ss) cDNA produced by reverse transcription (RT) of viral genomic RNA. Three separate primer pairs were used for RT/PCR of EAV genomic RNA, each pair producing only one band in agarose gels of the predicted size from the genomic nucleotide sequence. The viral origin of cDNA products was confirmed by hybridisation analysis with EAV-specific probes. RT/PCR analysis of clinical material indicates the methodology is sensitive enough to detect 600 pfu/ml EAV in seminal plasma.