Arch Virol (1994) 135:89-99 _Archives Virology © Springer-Verlag 1994 Printed in Austria Direct detection of the porcine reproductive and respiratory syndrome (PRRS) virus by reverse polymerase chain reaction (RT- PCR) P. Sufirez 1, R. Zardoya z, C. Prieto ~, A. Solana 1, E. Tabar6s 3, J. M. Bautista 2, and J. M. Castro ~ 1Departamento de Patologia Animal I and 2Departamento de Bioquimica y Biologia Molecular IV, Universidad Complutense de Madrid, Facultad de Veterinaria, Madrid, and 3Departamento de Microbiologia, Univergidad Autonoma de Madrid, Facultad de Medicina, Madrid, Spain Accepted October 18, 1993 Summary. A method for direct detection of the porcine reproductive and respiratory syndrome (PRRS) virus was developed, based on reverse transcrip- tion of the viral RNA coupled to DNA amplification by polymerase chain reaction. A set of primers was designed from Lelystad virus sequence within ORF 7 encoding nucleocapsid protein. From seven Spanish field isolated strains the 312bp amplified fragment was cloned and sequenced. Alignment with Lelystad virus sequence revealed a 96 97~o homology. A maximum sensitivity of 6.7 TCIDso was achieved with the reported procedure in experimentally infected swine alveolar macrophages cultures. The sensitivity obtained in crude clinical samples from experimentally infected 3-weeks old pigs was approxi- mately 102 TCIDso. High specificity for the PRRS virus was demonstrated for the method, as none of the seven common swine virus assayed rendered DNA amplification product. Introduction What was initially referred to as "mystery swine disease", which causes reproductive failures in sows and respiratory disorders in piglets [9], was first reported in North America in 1987 [8]. The etiological agent was first isolated in 1990 in Europe [16] and designated Lelystad virus (LV), and then in 1992 in the U.S.A. [4], being both virus antigenically distinct but structurally related [17]. This disease is also known by other different names, such as "swine infertility and respiratory syndrome" (SIRS) and "porcine reproductive and respiratory syndrome" (PRRS). It has been reported that the causative viral agent of PRRS preferentially replicates in vitro in primary cultures of swine alveolar lung macrophages (SAM) [2]. Electron microscopic studies have shown that the PRRS virus is