Rapid and Simple Detection of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) by Reverse Transcription Loop-Mediated Isothermal Amplification

Given the economic consequences of infection with porcine reproductive and respiratory syndrome virus (PRRSV) for the global swine industry a rapid and practical early detection assay is required. This study established a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay to detect PRRSV. Four primers were designed targeting the ORF5 gene of PRRSV were designed. Reverse transcription and amplification of the viral RNA using AMV reverse transcriptase was optimal at a constant temperature of 65 degrees C. The output of the RT-LAMP assay was visualized using 1% agarose gel electrophoresis or color change after the addition of the SYBR Green I dye. The RT-LAMP method was approximately 100-fold more sensitive than RT-PCR for PRRSV detection. The assay was also specific for PRRSV and did not cross react with CSFV, PCV-2, PPV or PRV. In clinical samples (n=5) RT-PCR and RT-LAMP both identified the same 2 PRRSV infected samples. Thus, the novel RT-LAMP assay described here is a rapid, simple, sensitive, specific test for PRRSV that can potentially be applied in clinical settings.