A novel RT-LAMP assay for rapid and simple detection of classical swine fever virus

Abstract

A simple and rapid assay for the detection of Classical swine fever virus (CSFV) was established using reverse transcription loop-mediated isothermal amplification (RT-LAMP). This study describes the amplification of the genomic RNA of CSFV under isothermal conditions (63 °C) within one hour, using a set of six primers (two outer primers, two inner primers and two loop primers). This RT-LAMP assay showed 100-fold higher sensitivity than the standard RT-PCR method and identified eighteen additional positive cases that were negative when tested by RT-PCR. This RT-LAMP was able to detect all the 13 strains of CSFV but not the BVDV. PRRSV. SIV. PRV-PCV, thus showed a good specificity. Products amplified by RT-LAMP can be visualized by agarose gel electrophoresis and in addition, either as a white precipitate at the bottom of the tube after a pulse spin or as a color change when dyed with SYBR Green I which are visible to the naked eye. Because RT-LAMP is low-cost and produces rapid results, it has the potential to be an excellent tool for CSFV surveillance in the field, especially in developing countries.