Visualized detection of wild-type classical swine fever virus using RT-LAMP

To develop a rapid and practical method to differentiate wild-type strains of classical swine fever virus(CSFV) and the attenuated C-strain,a reverse transcription-loop-mediated isothermal amplification(RT-LAMP) assay was established with a set of primers based on NS5B gene sequence.The viral cDNA generated by reverse transcription was amplified with Bst DNA polymerase at a constant temperature of 62 ℃,and the products could be visualized under the UV light with SYBR Green I dye.The RT-LAMP was able to detect different genotypes of wild-type CSFV strains,but not for the C-strain,bovine viral diarrhea virus or other swine viruses.The detection limit of the RT-LAMP assay was 2.5 TCID50 of CSFV,comparable to the sensitivity of the real-time RT-PCR.The agreement rate between RT-LAMP and the real-time RT-PCR or the primer-probe energy transfer real-time PCR was 100 % or 98.4 % in detecting 126 samples.Thus,the assay is a rapid,sensitive,simple and practical method for the detection of wild-type CSFV in the field.