A multiplex nested RT-PCR for the detection and differentiation of wild-type viruses from C-strain vaccine of classical swine fever virus

作者：Li Yan; Zhao Jian Jun; Li Na; Shi Zixue; Cheng Dan; Zhu Qing Hu; Tu Changchun; Tong Guang Zhi; Qiu Hua Ji^*

来源：Journal of Virological Methods, 2007, 143(1): 16-22.

DOI：10.1016/j.jviromet.2007.01.032

• classical swine fever virus
• nested RT-PCR
• restrictive fragment length polyniorphism
• differential diagnosis

A multiplex nested RT-PCR (RT-nPCR) was developed for the detection and differentiation of classical swine fever virus (CSFV). A fragment of 447 or 343 bp was amplified from the genomic RNA of C-strain or virulent Shimen strain, respectively, and two fragments of 447 and 343 bp were simultaneously amplified from the mixed samples of C-strain and Shimen. When detecting several wild-type isolates representative of different subgroups (1.1, 2.1, 2.2, and 2.3) circulating in Mainland China and samples from pigs experimentally infected with Shimen strain, the RT-nPCR resulted in an amplification pattern similar to Shimen. No amplification was achieved for uninfected cells, or cells infected with bovine viral diarrhea virus (BVDV), and other viruses of porcine origin. The RT-nPCR was able to detect as little as 0.04 pg of CSFV RNA. The restrictive fragment length polymorphism (RFLP) demonstrated unique patterns of wild-type viruses and C-strain. Among the 133 field samples, 42 were tested to contain wild-type viruses and 18 showing presence of C-strain. The RT-nPCR can be used to detect and differentiate pigs infected with wild-type CSFV from those vaccinated with C-strain vaccine, thus minimizing the risk of culling vaccinates during outbreaks.

• 出版日期2007-7
• 单位中国农业科学院哈尔滨兽医研究所; 中国农业科学院特产研究所; 中国人民解放军军事医学科学院