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

被引:47
作者
Li, Yan
Zhao, Jian-Jun
Li, Na
Shi, Zixue
Cheng, Dan
Zhu, Qing-Hu
Tu, Changchun
Tong, Guang-Zhi
Qiu, Hua-Ji
机构
[1] Acad Mil Med Sci, Inst Vet Sci, Changchun 130062, Peoples R China
[2] Chinese Acad Agr Sci, Inst Special Econ Anim & Plant Sci, Dept Vet Prevent Med, Jilin 123109, Peoples R China
关键词
classical swine fever virus; nested RT-PCR; restrictive fragment length polyniorphism; differential diagnosis;
D O I
10.1016/j.jviromet.2007.01.032
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
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. (C) 2007 Elsevier B.V. All rights reserved.
引用
收藏
页码:16 / 22
页数:7
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