Evaluation of a multiplex real-time RT-PCR for quantitative and differential detection of wild-type viruses and C-strain vaccine of Classical swine fever virus

被引：106

作者：

Zhao, Jian-Jun ^{[1
]}

Cheng, Dan ^{[1
,3
]}

Li, Na ^{[1
]}

Sun, Yuan ^{[1
]}

Shi, Zixue ^{[2
]}

Zhu, Qing-Hu ^{[1
]}

Tu, Changchun ^{[2
]}

Tong, Guang-Zhi ^{[1
]}

Qiu, Hua-Ji ^{[1
]}

机构：

[1] Chinese Acad Agr Sci, Harbin Vet Res Inst, Natl Key Lab Vet Biotechnol, Div Swine Infect Dis, Harbin 150001, Heilongjiang, Peoples R China

[2] Acad Mil Med Sci, Inst Vet Sci, Changchun 130062, Peoples R China

[3] Xinjiang Agr Univ, Coll Vet Med, Urumqi 830052, Peoples R China

来源：

VETERINARY MICROBIOLOGY | 2008年 / 126卷 / 1-3期

关键词：

Classical swine fever virus; TaqMan real-time RT-PCR; multiplex PCR; differential diagnosis; C-strain vaccine;

D O I：

10.1016/j.vetmic.2007.04.046

中图分类号：

Q93 [微生物学];

学科分类号：

071005 ; 100705 ;

摘要：

Classical swine fever virus (CSFV) is the causative agent of classical swine fever (CSF), one of OIE listed diseases. Most of the currently available detection methods do not allow discrimination between wild-type CSF viruses and the vaccine strains. This study was designed to develop a multiplex real-time RT-PCR for the quantitative and differential detection of wild-type viruses and C-strain vaccine widely used in China. CSFV specific primers and two differently labeled TaqMan probes for the differentiation of wild-type viruses from C-strain vaccine were designed in the 5'-untranslated region of the viral genome of CSFV The two TaqMan probes specifically hybridize wild-type viruses of different subgroups and C-strain vaccine, respectively, in the multiplex real-time RT-PCR, with no cross-reaction to a number of non-CSFV porcine viruses. The sensitivity of the assay for detecting wild-type and C-strain-type vaccine viruses was determined to be 41.8 and 81.5 copies/mu L viral RNA, respectively. Completely correct differentiation of wild-type viruses from C-strain vaccine was achieved when testing reference strains and characterized field isolates of CSFV in China. The multiplex real-time RT-PCR was able to detect the viral RNA in the whole blood samples of experimentally infected pigs as early as 2 days post-infection, 3 to 4 days prior to the onset of clinical signs in co-housed pigs. The agreements between the multiplex real-time RT-PCR and a multiplex RT-nested PCR for detection of wild-type and C-strain-type viruses were 96.9% and 100%, respectively, when detecting 106 different field samples. There is a positive correlation between the titers of C-strain vaccines titrated in rabbits and RNA copies quantitated by the multiplex real-time RT PCR. The novel assay described here is rapid and sensitive, and is useful for differentiating field strains and C-strain of CSFV in China. (C) 2007 Elsevier B.V. All rights reserved.

引用

页码：1 / 10

页数：10

共 31 条

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