Indian Journal of Animal Research

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Research Article

Indian Journal of Animal Research, volume 50 issue 6 (december 2016) : 979-982

Development and application of real-time TaqMan RT-PCR assay for improved detection of classical swine fever virus in slaughtered pigs 

M. Rout*, G. Saikumar1
1<p>Division of Pathology,&nbsp;Indian Veterinary Research Institute, Izatnagar - 243 122, India.</p>
Cite article:- Rout* M., Saikumar1 G. (2016). Development and application of real-time TaqMan RT-PCR assay for improved detection of classical swine fever virus in slaughtered pigs . Indian Journal of Animal Research. 50(6): 979-982. doi: 10.18805/ijar.v0iOF.6667.

ABSTRACT

Classical swine fever (CSF) is an economically devastating disease of pigs. Instrumental to the control of CSF is a well-characterized sensitive assay that can deliver a rapid and accurate diagnosis before the onset of clinical signs. With this objective, a real-time fluorogenic-probe hydrolysis (TaqMan) reverse transcription-polymerase chain reaction (RT-PCR) assay was developed for rapid and specific detection of classical swine fever virus (CSFV) and applied on samples derived from infected slaughtered pigs. A pair of PCR primers targeting 5’ -non-coding region (CSFL1 and CSFR1) in conjunction with a CSFV-specific fluorogenic probe (CSFP1) was designed and assessed in real-time PCR. During PCR, when the target of interest was present, the CSFV specific FAM-labeled TaqMan probe annealed to the amplicon between the forward and reverse primers and was subsequently cleaved via the 5¢-3¢  exonuclease activity of the DNA polymerase resulting in the release of the fluorescent reporter dye. This assay was found to be rapid and strain-specific for CSFV detection. 

REFERENCES


  1. Ciglenecki, U.J., Grom, J., Toplak, I., Jemersic, L. and Barlic-Maganja, D. (2008). Real-time RT-PCR assay for rapid and specific detection of classical swine fever virus: Comparison of SYBR Green and TaqMan MGB detection methods using novel MGB probes. J. Virol. Methods., 147: 257-264

  2. Clavijo, A., Zhou, E.M., Vydelingum, S. and Heckert. R. (1998). Development and evaluation of a novel antigen capture assay for the detection of classical swine fever virus antigens. Vet. Microbiol., 60: 155-168.

  3. De Smit, A.J., Eble, P.L., de Kluijver, E.P., Bloemraad, M. and Bouma. A. (1999). Laboratory decision-making during the classical swine fever epidemic of 1997-1998 in The Netherlands. Prev. Vet. Med., 42: 185-199. 

  4. Gava, D., Souza, C.K., Schaefer, R., Vincent, A.L., Cantao, M.E., Coldebella, A., and Ciacci-Zanella, J.R. (2015). A TaqMan-based real-time PCR for detection and quantification of porcine parvovirus 4. J. Virol. Methods., 219: 14-17. 

  5. Harding, M., Lutze-Wallace, C., Prud’Homme, I., Zhong, X. and Rola, J. (1994). Reverse transcriptase-PCR assay for detection of hog cholera virus. J. Clin. Microbiol., 32: 2600-2602.

  6. Hoffmann, B., Beer, M., Schelp, C., Schirrmeier, H. and Depner, K. (2005). Validation of a real-time RT-PCR assay for sensitive and specific detection of classical swine fever. J. Virol. Methods., 130: 36-44.

  7. Leifer, I., Hoffmann, B., Hoper, D., Rasmuseen, T.B., Blome, S., Strebelow, G., Horeth-Bontgen, D., Staubach, C. and Beer, M. (2010) Molecular epidemiology of current classical swine fever virus isolates of wild boar in Germany. J. Gen. Virol., 91: 2687-2697.

  8. Letellier, C. and Kerkhofs, P. (2003). Real-time PCR for simultaneous detection and genotyping of bovine viral diarrhea virus. J. Virol. Methods., 114: 21-27.

  9. Li, B., Xiao, S., Ma, J., Liu, Y., Mao, L., Wen, L., Mao, A., Zhang, X., Ni, Y., Guo, R., Zhou, J., Yu, Z., Lv, L., Wang, X., Fang, L., Chen, H. and He, K. (2011). Development of a novel TaqMan-based real-time PCR assay for the detection of porcine boca-like virus (Pbo-likeV). Virol J., 8: 357. 

  10. Li, Y., Zhou, F., Li, X., Wang, J., Zhao, X. and Huang, J. (2013). Development of TaqMan-based qPCR method for detection of caprine arthritis-encephalitis virus (CAEV) infection. Arch. Virol., 158: 2135-2141. 

  11. McGoldrick, A., Bensaude, E., Ibata, G., Sharp, G. and Paton, D.J. (1999). Closed one-tube reverse transcription nested polymerase chain reaction for the detection of pestiviral RNA with fluorescent probes. J. Virol. Methods., 79: 85-95.

  12. McGoldrick, A., Lowings, J.P., Ibata, G., Sands, J.J., Belak, S. and Paton, D.J. (1998). A novel approach to the detection of classical swine fever virus by RT-PCR with a fluorogenic probe (TaqMan). J. Virol. Methods., 72: 125-135.

  13. Moennig, V. and Plagemann, P.G. (1992). The pestiviruses. Adv. Virus Res., 41: 53-98.

  14. Moennig, V., Floegel-Niesmann, G. and Greiser-Wilke, I. (2003). Clinical signs and epidemiology of classical swine fever: A review of new knowledge. Vet. J., 165: 11-20.

  15. Ressang, A.A. and de Boer, G.F. (1968). A comparison between the cell culture, frozen tissue section, impression and mucosal smear techniques for fluorescent antibody in the diagnosis of hog cholera. Neth. J. Vet. Sci., 1: 72.

  16. Shannon, A.D., Morrissy, C. Mackintosh, S.G. and Westbury. H.A. (1993). Detection of hog cholera virus antigens in experimentally infected pigs using an antigen-capture ELISA. Vet. Microbiol., 34: 233-248.

  17. Song, C., Zhu, C., Zhang, C. and Cui, S. (2010). Detection of porcine parvovirus using a TaqMan-based real-time PCR with primers and probe designed for the NS1 gene. Virol J., 7: 353.

  18. Terpstra, C. (1991). Hog cholera: an update of present knowledge. Br. Vet. J., 147: 397-406.

  19. Van Oirschot, J.T. (1999). Hog Cholera/Classical swine fever. In: Disease of swine, 5th edn, edited by B. E. Shaw, Ames, Iowa State University Press, pp. 289-300.

  20. Wirtz, B., Tratchin, J.D., Muller, H.K. and Mitchell, D.B. (1993). Detection of hog cholera virus and differentiation from other pestiviruses by polymerase chain reaction. J. Clin. Microbiol., 31: 1148-1154.

  21. Zhao, K., Han, F., Zou, Y., Zhu, L., Li, C., Xu, Y., Zhang, C., Tan, F., Wang, J., Tao, S., He, X., Zhou, Z. and Tang, X. (2010). Rapid detection of porcine circovirus type 2 using a TaqMan-based real-time PCR. Virol J., 7: 374. 
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