Detection of human orthopoxvirus infections and differentiation of smallpox virus with real-time PCR
Article first published online: 21 NOV 2008
Copyright © 2008 Wiley-Liss, Inc.
Journal of Medical Virology
Volume 81, Issue 1, pages 146–152, January 2009
How to Cite
Putkuri, N., Piiparinen, H., Vaheri, A. and Vapalahti, O. (2009), Detection of human orthopoxvirus infections and differentiation of smallpox virus with real-time PCR. J. Med. Virol., 81: 146–152. doi: 10.1002/jmv.21385
- Issue published online: 21 NOV 2008
- Article first published online: 21 NOV 2008
- Manuscript Accepted: 22 SEP 2008
- Finnish Scientific Advisory Board for Defence (MATINE)
- Hospital District of Helsinki and Uusimaa. Grant Numbers: EVO TYH4211, TYH6215
- real-time PCR;
- cowpox virus
We developed a real-time PCR protocol to detect orthopoxviruses (OPVs) from different clinical specimens and to separate variola virus from other OPVs. In our protocol, we used automated nucleic acid extraction system together with real-time PCR to create a simple, safe and fast procedure to obtain an initial result. The sensitivity was better by using designed hybridization probes as compared to SYBR green I for detection. The detection limit ranged from 13 to 1,300 copies per 20 µl reaction volume depending on the sample type. The PCR detected all OPVs pathogenic to human (variola, cowpox, monkeypox, vaccinia) as well as camelpox and ectromelia viruses. Amplification of variola virus sequences could be distinguished from other OPVs by melting curve analysis. We also demonstrated the applicability of the assay in human cases of cowpox and vaccinia virus infections. J. Med. Virol. 81:146–152, 2009. © 2008 Wiley-Liss, Inc.