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Journal of Clinical Microbiology, October 2002, p. 3666-3670, Vol. 40, No. 10
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.10.3666-3670.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Rapid and Sensitive Routine Detection of All Members of the Genus Enterovirus in Different Clinical Specimens by Real-Time PCR

Monique Nijhuis, Noortje van Maarseveen, Rob Schuurman, Sandra Verkuijlen, Machiel de Vos, Karin Hendriksen, and Anton M. van Loon*

Department of Virology, Eijkman Winkler Center, University Medical Center, Utrecht, The Netherlands

Received 8 May 2002/ Accepted 15 June 2002

We developed a rapid and sensitive method for the routine detection of all members of the enterovirus genus in different clinical specimens by using real-time TaqMan quantitative PCR. Multiple primer and probe sets were selected in the highly conserved 5'-untranslated region of the enterovirus genome. Our assay detected all 60 different enterovirus species tested, whereas no reactivity was observed with the viruses from the other genera of the picornaviridae family, e.g., hepatovirus and parechovirus. Weak cross-reactivity was observed with 7 of the 90 different high-titer rhinovirus stocks but not with rhinovirus-positive clinical isolates. Analysis of a well-characterized reference panel containing different enteroviruses at various concentrations demon-strated that the enterovirus real-time TaqMan PCR is as sensitive as most of the currently used molecular detection assays. Evaluation of clinical isolates demonstrated that the assay is more sensitive than the "gold standard" method, i.e., viral culture. Moreover, the PCR assay can be used on different clinical specimens, such as plasma, serum, nose and throat swabs, cerebrospinal fluid, and bronchoalveolar lavage, without apparent inhibition. Our data demonstrate that the real-time TaqMan PCR is a rapid and sensitive assay for the detection of enterovirus infection. The assay has a robust character and is easily standardized, which makes it an excellent alternative for the conventional time-consuming viral culture.

* Corresponding author. Mailing address: Eijkman-Winkler Center for Microbiology, Infectious Diseases and Inflammation, Department of Virology G04.614, University Medical Center Utrecht, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands. Phone: 31-30-2506526. Fax: 31-30-2505426. E-mail: a.m.vanloon{at}azu.nl.

Journal of Clinical Microbiology, October 2002, p. 3666-3670, Vol. 40, No. 10
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.10.3666-3670.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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