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Journal of Clinical Microbiology, June 2009, p. 1863-1870, Vol. 47, No. 6
0095-1137/09/$08.00+0     doi:10.1128/JCM.02226-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Typing of Enteroviruses by Use of Microwell Oligonucleotide Arrays{triangledown} ,{dagger}

P. Susi,1 L. Hattara,2 M. Waris,1 T. Luoma-aho,2 H. Siitari,2 T. Hyypiä,1 and P. Saviranta2*

Department of Virology, University of Turku, Kiinamyllynkatu 13,1 Medical Biotechnology Centre, VTT Technical Research Centre of Finland, Itäinen Pitkäkatu 4C, 20520 Turku, Finland2

Received 19 November 2008/ Returned for modification 15 January 2009/ Accepted 27 March 2009

We have developed a straightforward assay for the rapid typing of enteroviruses using oligonucleotide arrays in microtiter wells. The viral nucleic acids are concomitantly amplified and labeled during reverse transcription-PCR, and unpurified PCR products are used for hybridization. DNA strands are separated by alkaline denaturation, and hybridization is started by neutralization. The microarray hybridization reactions and the subsequent washes are performed in standard 96-well microtiter plates, which makes the method easily adaptable to high-throughput analysis. We describe here the assay principle and its potential in clinical laboratory use by correctly identifying 10 different enterovirus reference strains. Furthermore, we explore the detection of unknown sequence variants using serotype consensus oligonucleotide probes. With just two consensus probes for the coxsackievirus A9 (CVA9) serotype, we detected 23 out of 25 highly diverse CVA9 isolates. Overall, the assay involves several features aiming at ease of performance, robustness, and applicability to large-scale studies.

* Corresponding author. Mailing address: VTT Technical Research Centre of Finland, Medical Biotechnology, Itäinen Pitkäkatu 4C, FIN-20520 Turku, Finland. Phone: 358-20-722 2831. Fax: 358-20-722 2840. E-mail: petri.saviranta{at}vtt.fi

{triangledown} Published ahead of print on 8 April 2009.

{dagger} Supplemental material for this article may be found at http://jcm.asm.org/.

Journal of Clinical Microbiology, June 2009, p. 1863-1870, Vol. 47, No. 6
0095-1137/09/$08.00+0     doi:10.1128/JCM.02226-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Zhou, F., Kong, F., McPhie, K., Ratnamohan, M., Donovan, L., Zeng, F., Gilbert, G. L., Dwyer, D. E. (2009). Identification of 20 Common Human Enterovirus Serotypes by Use of a Reverse Transcription-PCR-Based Reverse Line Blot Hybridization Assay. J. Clin. Microbiol. 47: 2737-2743 [Abstract] [Full Text]  


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