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Journal of Clinical Microbiology, January 2008, p. 164-170, Vol. 46, No. 1
0095-1137/08/$08.00+0     doi:10.1128/JCM.01316-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Novel Light-Upon-Extension Real-Time PCR Assays for Detection and Quantification of Genogroup I and II Noroviruses in Clinical Specimens{triangledown}

Johan Nordgren,1,4 Filemón Bucardo,2 Olaf Dienus,3 Lennart Svensson,1 and Per-Eric Lindgren4*

Division of Molecular Virology, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden,1 Department of Microbiology, University of León, León, Nicaragua,2 Microbiological Laboratory, Ryhov County Hospital, Jönköping, Sweden,3 Division of Medical Microbiology, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden4

Received 1 July 2007/ Returned for modification 31 August 2007/ Accepted 16 October 2007

Norovirus is now recognized as the leading cause of nonbacterial acute gastroenteritis in adults, causing numerous outbreaks worldwide. We have developed two novel light-upon-extension (LUX) real-time PCR assays for detection and quantification of norovirus genogroups I and II. The LUX system uses a fluorophore attached to one primer having a self-quenching hairpin structure, making it cost-effective and specific. The assays were evaluated against clinical stool specimens (n = 103) from Sweden and Nicaragua and compared to established methods. The norovirus assay detected more positive stool specimens (47/103) than conventional PCR (39/103) and corresponded to a TaqMan real-time PCR, with the exception of one specimen. Furthermore, the assays correctly identified all (n = 11) coded control specimens in a reference panel containing various genogroups and genotypes. Both LUX real-time PCR assays had a wide dynamic range, detecting from ≤101 to 107 genes per reaction, resulting in a theoretical lower limit of ≤~20 000 viruses per gram of stool. No cross-reactivity was noticed with specimens containing other enteric viruses, and by using melting curve analysis we could differentiate between norovirus genogroups I and II.

* Corresponding author. Mailing address: Department of Clinical and Experimental Medicine, Linköping University, 58185 Linköping, Sweden. Phone: 4613228586. Fax: 4613224789. E-mail: perli{at}imk.liu.se

{triangledown} Published ahead of print on 24 October 2007.

Journal of Clinical Microbiology, January 2008, p. 164-170, Vol. 46, No. 1
0095-1137/08/$08.00+0     doi:10.1128/JCM.01316-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Bucardo, F., Nordgren, J., Carlsson, B., Paniagua, M., Lindgren, P.-E., Espinoza, F., Svensson, L. (2008). Pediatric Norovirus Diarrhea in Nicaragua. J. Clin. Microbiol. 46: 2573-2580 [Abstract] [Full Text]  


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