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Journal of Clinical Microbiology, September 2009, p. 2855-2862, Vol. 47, No. 9
0095-1137/09/$08.00+0     doi:10.1128/JCM.00448-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

New Real-Time PCR Detects Prolonged Norovirus Excretion in Highly Immunosuppressed Patients and Children {triangledown}

C. Henke-Gendo,1* G. Harste,1 B. Juergens-Saathoff,1 F. Mattner,2 H. Deppe,1 and A. Heim1

Institute of Virology, Medical School Hannover, Hannover,1 Institute of Medical Microbiology and Hygiene, Universitätsklinikum Schleswig-Holstein, Campus Lübeck, Lübeck, Germany2

Received 3 March 2009/ Returned for modification 23 April 2009/ Accepted 14 July 2009

Noroviruses (NoV) are a major cause of epidemic nonbacterial gastroenteritis and affect all age groups worldwide. Three of five NoV genogroups, namely, genogroup I (GI), GII, and GIV, are associated with human disease. Unfortunately, these genogroups demonstrate a high degree of sequence diversity, complicating the design of pan-NoV diagnostic PCR tests. To decrease the risk of false-negative test results, we have developed a new one-step real-time TaqMan reverse transcription-PCR protocol. This protocol detects all human NoV genogroups in one reaction with a sensitivity of 400 virus genome equivalents/reaction for both GI and GII. The use of in vitro-transcribed NoV RNA as an external standard allows (semi)quantification of viral loads in samples. In a retrospective analysis of 206 stool samples from 77 patient episodes, the duration of NoV excretion and the amount of virus excreted were determined. Twenty (26.0%) of these episodes lasted longer than 10 days. Univariate risk factor analysis revealed the patient status after organ transplantation (odds ratio [OR], 7.49 [95% confidence interval, 2.06 to 28.32]; P < 0.001), immunosuppression (OR, 9.19 [95% confidence interval, 2.50 to 35.39]; P < 0.001), and age of less than 10 years (OR, 4.58 [95% confidence interval, 1.36 to 15.77]; P = 0.004) as risk factors for a NoV excretion period of more than 10 days. These findings were confirmed by time-dependent Kaplan-Meier analyses, whereas multivariate Cox regression analyses identified immunosuppression as the sole risk factor. Surprisingly, in contrast to the excretion periods, the viral loads in stools did not increase in connection with age or immunosuppressive status. This fact may be one important piece in the pattern of high-level NoV transmissibility and may have an impact on the development of transmission prevention strategies.

* Corresponding author. Mailing address: Institute of Virology, Medical School Hannover, Carl-Neuberg-Str. 1, D-30625 Hannover, Germany. Phone: 49 511 532-4310. Fax: 49 511 532-8736. E-mail: Henke-Gendo.Cornelia{at}mh-hannover.de

{triangledown} Published ahead of print on 22 July 2009.

Journal of Clinical Microbiology, September 2009, p. 2855-2862, Vol. 47, No. 9
0095-1137/09/$08.00+0     doi:10.1128/JCM.00448-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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