Comparative evaluation of RT-PCR, nucleic acid sequence-based amplification (NASBA) and real-time RT-PCR for detection of noroviruses in faecal material

Journal of Virological Methods
Alain HoudeYvon-Louis Trottier

Abstract

Using different primer and probe sets, RT-PCR, NASBA and TaqMan RT-PCR molecular methods were compared to detect NoV GII in 13 clinical stool samples. The RT-PCR results observed by gel electrophoresis (Ando, Kageyama and Anderson amplification and probe systems), dot blot hybridization (Ando and Kageyama) and real-time TaqMan assay (Ando and Kageyama) were shown to be consistent and reproducible for the detection of NoV GII. Whereas, the NASBA assay using Ando primers showed some reproducibility discrepancies. Detection limits of the NoV GII/Kageyama system were found to be equal or significantly higher than the Ando system. Real-time TaqMan RT-PCR assay showed similar detection limits to that of NASBA with the Kageyama amplification and detection system, while it was 1log less sensitive than the Ando system. In a clinical context, RT-PCR, NASBA and real-time TaqMan RT-PCR methods using undiluted samples were all suitable for the detection of NoV GII, however the NASBA assay provided less consistent signals. The NoV GII Kageyama real-time TaqMan RT-PCR assay was reliable with a high analytical sensitivity and has shown the capability of detecting one genomic equivalent copy.

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Citations

Aug 28, 2010·The Journal of Microbiology·Heetae LeeGwangPyo Ko
May 16, 2009·BMC Infectious Diseases·Gemma PhillipsJim Gray
Sep 13, 2012·Sensors·Chia-Chen ChangChii-Wann Lin
Feb 16, 2013·Food and Environmental Virology·Ambroos StalsMieke Uyttendaele
Oct 22, 2011·International Journal of Food Microbiology·L BaertM Uyttendaele
Nov 13, 2010·Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology·Kirsten MattisonTim F Booth
Apr 28, 2009·International Journal of Food Microbiology·Kirsten MattisonYvon-Louis Trottier
Mar 26, 2009·Journal of Microbiological Methods·Ambroos StalsEls Van Coillie
Dec 13, 2006·Journal of Virological Methods·Alain HoudeYvon-Louis Trottier
Jan 21, 2015·Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases·Qing-Bin LuWu-Chun Cao
Aug 21, 2012·Critical Reviews in Microbiology·Angus KnightLee-Ann Jaykus
Jan 9, 2015·Clinical Microbiology Reviews·Elizabeth RobilottiBenjamin A Pinsky
Jun 24, 2016·The Open Microbiology Journal·Haifeng Chen, Yuan Hu
Mar 3, 2021·Food and Environmental Virology·Philippe RaymondLouise Deschênes

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