2017 ; 27(4):
|Author||Ju Eun Yoo, Cheonghoon Lee, SungJun Park, GwangPyo Ko|
|Affiliation||Department of Environmental Health Sciences, Graduate School of Public Health,|
|Title||Evaluation of Various Real-Time Reverse Transcription Quantitative PCR Assays for Norovirus Detection|
J. Microbiol. Biotechnol.2017 ; 27(4):
|Abstract||Human noroviruses are widespread and contagious viruses causing nonbacterial
gastroenteritis. Real-time reverse transcription quantitative PCR (real-time RT-qPCR) is
currently the gold standard for the sensitive and accurate detection of these pathogens and
serves as a critical tool in outbreak prevention and control. Different surveillance teams,
however, may use different assays, and variability in specimen conditions may lead to
disagreement in results. Furthermore, the norovirus genome is highly variable and
continuously evolving. These issues necessitate the re-examination of the real-time RT-qPCR’s
robustness in the context of accurate detection as well as the investigation of practical
strategies to enhance assay performance. Four widely referenced real-time RT-qPCR assays
(Assays A-D) were simultaneously performed to evaluate characteristics such as PCR
efficiency, detection limit, and sensitivity and specificity with RT-PCR, and to assess the most
accurate method for detecting norovirus genogroups I and II. Overall, Assay D was evaluated
to be the most precise and accurate assay in this study. A ZEN internal quencher, which
decreases nonspecific fluorescence during the PCR, was added to Assay D’s probe, which
further improved the assay performance. This study compared several detection assays for
noroviruses, and an improvement strategy based on such comparisons provided useful
characterizations of a highly optimized real-time RT-qPCR assay for norovirus detection.|
|Keywords||Norovirus, RT-qPCR, detection limit, sensitivity, internal quencher|
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