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Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus

The most prevalent viruses currently causing diarrhea are norovirus and rotavirus, and rapid and sensitive detection methods are essential for the early diagnosis of disease. The purpose of this study was to establish a sensitive single‐tube two‐stage nucleic acid amplification method—reverse transc...

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Autores principales: Li, Feng‐yu, Guo, Ying‐hui, Sun, Zhen‐lu, Liu, Hong, Zhao, Meng‐chuan, Cui, Jia, Jiang, Yue, Shen, Xin‐xin, Ma, Xue‐jun, Feng, Zhi‐shan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10098066/
https://www.ncbi.nlm.nih.gov/pubmed/36916770
http://dx.doi.org/10.1002/jcla.24858
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author Li, Feng‐yu
Guo, Ying‐hui
Sun, Zhen‐lu
Liu, Hong
Zhao, Meng‐chuan
Cui, Jia
Jiang, Yue
Shen, Xin‐xin
Ma, Xue‐jun
Feng, Zhi‐shan
author_facet Li, Feng‐yu
Guo, Ying‐hui
Sun, Zhen‐lu
Liu, Hong
Zhao, Meng‐chuan
Cui, Jia
Jiang, Yue
Shen, Xin‐xin
Ma, Xue‐jun
Feng, Zhi‐shan
author_sort Li, Feng‐yu
collection PubMed
description The most prevalent viruses currently causing diarrhea are norovirus and rotavirus, and rapid and sensitive detection methods are essential for the early diagnosis of disease. The purpose of this study was to establish a sensitive single‐tube two‐stage nucleic acid amplification method—reverse transcription recombinase‐assisted PCR (RT‐RAP)—for simultaneous detection of norovirus GII and group A Rotavirus, with the first stage consisting of isothermal reverse transcription recombinase‐aided amplification (RT‐RAA) and the second stage consisting of qPCR (quantitative PCR). RT‐RAP is more sensitive than either RT‐RAA or qRT‐PCR (quantitative RT‐PCR) alone. And the addition of a barrier that can be disassembled after heating enabled the detection of samples within 1 h in a single closed tube. Sensitivity was 10 copies/reaction of norovirus (Novs) GII and group A rotavirus (RVA). In parallel, two hundred fecal specimens were used to evaluate the method and compare it with a commercial fluorescent quantitative RT‐PCR. The data showed kappa values of 0.957 and 0.98 (p < 0.05) for detecting Novs GII and RVA by the two methods, indicating the potential of the newly established assay to be applied to clinical and laboratory testing.
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spelling pubmed-100980662023-04-14 Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus Li, Feng‐yu Guo, Ying‐hui Sun, Zhen‐lu Liu, Hong Zhao, Meng‐chuan Cui, Jia Jiang, Yue Shen, Xin‐xin Ma, Xue‐jun Feng, Zhi‐shan J Clin Lab Anal Research Articles The most prevalent viruses currently causing diarrhea are norovirus and rotavirus, and rapid and sensitive detection methods are essential for the early diagnosis of disease. The purpose of this study was to establish a sensitive single‐tube two‐stage nucleic acid amplification method—reverse transcription recombinase‐assisted PCR (RT‐RAP)—for simultaneous detection of norovirus GII and group A Rotavirus, with the first stage consisting of isothermal reverse transcription recombinase‐aided amplification (RT‐RAA) and the second stage consisting of qPCR (quantitative PCR). RT‐RAP is more sensitive than either RT‐RAA or qRT‐PCR (quantitative RT‐PCR) alone. And the addition of a barrier that can be disassembled after heating enabled the detection of samples within 1 h in a single closed tube. Sensitivity was 10 copies/reaction of norovirus (Novs) GII and group A rotavirus (RVA). In parallel, two hundred fecal specimens were used to evaluate the method and compare it with a commercial fluorescent quantitative RT‐PCR. The data showed kappa values of 0.957 and 0.98 (p < 0.05) for detecting Novs GII and RVA by the two methods, indicating the potential of the newly established assay to be applied to clinical and laboratory testing. John Wiley and Sons Inc. 2023-03-14 /pmc/articles/PMC10098066/ /pubmed/36916770 http://dx.doi.org/10.1002/jcla.24858 Text en © 2023 The Authors. Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC. https://creativecommons.org/licenses/by-nc/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ (https://creativecommons.org/licenses/by-nc/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
spellingShingle Research Articles
Li, Feng‐yu
Guo, Ying‐hui
Sun, Zhen‐lu
Liu, Hong
Zhao, Meng‐chuan
Cui, Jia
Jiang, Yue
Shen, Xin‐xin
Ma, Xue‐jun
Feng, Zhi‐shan
Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title_full Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title_fullStr Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title_full_unstemmed Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title_short Rapid two‐stage amplification in a single tube for simultaneous detection of norovirus GII and group a rotavirus
title_sort rapid two‐stage amplification in a single tube for simultaneous detection of norovirus gii and group a rotavirus
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10098066/
https://www.ncbi.nlm.nih.gov/pubmed/36916770
http://dx.doi.org/10.1002/jcla.24858
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