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Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens

Human enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD) worldwide and has been associated with neurological complications which resulted in fatalities during recent outbreak in Asia pacific region. A direct reverse transcription loop-mediated isothermal ampli...

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Autores principales: Nie, Kai, Qi, Shun-xiang, Zhang, Yong, Luo, Le, Xie, Yun, Yang, Meng-jie, Zhang, Yi, Li, Jin, Shen, Hongwei, Li, Qi, Ma, Xue-jun
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3525532/
https://www.ncbi.nlm.nih.gov/pubmed/23272248
http://dx.doi.org/10.1371/journal.pone.0052486
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author Nie, Kai
Qi, Shun-xiang
Zhang, Yong
Luo, Le
Xie, Yun
Yang, Meng-jie
Zhang, Yi
Li, Jin
Shen, Hongwei
Li, Qi
Ma, Xue-jun
author_facet Nie, Kai
Qi, Shun-xiang
Zhang, Yong
Luo, Le
Xie, Yun
Yang, Meng-jie
Zhang, Yi
Li, Jin
Shen, Hongwei
Li, Qi
Ma, Xue-jun
author_sort Nie, Kai
collection PubMed
description Human enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD) worldwide and has been associated with neurological complications which resulted in fatalities during recent outbreak in Asia pacific region. A direct reverse transcription loop-mediated isothermal amplification (direct RT-LAMP) assay using heat-treated samples without RNA extraction was developed and evaluated for the detection of EV71 subgenotype C4 in nasopharyngeal swab specimens. The analytical sensitivity and specificity of the direct RT-LAMP assay were examined. The detection limit of the direct RT-LAMP assays was 1.6 of a 50% tissue culture infective dose (TCID(50)) per reaction and no cross-reaction was observed with control viruses including Cosackievirus A (CVA) viruses (CVA2,4,5,7,9,10,14,16, and 24), Coxsackievirus B (CVB) viruses (CVB1,2,3,4, and 5) or ECHO viruses (ECHO3,6,11, and 19). The direct RT-LAMP assay was evaluated and compared to both RT-LAMP and quantitative real-time PCR (qRT-PCR) in detecting EV71 infection with 145 nasopharyngeal swab specimens. The clinical performance demonstrated the sensitivity and specificity of direct RT-LAMP was reported to be 90.3% and 100% respectively, compared to RT-LAMP, and 86.83% and 100% respectively, compared to qRT-PCR. These data demonstrated that the direct RT-LAMP assay can potentially be developed for the point of care screening of EV71 infection in China.
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spelling pubmed-35255322012-12-27 Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens Nie, Kai Qi, Shun-xiang Zhang, Yong Luo, Le Xie, Yun Yang, Meng-jie Zhang, Yi Li, Jin Shen, Hongwei Li, Qi Ma, Xue-jun PLoS One Research Article Human enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD) worldwide and has been associated with neurological complications which resulted in fatalities during recent outbreak in Asia pacific region. A direct reverse transcription loop-mediated isothermal amplification (direct RT-LAMP) assay using heat-treated samples without RNA extraction was developed and evaluated for the detection of EV71 subgenotype C4 in nasopharyngeal swab specimens. The analytical sensitivity and specificity of the direct RT-LAMP assay were examined. The detection limit of the direct RT-LAMP assays was 1.6 of a 50% tissue culture infective dose (TCID(50)) per reaction and no cross-reaction was observed with control viruses including Cosackievirus A (CVA) viruses (CVA2,4,5,7,9,10,14,16, and 24), Coxsackievirus B (CVB) viruses (CVB1,2,3,4, and 5) or ECHO viruses (ECHO3,6,11, and 19). The direct RT-LAMP assay was evaluated and compared to both RT-LAMP and quantitative real-time PCR (qRT-PCR) in detecting EV71 infection with 145 nasopharyngeal swab specimens. The clinical performance demonstrated the sensitivity and specificity of direct RT-LAMP was reported to be 90.3% and 100% respectively, compared to RT-LAMP, and 86.83% and 100% respectively, compared to qRT-PCR. These data demonstrated that the direct RT-LAMP assay can potentially be developed for the point of care screening of EV71 infection in China. Public Library of Science 2012-12-18 /pmc/articles/PMC3525532/ /pubmed/23272248 http://dx.doi.org/10.1371/journal.pone.0052486 Text en © 2012 Nie et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Nie, Kai
Qi, Shun-xiang
Zhang, Yong
Luo, Le
Xie, Yun
Yang, Meng-jie
Zhang, Yi
Li, Jin
Shen, Hongwei
Li, Qi
Ma, Xue-jun
Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title_full Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title_fullStr Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title_full_unstemmed Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title_short Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
title_sort evaluation of a direct reverse transcription loop-mediated isothermal amplification method without rna extraction for the detection of human enterovirus 71 subgenotype c4 in nasopharyngeal swab specimens
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3525532/
https://www.ncbi.nlm.nih.gov/pubmed/23272248
http://dx.doi.org/10.1371/journal.pone.0052486
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