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A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan
The rapid identification of Influenza A virus and its variants, which cause severe respiratory diseases, is imperative to providing timely treatment and improving patient outcomes. Conventionally, two separate assays (total test duration of up to 6 h) are required to initially differentiate Influenz...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10035893/ https://www.ncbi.nlm.nih.gov/pubmed/36952488 http://dx.doi.org/10.1371/journal.pone.0283074 |
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author | Yang, Hui-Hua Huang, I-Tsong Wu, Ren-Chieh Chen, Li-Kuang |
author_facet | Yang, Hui-Hua Huang, I-Tsong Wu, Ren-Chieh Chen, Li-Kuang |
author_sort | Yang, Hui-Hua |
collection | PubMed |
description | The rapid identification of Influenza A virus and its variants, which cause severe respiratory diseases, is imperative to providing timely treatment and improving patient outcomes. Conventionally, two separate assays (total test duration of up to 6 h) are required to initially differentiate Influenza A and B viruses and subsequently distinguish the pdm H1N1 and H3N2 serotypes of Influenza A virus. In this study, we developed a multiplex real-time RT-PCR method for simultaneously detecting Influenza A and B viruses and subtyping Influenza A virus, with a substantially reduced test duration. Clinical specimens from hospitalized patients and outpatients with influenza-like symptoms in Eastern Taiwan were collected between 2011 and 2015, transported to Hualien Tzu Chi Hospital, and analyzed. Conventional RT-PCR was used to subtype the isolated Influenza A viruses. Thereafter, for rapid identification, the multiplex real-time RT-PCR method was developed and applied to identify the conserved regions that aligned with the available primers and probes. Accordingly, a multiplex RT-PCR assay with three groups of primers and probes (MAF and MAR primers and MA probe; InfAF and InfAR primers and InfA probe; and MBF and MBR primers and MB probe) was established to distinguish these viruses in the same reaction. Thus, with this multiplex RT-PCR assay, Influenza B, Influenza A pdm H1N1, and Influenza A H3N2 viruses were accurately detected and differentiated within only 2.5 h. This multiplex RT-PCR assay showed similar analytical sensitivity to the conventional singleplex assay. Further, the phylogenetic analyses of our samples revealed that the characteristics of these viruses were different from those reported previously using samples collected during 2012–2013. In conclusion, we developed a multiplex real-time RT-PCR method for highly efficient and accurate detection and differentiation of Influenza A and B viruses and subtyping Influenza A virus with a substantially reduced test duration for diagnosis. |
format | Online Article Text |
id | pubmed-10035893 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-100358932023-03-24 A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan Yang, Hui-Hua Huang, I-Tsong Wu, Ren-Chieh Chen, Li-Kuang PLoS One Research Article The rapid identification of Influenza A virus and its variants, which cause severe respiratory diseases, is imperative to providing timely treatment and improving patient outcomes. Conventionally, two separate assays (total test duration of up to 6 h) are required to initially differentiate Influenza A and B viruses and subsequently distinguish the pdm H1N1 and H3N2 serotypes of Influenza A virus. In this study, we developed a multiplex real-time RT-PCR method for simultaneously detecting Influenza A and B viruses and subtyping Influenza A virus, with a substantially reduced test duration. Clinical specimens from hospitalized patients and outpatients with influenza-like symptoms in Eastern Taiwan were collected between 2011 and 2015, transported to Hualien Tzu Chi Hospital, and analyzed. Conventional RT-PCR was used to subtype the isolated Influenza A viruses. Thereafter, for rapid identification, the multiplex real-time RT-PCR method was developed and applied to identify the conserved regions that aligned with the available primers and probes. Accordingly, a multiplex RT-PCR assay with three groups of primers and probes (MAF and MAR primers and MA probe; InfAF and InfAR primers and InfA probe; and MBF and MBR primers and MB probe) was established to distinguish these viruses in the same reaction. Thus, with this multiplex RT-PCR assay, Influenza B, Influenza A pdm H1N1, and Influenza A H3N2 viruses were accurately detected and differentiated within only 2.5 h. This multiplex RT-PCR assay showed similar analytical sensitivity to the conventional singleplex assay. Further, the phylogenetic analyses of our samples revealed that the characteristics of these viruses were different from those reported previously using samples collected during 2012–2013. In conclusion, we developed a multiplex real-time RT-PCR method for highly efficient and accurate detection and differentiation of Influenza A and B viruses and subtyping Influenza A virus with a substantially reduced test duration for diagnosis. Public Library of Science 2023-03-23 /pmc/articles/PMC10035893/ /pubmed/36952488 http://dx.doi.org/10.1371/journal.pone.0283074 Text en © 2023 Yang et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Yang, Hui-Hua Huang, I-Tsong Wu, Ren-Chieh Chen, Li-Kuang A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title | A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title_full | A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title_fullStr | A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title_full_unstemmed | A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title_short | A highly efficient and accurate method of detecting and subtyping Influenza A pdm H1N1 and H3N2 viruses with newly emerging mutations in the matrix gene in Eastern Taiwan |
title_sort | highly efficient and accurate method of detecting and subtyping influenza a pdm h1n1 and h3n2 viruses with newly emerging mutations in the matrix gene in eastern taiwan |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10035893/ https://www.ncbi.nlm.nih.gov/pubmed/36952488 http://dx.doi.org/10.1371/journal.pone.0283074 |
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