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Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)

Rapid diagnosis is an important intervention in managing the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) outbreak. Real time reverse transcription polymerase chain reaction (RT-qPCR) remains the primary means for diagnosing the new virus strain but it is time consuming and costly. R...

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Autores principales: Lau, Yee Ling, Ismail, Ilyiana binti, Mustapa, Nur Izati binti, Lai, Meng Yee, Tuan Soh, Tuan Suhaila, Haji Hassan, Afifah, Peariasamy, Kalaiarasu M., Lee, Yee Leng, Abdul Kahar, Maria Kahar Bador, Chong, Jennifer, Goh, Pik Pin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7787525/
https://www.ncbi.nlm.nih.gov/pubmed/33406112
http://dx.doi.org/10.1371/journal.pone.0245164
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author Lau, Yee Ling
Ismail, Ilyiana binti
Mustapa, Nur Izati binti
Lai, Meng Yee
Tuan Soh, Tuan Suhaila
Haji Hassan, Afifah
Peariasamy, Kalaiarasu M.
Lee, Yee Leng
Abdul Kahar, Maria Kahar Bador
Chong, Jennifer
Goh, Pik Pin
author_facet Lau, Yee Ling
Ismail, Ilyiana binti
Mustapa, Nur Izati binti
Lai, Meng Yee
Tuan Soh, Tuan Suhaila
Haji Hassan, Afifah
Peariasamy, Kalaiarasu M.
Lee, Yee Leng
Abdul Kahar, Maria Kahar Bador
Chong, Jennifer
Goh, Pik Pin
author_sort Lau, Yee Ling
collection PubMed
description Rapid diagnosis is an important intervention in managing the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) outbreak. Real time reverse transcription polymerase chain reaction (RT-qPCR) remains the primary means for diagnosing the new virus strain but it is time consuming and costly. Recombinase polymerase amplification (RPA) is an isothermal amplification assay that does not require a PCR machine. It is an affordable, rapid, and simple assay. In this study, we developed and optimized a sensitive reverse transcription (RT)-RPA assay for the rapid detection of SARS-CoV-2 using SYBR Green I and/or lateral flow (LF) strip. The analytical sensitivity and specificity of the RT-RPA assay were tested by using 10-fold serial diluted synthetic RNA and genomic RNA of similar viruses, respectively. Clinical sensitivity and specificity of the RT-RPA assay were carried out using 78 positive and 35 negative nasopharyngeal samples. The detection limit of both RPA and RT-qPCR assays was 7.659 and 5 copies/μL RNA, respectively with no cross reactivity with other viruses. The clinical sensitivity and specificity of RT-RPA were 98% and 100%, respectively. Our study showed that RT-RPA represents a viable alternative to RT-qPCR for the detection of SARS-CoV-2, especially in areas with limited infrastructure.
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spelling pubmed-77875252021-01-14 Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Lau, Yee Ling Ismail, Ilyiana binti Mustapa, Nur Izati binti Lai, Meng Yee Tuan Soh, Tuan Suhaila Haji Hassan, Afifah Peariasamy, Kalaiarasu M. Lee, Yee Leng Abdul Kahar, Maria Kahar Bador Chong, Jennifer Goh, Pik Pin PLoS One Research Article Rapid diagnosis is an important intervention in managing the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) outbreak. Real time reverse transcription polymerase chain reaction (RT-qPCR) remains the primary means for diagnosing the new virus strain but it is time consuming and costly. Recombinase polymerase amplification (RPA) is an isothermal amplification assay that does not require a PCR machine. It is an affordable, rapid, and simple assay. In this study, we developed and optimized a sensitive reverse transcription (RT)-RPA assay for the rapid detection of SARS-CoV-2 using SYBR Green I and/or lateral flow (LF) strip. The analytical sensitivity and specificity of the RT-RPA assay were tested by using 10-fold serial diluted synthetic RNA and genomic RNA of similar viruses, respectively. Clinical sensitivity and specificity of the RT-RPA assay were carried out using 78 positive and 35 negative nasopharyngeal samples. The detection limit of both RPA and RT-qPCR assays was 7.659 and 5 copies/μL RNA, respectively with no cross reactivity with other viruses. The clinical sensitivity and specificity of RT-RPA were 98% and 100%, respectively. Our study showed that RT-RPA represents a viable alternative to RT-qPCR for the detection of SARS-CoV-2, especially in areas with limited infrastructure. Public Library of Science 2021-01-06 /pmc/articles/PMC7787525/ /pubmed/33406112 http://dx.doi.org/10.1371/journal.pone.0245164 Text en © 2021 Lau 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 (http://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
Lau, Yee Ling
Ismail, Ilyiana binti
Mustapa, Nur Izati binti
Lai, Meng Yee
Tuan Soh, Tuan Suhaila
Haji Hassan, Afifah
Peariasamy, Kalaiarasu M.
Lee, Yee Leng
Abdul Kahar, Maria Kahar Bador
Chong, Jennifer
Goh, Pik Pin
Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title_full Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title_fullStr Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title_full_unstemmed Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title_short Development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)
title_sort development of a reverse transcription recombinase polymerase amplification assay for rapid and direct visual detection of severe acute respiratory syndrome coronavirus 2 (sars-cov-2)
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7787525/
https://www.ncbi.nlm.nih.gov/pubmed/33406112
http://dx.doi.org/10.1371/journal.pone.0245164
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