Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems

Objective: When using high-throughput batched diagnostic platforms based on RT-PCR for SARS-CoV-2 detection, avoidance of the conventional nucleic acid extraction step can help to reduce the turnaround time and increase processivity. This approach can also spare reagents and plasticware, which have...

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Autores principales:	Fabio Morecchiato, Marco Coppi, Ilaria Baccani, Niccolò Maggini, Nunziata Ciccone, Alberto Antonelli, Gian Maria Rossolini
Formato:	article
Lenguaje:	EN
Publicado:	Elsevier 2021
Materias:	SARS-CoV-2 diagnostic platform extraction-free method real-time polymerase chain reaction molecular diagnostics Infectious and parasitic diseases RC109-216
Acceso en línea:	https://doaj.org/article/efb90aaf7c7e450286210d23b1e98590
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spelling	oai:doaj.org-article:efb90aaf7c7e450286210d23b1e985902021-11-30T04:14:22ZEvaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems1201-971210.1016/j.ijid.2021.09.046https://doaj.org/article/efb90aaf7c7e450286210d23b1e985902021-11-01T00:00:00Zhttp://www.sciencedirect.com/science/article/pii/S1201971221007542https://doaj.org/toc/1201-9712Objective: When using high-throughput batched diagnostic platforms based on RT-PCR for SARS-CoV-2 detection, avoidance of the conventional nucleic acid extraction step can help to reduce the turnaround time and increase processivity. This approach can also spare reagents and plasticware, which have experienced a shortage during the initial waves of the pandemic, reducing the overall testing costs. Methods: This study evaluated the performance of extraction-free protocols based on simple dilution of the specimen in sterile RNAse free water (with or without a heating step) in comparison to standard RNA extraction protocols, using two commercial kits for molecular detection of SARS-CoV-2 (Allplex™ SARS-CoV-2 assay and Allplex™ SARS-CoV-2/FluA/FluB/RSV assay) in nasopharyngeal swabs (NPS). Results: Compared with conventional protocols, extraction-free protocols based on sample dilution without a heating step exhibited a lower analytical sensitivity: 74.0% and 82.1% with the Allplex™ SARS-CoV-2 assay (tested with 139 NPS samples) and the Allplex™ SARS-CoV-2/FluA/FluB/RSV assay (tested with 69 NPS samples), with a mean increase of Ct values of +2.04 and +1.32, respectively. Most false negative results were observed with sampled low viral load. Including a step of heat exposure did not improve but actually decreased the analytical sensitivity of the assay. Conclusions: Results confirmed that extraction-free protocols could be a faster and cheaper approach to SARS-CoV-2 detection in NPS samples, which could improve processivity of diagnostic platforms.Fabio MorecchiatoMarco CoppiIlaria BaccaniNiccolò MagginiNunziata CicconeAlberto AntonelliGian Maria RossoliniElsevierarticleSARS-CoV-2diagnostic platformextraction-free methodreal-time polymerase chain reactionmolecular diagnosticsInfectious and parasitic diseasesRC109-216ENInternational Journal of Infectious Diseases, Vol 112, Iss , Pp 264-268 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	SARS-CoV-2 diagnostic platform extraction-free method real-time polymerase chain reaction molecular diagnostics Infectious and parasitic diseases RC109-216
spellingShingle	SARS-CoV-2 diagnostic platform extraction-free method real-time polymerase chain reaction molecular diagnostics Infectious and parasitic diseases RC109-216 Fabio Morecchiato Marco Coppi Ilaria Baccani Niccolò Maggini Nunziata Ciccone Alberto Antonelli Gian Maria Rossolini Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
description	Objective: When using high-throughput batched diagnostic platforms based on RT-PCR for SARS-CoV-2 detection, avoidance of the conventional nucleic acid extraction step can help to reduce the turnaround time and increase processivity. This approach can also spare reagents and plasticware, which have experienced a shortage during the initial waves of the pandemic, reducing the overall testing costs. Methods: This study evaluated the performance of extraction-free protocols based on simple dilution of the specimen in sterile RNAse free water (with or without a heating step) in comparison to standard RNA extraction protocols, using two commercial kits for molecular detection of SARS-CoV-2 (Allplex™ SARS-CoV-2 assay and Allplex™ SARS-CoV-2/FluA/FluB/RSV assay) in nasopharyngeal swabs (NPS). Results: Compared with conventional protocols, extraction-free protocols based on sample dilution without a heating step exhibited a lower analytical sensitivity: 74.0% and 82.1% with the Allplex™ SARS-CoV-2 assay (tested with 139 NPS samples) and the Allplex™ SARS-CoV-2/FluA/FluB/RSV assay (tested with 69 NPS samples), with a mean increase of Ct values of +2.04 and +1.32, respectively. Most false negative results were observed with sampled low viral load. Including a step of heat exposure did not improve but actually decreased the analytical sensitivity of the assay. Conclusions: Results confirmed that extraction-free protocols could be a faster and cheaper approach to SARS-CoV-2 detection in NPS samples, which could improve processivity of diagnostic platforms.
format	article
author	Fabio Morecchiato Marco Coppi Ilaria Baccani Niccolò Maggini Nunziata Ciccone Alberto Antonelli Gian Maria Rossolini
author_facet	Fabio Morecchiato Marco Coppi Ilaria Baccani Niccolò Maggini Nunziata Ciccone Alberto Antonelli Gian Maria Rossolini
author_sort	Fabio Morecchiato
title	Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
title_short	Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
title_full	Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
title_fullStr	Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
title_full_unstemmed	Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems
title_sort	evaluation of extraction-free rt-pcr methods for faster and cheaper detection of sars-cov-2 using two commercial systems
publisher	Elsevier
publishDate	2021
url	https://doaj.org/article/efb90aaf7c7e450286210d23b1e98590
work_keys_str_mv	AT fabiomorecchiato evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT marcocoppi evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT ilariabaccani evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT niccolomaggini evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT nunziataciccone evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT albertoantonelli evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems AT gianmariarossolini evaluationofextractionfreertpcrmethodsforfasterandcheaperdetectionofsarscov2usingtwocommercialsystems
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Evaluation of extraction-free RT-PCR methods for faster and cheaper detection of SARS-CoV-2 using two commercial systems

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