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SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods

The COVID-19 pandemic has forced diagnostic laboratories to focus on the early diagnostics of SARS-CoV-2. The positivity of a molecular test cannot respond to the question regarding the viral capability to replicate, spread, and give different clinical effects. Despite the fact that some targets are...

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Autores principales: Zollo, Massimo, Ferrucci, Veronica, Izzo, Barbara, Quarantelli, Fabrizio, Domenico, Carmela Di, Comegna, Marika, Paolillo, Carmela, Amato, Felice, Siciliano, Roberto, Castaldo, Giuseppe, Capoluongo, Ettore
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7923082/
https://www.ncbi.nlm.nih.gov/pubmed/33673182
http://dx.doi.org/10.3390/diagnostics11020288
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author Zollo, Massimo
Ferrucci, Veronica
Izzo, Barbara
Quarantelli, Fabrizio
Domenico, Carmela Di
Comegna, Marika
Paolillo, Carmela
Amato, Felice
Siciliano, Roberto
Castaldo, Giuseppe
Capoluongo, Ettore
author_facet Zollo, Massimo
Ferrucci, Veronica
Izzo, Barbara
Quarantelli, Fabrizio
Domenico, Carmela Di
Comegna, Marika
Paolillo, Carmela
Amato, Felice
Siciliano, Roberto
Castaldo, Giuseppe
Capoluongo, Ettore
author_sort Zollo, Massimo
collection PubMed
description The COVID-19 pandemic has forced diagnostic laboratories to focus on the early diagnostics of SARS-CoV-2. The positivity of a molecular test cannot respond to the question regarding the viral capability to replicate, spread, and give different clinical effects. Despite the fact that some targets are covered by commercially-available assays, the identification of new biomarkers is desired in order to improve the quality of the information given by these assays. Therefore, since the subgenomic transcripts (sgN and sgE) are considered markers of viral activity, we evaluated these subgenomic transcripts in relation to the genomic amplification obtained using five different commercial CE-IVD tools. Methods: Five CE-IVD kits were compared in terms of their capability to detect both synthetic SARS-CoV-2 viral constructs (spiked in TMB or PBS medium) and targets (N, E, RdRp and Orf1ab genes) in twenty COVID-19–positive patients’ swabs. The sgN and sgE were assayed by real-time RT-qPCR and digital PCR. Results: None of the diagnostic kits missed the viral target genes when they were applied to targets spiked in TMB or PBS (at dilutions ranging from 100 pg to 0.1 pg). Nevertheless, once they were applied to RNA extracted from the patients’ swabs, the superimposability ranged from 50% to 100%, regardless of the extraction procedure. The sgN RNA transcript was detected only in samples with a higher viral load (Ct ≤ 22.5), while sgE was within all of the Ct ranges. Conclusions: The five kits show variable performances depending on the assay layout. It is worthy of note that the detection of the sgN transcript is associated with a higher viral load, thus representing a new marker of early and more severe infection.
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spelling pubmed-79230822021-03-03 SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods Zollo, Massimo Ferrucci, Veronica Izzo, Barbara Quarantelli, Fabrizio Domenico, Carmela Di Comegna, Marika Paolillo, Carmela Amato, Felice Siciliano, Roberto Castaldo, Giuseppe Capoluongo, Ettore Diagnostics (Basel) Article The COVID-19 pandemic has forced diagnostic laboratories to focus on the early diagnostics of SARS-CoV-2. The positivity of a molecular test cannot respond to the question regarding the viral capability to replicate, spread, and give different clinical effects. Despite the fact that some targets are covered by commercially-available assays, the identification of new biomarkers is desired in order to improve the quality of the information given by these assays. Therefore, since the subgenomic transcripts (sgN and sgE) are considered markers of viral activity, we evaluated these subgenomic transcripts in relation to the genomic amplification obtained using five different commercial CE-IVD tools. Methods: Five CE-IVD kits were compared in terms of their capability to detect both synthetic SARS-CoV-2 viral constructs (spiked in TMB or PBS medium) and targets (N, E, RdRp and Orf1ab genes) in twenty COVID-19–positive patients’ swabs. The sgN and sgE were assayed by real-time RT-qPCR and digital PCR. Results: None of the diagnostic kits missed the viral target genes when they were applied to targets spiked in TMB or PBS (at dilutions ranging from 100 pg to 0.1 pg). Nevertheless, once they were applied to RNA extracted from the patients’ swabs, the superimposability ranged from 50% to 100%, regardless of the extraction procedure. The sgN RNA transcript was detected only in samples with a higher viral load (Ct ≤ 22.5), while sgE was within all of the Ct ranges. Conclusions: The five kits show variable performances depending on the assay layout. It is worthy of note that the detection of the sgN transcript is associated with a higher viral load, thus representing a new marker of early and more severe infection. MDPI 2021-02-12 /pmc/articles/PMC7923082/ /pubmed/33673182 http://dx.doi.org/10.3390/diagnostics11020288 Text en © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Zollo, Massimo
Ferrucci, Veronica
Izzo, Barbara
Quarantelli, Fabrizio
Domenico, Carmela Di
Comegna, Marika
Paolillo, Carmela
Amato, Felice
Siciliano, Roberto
Castaldo, Giuseppe
Capoluongo, Ettore
SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title_full SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title_fullStr SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title_full_unstemmed SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title_short SARS-CoV-2 Subgenomic N (sgN) Transcripts in Oro-Nasopharyngeal Swabs Correlate with the Highest Viral Load, as Evaluated by Five Different Molecular Methods
title_sort sars-cov-2 subgenomic n (sgn) transcripts in oro-nasopharyngeal swabs correlate with the highest viral load, as evaluated by five different molecular methods
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7923082/
https://www.ncbi.nlm.nih.gov/pubmed/33673182
http://dx.doi.org/10.3390/diagnostics11020288
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