False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors
Autor(a) principal: | |
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Data de Publicação: | 2020 |
Outros Autores: | , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) |
Texto Completo: | http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1676-24442020000100427 |
Resumo: | ABSTRACT Introduction: Although reverse transcription-polymerase chain reaction (rRT-PCR) is the gold standard method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), some factors, such as the presence of amplification inhibitors, lead to false-negative results. Objective: Here we describe the differences between rRT-PCR results for SARS-CoV-2 infection in normal and diluted samples, simulating the need for dilution due to the presence of amplification inhibitors. Material and method: Viral ribonucleic acid (RNA) from samples of nasopharyngeal swabs from 20 patients previously detected as “Negative” and 21 patients detected as “Positive” for SARS-CoV-2 was performed with the EasyExtract DNA-RNA kit (Interprise®). The rRT-PCR was performed with the OneStep/COVID-19 kit (IBMP), with normal and diluted (80 µl of H2O RNAse free) samples, totaling 82 tests. Results: The results indicate that there is an average variation (a < 0.05) delaying the Cq between the results of amplification of the internal control (IC), N gene (NG), and ORF1ab (OF), 1.811 Cq, 3.840 Cq, and 3.842 Cq, respectively. Discussion: The extraction kit does not completely purify the inhibitor compounds; therefore, no amplified product result may occur. In this study, we obtained a 19.04% false-negative diagnosis after sample dilution; this process reduces the efficiency of rRT-PCR to 29.8% in detecting SARS-CoV-2. Conclusion: Knowing the rRT-PCR standards of diluted samples can assist in the identification of false-negative cases and, consequently, avoid incorrect diagnosis. |
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False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitorsCOVID-19rRT-PCRdilutionviral diagnosisRNA extractionABSTRACT Introduction: Although reverse transcription-polymerase chain reaction (rRT-PCR) is the gold standard method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), some factors, such as the presence of amplification inhibitors, lead to false-negative results. Objective: Here we describe the differences between rRT-PCR results for SARS-CoV-2 infection in normal and diluted samples, simulating the need for dilution due to the presence of amplification inhibitors. Material and method: Viral ribonucleic acid (RNA) from samples of nasopharyngeal swabs from 20 patients previously detected as “Negative” and 21 patients detected as “Positive” for SARS-CoV-2 was performed with the EasyExtract DNA-RNA kit (Interprise®). The rRT-PCR was performed with the OneStep/COVID-19 kit (IBMP), with normal and diluted (80 µl of H2O RNAse free) samples, totaling 82 tests. Results: The results indicate that there is an average variation (a < 0.05) delaying the Cq between the results of amplification of the internal control (IC), N gene (NG), and ORF1ab (OF), 1.811 Cq, 3.840 Cq, and 3.842 Cq, respectively. Discussion: The extraction kit does not completely purify the inhibitor compounds; therefore, no amplified product result may occur. In this study, we obtained a 19.04% false-negative diagnosis after sample dilution; this process reduces the efficiency of rRT-PCR to 29.8% in detecting SARS-CoV-2. Conclusion: Knowing the rRT-PCR standards of diluted samples can assist in the identification of false-negative cases and, consequently, avoid incorrect diagnosis.Sociedade Brasileira de Patologia Clínica2020-01-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1676-24442020000100427Jornal Brasileiro de Patologia e Medicina Laboratorial v.56 2020reponame:Jornal Brasileiro de Patologia e Medicina Laboratorial (Online)instname:Sociedade Brasileira de Patologia (SBP)instacron:SBP10.5935/1676-2444.20200060info:eu-repo/semantics/openAccessFruehwirth,MarceloRivas,Açucena V.Fitz,Andressa F. R.Batista,Aline Cristiane C. A.Silveira,Cleypson ViniciusDelai,Robson M.eng2020-11-25T00:00:00Zoai:scielo:S1676-24442020000100427Revistahttp://www.scielo.br/jbpmlhttps://old.scielo.br/oai/scielo-oai.php||jbpml@sbpc.org.br1678-47741676-2444opendoar:2020-11-25T00:00Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) - Sociedade Brasileira de Patologia (SBP)false |
dc.title.none.fl_str_mv |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
title |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
spellingShingle |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors Fruehwirth,Marcelo COVID-19 rRT-PCR dilution viral diagnosis RNA extraction |
title_short |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
title_full |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
title_fullStr |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
title_full_unstemmed |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
title_sort |
False-negative result in molecular diagnosis of SARS-CoV-2 in samples with amplification inhibitors |
author |
Fruehwirth,Marcelo |
author_facet |
Fruehwirth,Marcelo Rivas,Açucena V. Fitz,Andressa F. R. Batista,Aline Cristiane C. A. Silveira,Cleypson Vinicius Delai,Robson M. |
author_role |
author |
author2 |
Rivas,Açucena V. Fitz,Andressa F. R. Batista,Aline Cristiane C. A. Silveira,Cleypson Vinicius Delai,Robson M. |
author2_role |
author author author author author |
dc.contributor.author.fl_str_mv |
Fruehwirth,Marcelo Rivas,Açucena V. Fitz,Andressa F. R. Batista,Aline Cristiane C. A. Silveira,Cleypson Vinicius Delai,Robson M. |
dc.subject.por.fl_str_mv |
COVID-19 rRT-PCR dilution viral diagnosis RNA extraction |
topic |
COVID-19 rRT-PCR dilution viral diagnosis RNA extraction |
description |
ABSTRACT Introduction: Although reverse transcription-polymerase chain reaction (rRT-PCR) is the gold standard method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), some factors, such as the presence of amplification inhibitors, lead to false-negative results. Objective: Here we describe the differences between rRT-PCR results for SARS-CoV-2 infection in normal and diluted samples, simulating the need for dilution due to the presence of amplification inhibitors. Material and method: Viral ribonucleic acid (RNA) from samples of nasopharyngeal swabs from 20 patients previously detected as “Negative” and 21 patients detected as “Positive” for SARS-CoV-2 was performed with the EasyExtract DNA-RNA kit (Interprise®). The rRT-PCR was performed with the OneStep/COVID-19 kit (IBMP), with normal and diluted (80 µl of H2O RNAse free) samples, totaling 82 tests. Results: The results indicate that there is an average variation (a < 0.05) delaying the Cq between the results of amplification of the internal control (IC), N gene (NG), and ORF1ab (OF), 1.811 Cq, 3.840 Cq, and 3.842 Cq, respectively. Discussion: The extraction kit does not completely purify the inhibitor compounds; therefore, no amplified product result may occur. In this study, we obtained a 19.04% false-negative diagnosis after sample dilution; this process reduces the efficiency of rRT-PCR to 29.8% in detecting SARS-CoV-2. Conclusion: Knowing the rRT-PCR standards of diluted samples can assist in the identification of false-negative cases and, consequently, avoid incorrect diagnosis. |
publishDate |
2020 |
dc.date.none.fl_str_mv |
2020-01-01 |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1676-24442020000100427 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1676-24442020000100427 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.5935/1676-2444.20200060 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
text/html |
dc.publisher.none.fl_str_mv |
Sociedade Brasileira de Patologia Clínica |
publisher.none.fl_str_mv |
Sociedade Brasileira de Patologia Clínica |
dc.source.none.fl_str_mv |
Jornal Brasileiro de Patologia e Medicina Laboratorial v.56 2020 reponame:Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) instname:Sociedade Brasileira de Patologia (SBP) instacron:SBP |
instname_str |
Sociedade Brasileira de Patologia (SBP) |
instacron_str |
SBP |
institution |
SBP |
reponame_str |
Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) |
collection |
Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) |
repository.name.fl_str_mv |
Jornal Brasileiro de Patologia e Medicina Laboratorial (Online) - Sociedade Brasileira de Patologia (SBP) |
repository.mail.fl_str_mv |
||jbpml@sbpc.org.br |
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1752122297649463296 |