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J Clin Virol. Performance and clinical validation of the RealStar? MERS-CoV Kit for detection of Middle East respiratory syndrome coronavirus RNA
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Journal of Clinical Virology, Available online 28 March 2014 - In Press, Accepted Manuscript. / Short Communication
Performance and clinical validation of the RealStar? MERS-CoV Kit for detection of Middle East respiratory syndrome coronavirus RNA
Victor Max Corman<SUP>a</SUP>, Stephan ?lschl?ger<SUP>b</SUP>, Clemens-Martin Wendtner<SUP>c</SUP>, Jan Felix Drexler<SUP>a</SUP><SUP> </SUP><SUP>1</SUP>, Markus Hess<SUP>b</SUP>, Christian Drosten<SUP>a</SUP>
<SUP></SUP>
<SUP>a</SUP> Institute of Virology, University of Bonn Medical Centre, 53127 Bonn, Germany; <SUP>b</SUP> altona Diagnostics GmbH, M?rkenstrasse 12, 22767 Hamburg, Germany; <SUP>c</SUP> Klinikum Schwabing, Munich, Germany
Received 4 February 2014, Revised 17 March 2014, Accepted 19 March 2014, Available online 28 March 2014
http://dx.doi.org/10.1016/j.jcv.2014.03.012
Abstract
Background
A highly pathogenic human coronavirus causing respiratory disease emerged in the Middle East region in 2012. In-house molecular diagnostic methods for this virus termed Middle East respiratory syndrome coronavirus (MERS-CoV) allowed sensitive MERS-CoV RNA detection in patient samples. Fast diagnosis is important to manage human cases and trace possible contacts.
Objectives
The aim of this study was to improve the availability of existing nucleic acid amplification-based diagnostic methods for MERS-CoV infections by providing a RT-PCR kit, including an internal control and two target regions recommended by the World Health Organization (WHO). And to validate this real-time RT-PCR kit (RealStar? MERS-CoV RT-PCR Kit 1.0, altona Diagnostics GmbH, Hamburg, Germany) using clinical samples of one MERS-CoV case from Munich and respiratory samples of patients with other respiratory diseases.
Study design
An internal amplification control was included into the RT-PCR assays targeting the genomic region upstream of the Envelope gene (upE) and within open reading frame (ORF) 1A. Based on these assays, a ready-to-use RT-PCR kit featuring both the upE and ORF1A assays was developed, validated and compared to the established in-house versions.
Results
The performance of both RT-PCR assays included in the kit is comparable to the in-house assays. They show high analytical sensitivity (upE: 5.3 copies/reaction; ORF1A: 9.3 copies/reaction), no cross-reactivity with other respiratory pathogens and detected MERS-CoV RNA in patient samples in almost the same manner as the in-house versions.
Conclusion
The kit is a valuable tool for assisting in the rapid diagnosis, patient management and epidemiology of suspected MERS-CoV cases.
Keywords: Middle East respiratory syndrome; MERS-CoV; Coronavirus; Diagnostic assay; PCR
Corresponding author at: Institute of Virology, University of Bonn Medical Centre, Sigmund-Freud-Str. 25, 53127 Bonn, Germany. Tel.: +49 228 287 13590; fax: +49 228 287 19144.
1 current address: Department of Viroscience, Erasmus Medical Centre, Rotterdam, The Netherlands
Copyright ? 2014 Published by Elsevier B.V.
_____
Note to users: Accepted manuscripts are Articles in Press that have been peer reviewed and accepted for publication by the Editorial Board of this journal. They have not yet been copy edited and/or formatted in the journal house style, and may not yet have the full ScienceDirect functionality, e.g., supplementary files may still need to be added, links to references may not resolve yet etc. The text could still change before final publication.
Although accepted manuscripts do not have all bibliographic details available yet, they can already be cited using the year of online publication and the DOI, as follows: author(s), article title, journal (year), DOI. Please consult the journal's reference style for the exact appearance of these elements, abbreviation of journal names and use of punctuation.
When the final article is assigned to an issue of the journal, the Article in Press version will be removed and the final version will appear in the associated published issue of the journal. The date the article was first made available online will be carried over.
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Journal of Clinical Virology, Available online 28 March 2014 - In Press, Accepted Manuscript. / Short Communication
Performance and clinical validation of the RealStar? MERS-CoV Kit for detection of Middle East respiratory syndrome coronavirus RNA
Victor Max Corman<SUP>a</SUP>, Stephan ?lschl?ger<SUP>b</SUP>, Clemens-Martin Wendtner<SUP>c</SUP>, Jan Felix Drexler<SUP>a</SUP><SUP> </SUP><SUP>1</SUP>, Markus Hess<SUP>b</SUP>, Christian Drosten<SUP>a</SUP>
<SUP></SUP>
<SUP>a</SUP> Institute of Virology, University of Bonn Medical Centre, 53127 Bonn, Germany; <SUP>b</SUP> altona Diagnostics GmbH, M?rkenstrasse 12, 22767 Hamburg, Germany; <SUP>c</SUP> Klinikum Schwabing, Munich, Germany
Received 4 February 2014, Revised 17 March 2014, Accepted 19 March 2014, Available online 28 March 2014
http://dx.doi.org/10.1016/j.jcv.2014.03.012
Abstract
Background
A highly pathogenic human coronavirus causing respiratory disease emerged in the Middle East region in 2012. In-house molecular diagnostic methods for this virus termed Middle East respiratory syndrome coronavirus (MERS-CoV) allowed sensitive MERS-CoV RNA detection in patient samples. Fast diagnosis is important to manage human cases and trace possible contacts.
Objectives
The aim of this study was to improve the availability of existing nucleic acid amplification-based diagnostic methods for MERS-CoV infections by providing a RT-PCR kit, including an internal control and two target regions recommended by the World Health Organization (WHO). And to validate this real-time RT-PCR kit (RealStar? MERS-CoV RT-PCR Kit 1.0, altona Diagnostics GmbH, Hamburg, Germany) using clinical samples of one MERS-CoV case from Munich and respiratory samples of patients with other respiratory diseases.
Study design
An internal amplification control was included into the RT-PCR assays targeting the genomic region upstream of the Envelope gene (upE) and within open reading frame (ORF) 1A. Based on these assays, a ready-to-use RT-PCR kit featuring both the upE and ORF1A assays was developed, validated and compared to the established in-house versions.
Results
The performance of both RT-PCR assays included in the kit is comparable to the in-house assays. They show high analytical sensitivity (upE: 5.3 copies/reaction; ORF1A: 9.3 copies/reaction), no cross-reactivity with other respiratory pathogens and detected MERS-CoV RNA in patient samples in almost the same manner as the in-house versions.
Conclusion
The kit is a valuable tool for assisting in the rapid diagnosis, patient management and epidemiology of suspected MERS-CoV cases.
Keywords: Middle East respiratory syndrome; MERS-CoV; Coronavirus; Diagnostic assay; PCR
Corresponding author at: Institute of Virology, University of Bonn Medical Centre, Sigmund-Freud-Str. 25, 53127 Bonn, Germany. Tel.: +49 228 287 13590; fax: +49 228 287 19144.
1 current address: Department of Viroscience, Erasmus Medical Centre, Rotterdam, The Netherlands
Copyright ? 2014 Published by Elsevier B.V.
_____
Note to users: Accepted manuscripts are Articles in Press that have been peer reviewed and accepted for publication by the Editorial Board of this journal. They have not yet been copy edited and/or formatted in the journal house style, and may not yet have the full ScienceDirect functionality, e.g., supplementary files may still need to be added, links to references may not resolve yet etc. The text could still change before final publication.
Although accepted manuscripts do not have all bibliographic details available yet, they can already be cited using the year of online publication and the DOI, as follows: author(s), article title, journal (year), DOI. Please consult the journal's reference style for the exact appearance of these elements, abbreviation of journal names and use of punctuation.
When the final article is assigned to an issue of the journal, the Article in Press version will be removed and the final version will appear in the associated published issue of the journal. The date the article was first made available online will be carried over.
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