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Comparison of commercial realtime reverse transcription PCR assays for the detection of SARS-CoV-2.

ABSTRACT: The emergence of a new coronavirus in Wuhan China has triggered a global need for accurate diagnostic assays. Initially, mostly laboratory developed molecular tests were available but shortly thereafter different commercial assays started to appear and are still increasing in number. Although independent performance evaluations are ongoing, available data is still scarce. Here we provide a direct comparison of key performance characteristics of 13 commercial RT-PCR assays. Thirteen RT-PCR assays were selected based on the criteria that they can be used following generic RNA extraction protocols, on common PCR platforms and availability. Using a 10-fold and 2-fold dilution series of a quantified SARS-CoV-2 cell-cultured virus stock, performance was assessed compared to our in house validated assay. Specificity was tested by using RNA extracted from cultured common human coronaviruses. All RT-PCR kits included in this study exhibited PCR efficiencies > 90%, except for the Sentinel Diagnostics B E-gene RUO assay (80%). Analytical sensitivity varied between 3.3 RNA copies to 330 RNA copies. Only one assay cross reacted with another human coronavirus (MERS). This study provides a technical baseline of 13 different commercial PCR assays for SARS-CoV-2 detection that can be used by laboratories interested in purchasing any of these for further full clinical validation.

SUBMITTER: Igloi Z

PROVIDER: S-EPMC7833356 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Comparison of commercial realtime reverse transcription PCR assays for the detection of SARS-CoV-2.

Iglói Zsófia Z Leven Margareta M Abdel-Karem Abou-Nouar Zain Z Weller Babette B Matheeussen Veerle V Matheeussen Veerle V Coppens Jasmine J Koopmans Marion M Molenkamp Richard R

Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 20200617

The emergence of a new coronavirus in Wuhan China has triggered a global need for accurate diagnostic assays. Initially, mostly laboratory developed molecular tests were available but shortly thereafter different commercial assays started to appear and are still increasing in number. Although independent performance evaluations are ongoing, available data is still scarce. Here we provide a direct comparison of key performance characteristics of 13 commercial RT-PCR assays. Thirteen RT-PCR assays ...[more]

PMID: 32570045

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Project description:Most individuals acutely infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exhibit mild symptoms. However, 10 to 20% of those infected develop long-term symptoms, referred to as post-coronavirus disease 2019 (COVID-19) condition (PCC). One hypothesis is that PCC might be exacerbated by viral persistence in tissue sanctuaries. Therefore, the accurate detection and quantification of SARS-CoV-2 are not only necessary for viral load monitoring but also crucial for detecting long-term viral persistence and determining whether viral replication is occurring in tissue reservoirs. In this study, the sensitivity and robustness of reverse transcription (RT)-droplet digital PCR (ddPCR) and RT-quantitative PCR (qPCR) techniques have been compared for the detection and quantification of SARS-CoV-2 genomic and subgenomic RNAs from oropharyngeal swabs taken from confirmed SARS-CoV-2-positive, SARS-CoV-2-exposed, and nonexposed individuals as well as from samples from mice infected with SARS-CoV-2. Our data demonstrated that both techniques presented equivalent results in the mid- and high-viral-load ranges. Additionally, RT-ddPCR was more sensitive than RT-qPCR in the low-viral-load range, allowing the accurate detection of positive results in individuals exposed to the virus. Overall, these data suggest that RT-ddPCR might be an alternative to RT-qPCR for detecting low viral loads in samples and for assessing viral persistence in samples from individuals with PCC. IMPORTANCE We developed one-step reverse transcription (RT)-droplet digital PCR (ddPCR) protocols to detect SARS-CoV-2 RNA and compared them to the gold-standard RT-quantitative PCR (RT-qPCR) method. RT-ddPCR was more sensitive than RT-qPCR in the low-viral-load range, while both techniques were equivalent in the mid- and high-viral-load ranges. Overall, these results suggest that RT-ddPCR might be a viable alternative to RT-qPCR when it comes to detecting low viral loads in samples, which is a highly relevant issue for determining viral persistence in as-yet-unknown tissue reservoirs in individuals suffering from post-COVID conditions or long COVID.

Dataset Information

Comparison of commercial realtime reverse transcription PCR assays for the detection of SARS-CoV-2.

Publications

Comparison of commercial realtime reverse transcription PCR assays for the detection of SARS-CoV-2.

Similar Datasets

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