Massive and rapid COVID-19 testing is feasible by extraction-free SARS-CoV-2 RT-PCR
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Abstract
Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is commonly diagnosed by reverse transcription polymerase chain reaction (RT-PCR) to detect viral RNA in patient samples, but RNA extraction constitutes a major bottleneck in current testing. Methodological simplification could increase diagnostic availability and efficiency, benefitting patient care and infection control. Here, we describe methods circumventing RNA extraction in COVID-19 testing by performing RT-PCR directly on heat-inactivated or lysed samples. Our data, including benchmarking using 597 clinical patient samples and a standardised diagnostic system, demonstrate that direct RT-PCR is viable option to extraction-based tests. Using controlled amounts of active SARS-CoV-2, we confirm effectiveness of heat inactivation by plaque assay and evaluate various generic buffers as transport medium for direct RT-PCR. Significant savings in time and cost are achieved through RNA-extraction-free protocols that are directly compatible with established PCR-based testing pipelines. This could aid expansion of COVID-19 testing.
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SciScore for 10.1101/2020.04.17.20067348: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement Consent: Informed consent was not obtained from the patients. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources SARS-CoV-2 in vitro assays: SARS-CoV-2 (GenBank: MT093571.1), originally from the Public Health Agency of Sweden, was propagated on 90% confluent Vero E6 cells (ATCC-CRL-1586) for 3 days at 37°C. Vero E6suggested: NoneResults from OddPub: Thank you for sharing your code and data.
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in …SciScore for 10.1101/2020.04.17.20067348: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement Consent: Informed consent was not obtained from the patients. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources SARS-CoV-2 in vitro assays: SARS-CoV-2 (GenBank: MT093571.1), originally from the Public Health Agency of Sweden, was propagated on 90% confluent Vero E6 cells (ATCC-CRL-1586) for 3 days at 37°C. Vero E6suggested: NoneResults from OddPub: Thank you for sharing your code and data.
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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