Validation and Comparison of a Modified CDC Assay with two Commercially Available Assays for the Detection of SARS-CoV-2 in Respiratory Specimen

  1. Amorce Lima
  2. Vicki Healer
  3. Elaine Vendrone
  4. Suzane Silbert

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Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (COVID-19), has spread rapidly around the globe since it was first identified in December of 2019 in Wuhan, China. In a race to contain the infection, researchers and healthcare officials have developed several assays to help diagnose individuals with COVID-19. To help laboratories in deciding what assay to bring into testing lines, factors such as assay availability, cost, throughput, and TAT should be considered. Here we validated a modified version of the CDC assay and used it as a reference to evaluate the performance of the NeuMoDx™ SARS-CoV-2 and DiaSorin Simplexa™ Covid-19 Direct assays. In silico analysis and clinical sample testing showed that the primesr/probes designed by the CDC were specific to the SARS-CoV-2 as they accurately detected all reactive samples with an assay LoD of 200 copies/ml. The performance of the three assays were analyzed using 161 nasopharyngeal swabs specimen tested within 24 hours or 5 days from routine testing. A 100% agreement was observed between the commercial assays and the modified CDC SARS-CoV-2 assay. A deeper look at the Ct values showed no significant difference between NeuMoDx and the modified CDC SARS-CoV-2 assay, whereas DiaSorin had lower overall Ct values than the modified CDC SARS-CoV-2 assay. NeuMoDx and DiaSorin workflows were much easier to perform. NeuMoDx has the highest throughput and shortest TAT, whereas although the modified CDC SARS-CoV-2 assay has comparable throughput to DiaSorin, it has the longest hands-on time, and highest TAT.

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  1. ScreenIT

    SciScore for 10.1101/2020.06.29.179192: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementIRB: Specimen enrollment: Residual respiratory specimens submitted for Covid-19 testing at Tampa General Hospital (TGH) Esoteric Testing Laboratory were used in this study according to the standard procedures and in compliance with the TGH Institutional Review Board (IRB) protocols.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    No key resources detected.

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    A limitation of this study was that the same samples were not tested by the three different assays, so a head-to-head comparison of the three assays was not performed. This was due to the limit of available kits for routine testing in patient care. However, we did a head-to-head comparison of the assay’s workflow. The NeuMoDx 96 molecular system is a sample-to-answer and random-access platform that automatically performs nucleic acid extraction, amplification, and signal detection and analysis requiring only very little human interaction for loading and scanning the samples (20). It has the shortest TAT and highest throughput of the three assays. DiaSorin Simplexa CoV-2 Direct assay involves a simple operation procedure that does not include an extraction step; it has, however, much lower throughput than NeuMoDx. The modified CDC SARS-CoV-2 assay is singleplex; each target must be run in different tubes, as opposed to the two commercial assays, which are multiplex. It requires separate extraction steps on the EasyMag which can extract up to 24 samples at a time, longer hands-on time, and has much lower throughput compared to NeuMoDx. It has comparable throughput to DiaSorin if nucleic acid extraction and PCR reaction tubes for the next sample batch are prepared before the PCR cycles of the previous batch ends. Therefore, although the three assays have the same accuracy, the overall workflow favors the commercial platforms. In conclusion, diagnostic laboratories around the wor...

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • No conflict of interest statement was detected. If there are no conflicts, we encourage authors to explicit state so.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.06.29.179192v1 on bioRxiv