Current RT-qPCR to detect SARS-CoV-2 may give positive results for related coronaviruses

  1. Antonio Martínez-Murcia
  2. Adrián García-Sirera
  3. Aaron Navarro
  4. Laura Pérez

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Abstract

Some weeks after the first CoVID-19 outbreak, the World Health Organization published some real-time PCR (qPCR) protocols developed by different health reference centers. These qPCR designs are being used worldwide to detect SARS-CoV-2 in the population, to monitor the prevalence of the virus during the pandemic. Moreover, some of these protocols to detect SARS-CoV-2 have widely been applied to environmental samples for epidemiological surveillance purposes. In the present work, the specificity of these currently used RT-qPCR designs was validated in vitro using SARS-CoV-2 and highly related coronaviral genomic sequences and compared to performance of the commercially available GPS™ CoVID-19 dtec-RT-qPCR Test. Assays performed with SARS-CoV-2-related genomes showed positive amplification when using some of these qPCR methods, indicating they may give SARS-CoV-2 false positives. This finding may be particularly relevant for SARS-CoV-2 monitoring of environmental samples, where an unknown pool of phylogenetically close-related viruses may exist.

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  1. Version published to 10.1007/s00203-022-03029-y
  2. ScreenIT

    SciScore for 10.1101/2021.02.08.21251332: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Experimental Models: Organisms/Strains
    SentencesResources
    Primers and probes of IP2 and IP4 designs were provided by IDT (California, USA), and these of RdRp-P2, N1 and N2 were obtained from Eurofins Genomics Germany GmbH (Ebersberg, Germany).
    N2
    suggested: None
    Software and Algorithms
    SentencesResources
    The primers and probes sequences of the qPCR designs subjected to analysis in the present study (CDC Atlanta, 2020; Corman et al., 2020; Institut Pasteur, 2020; MartÍnez-Murcia et al., 2020) were used to find similar regions between close biological sequences using the Basic Local Alignment Search Tool (BLAST) software available on the National Center for Biotechnology Information (NCBI, https://blast.ncbi.nlm.nih.gov/Blast.cgi) website databases (Bethesda, MD, USA).
    BLAST
    suggested: (BLASTX, RRID:SCR_001653)
    https://blast.ncbi.nlm.nih.gov/Blast.cgi
    suggested: (TBLASTX, RRID:SCR_011823)

    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: 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 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:
    • 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.

    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.

  3. Version published to 10.1101/2021.02.08.21251332 on medRxiv