Sources of variability in methods for processing, storing, and concentrating SARS-CoV-2 in influent from urban wastewater treatment plants

  1. Joshua A. Steele
  2. Amity G. Zimmer-Faust
  3. John F. Griffith
  4. Stephen B. Weisberg

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Abstract

The rapid emergence of wastewater based surveillance has led to a wide array of SARS-CoV-2 RNA quantification methodologies being employed. Here we compare methods to store samples, inactivate viruses, capture/concentrate viruses, and extract/measure viral RNA from primary influent into wastewater facilities. We found that heat inactivation of the viruses led to a 1-3 log 10 decrease compared to chemical inactivation. Freezing influent prior to concentration caused a 1-4 log 10 decrease compared to processing fresh samples, but viral capture by membrane adsorption prior to freezing was robust to freeze-thaw variability. Concentration vs. direct extraction, and PCR platform also affected outcome, but by a smaller amount. The choice of nucleocapsid gene target had nearly no effect. Pepper mild-mottle virus was much less sensitive to these methodological differences than was SARS-CoV-2, which challenges its use as a population-level control among studies using different methods. Better characterizing the variability associated with different methodologies, in particular the impact of methods on sensitivity, will aid decision makers in following the effects of vaccination campaigns, early detection of future outbreaks, and potentially monitoring the appearance of SARS-CoV-2 variants in the population.

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  1. ScreenIT

    SciScore for 10.1101/2021.06.16.21259063: (What is this?)

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

    Table 1: Rigor

    Ethicsnot detected.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot 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: 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.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

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  2. Version published to 10.1101/2021.06.16.21259063v1 on medRxiv