SARS-CoV-2 Detection in the Nasopharyngeal Swabs and Saliva of College Students using RT-qPCR and RT-LAMP

  1. D. A. Bikos
  2. C. Hwang
  3. K. A. Brileya
  4. A. Parker
  5. E. K. Loveday
  6. M. Rodriguez
  7. I. Thornton
  8. T. LeFevre
  9. J. N. Wilking
  10. M. Dills
  11. S. T. Walk
  12. A. K. Adams
  13. R. K. Plowright
  14. A. B. Hoegh
  15. J. R. Carter
  16. J. Morrow
  17. M. P. Taylor
  18. D. E. Keil
  19. M. W. Fields
  20. C. B. Chang

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Abstract

Background

Diagnostic testing can identify outbreaks and inform preventive strategies for slowing the spread of SARS-CoV-2, the virus that causes Covid-19. The “gold standard” method for detection of SARS-CoV-2 is reverse transcription quantitative polymerase chain reaction (RT-qPCR) performed on samples collected using nasopharyngeal (NP) swabs. While NP RT-qPCR achieves high sensitivity, it requires trained personnel to administer and suffers from lengthy time-to-result. Instead, rapid saliva-based reverse transcription loop-mediated amplification (RT-LAMP) screening methods may offer advantages in sample collection and speed.

Methods

Regardless of symptomatic presentation, a total of 233 individuals were tested for SARS-CoV-2 using NP RT-qPCR, alongside saliva-based RT-qPCR (SalivirDetect) and RT-LAMP (SLAMP), a simple and rapid fluorometric RT-LAMP assay performed directly on heat-inactivated saliva without any additional treatments or RNA extraction. SLAMP is conducted in triplicate and takes 45 min. Samples found negative using both saliva-based methods but positive under CDC NP RT-qPCR above the saliva method LoD were excluded from evaluation, suggesting significant differences in viral titer between sampling sites. Individuals who consumed potential inhibitors in the form of food, drink, and oral health products within 30 min of sampling were identified using a self-reported questionnaire.

Results

Of the 233 NP RT-qPCR tests, 58 were positive and 175 were negative. Comparatively, SLAMP resulted in 95% sensitivity and 98% specificity and SalivirDetect 97% sensitivity and 98% specificity. Prior consumption had no measurable effect on test outcomes, except for drinking, which lowered Ct values in saliva.

Conclusions

SLAMP requires less technician and instrument time than CDC-approved NP RT-qPCR and demonstrates that saliva-based RT-LAMP can enable frequent and rapid identification of pre-symptomatic and asymptomatic SARS-CoV-2 infections with high sensitivity and specificity.

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    SciScore for 10.1101/2021.03.31.21254634: (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.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    Dilutions are inoculated onto susceptible E6-Vero cells (ATCC) and incubated for 1 h to facilitate infection.
    E6-Vero
    suggested: None
    Software and Algorithms
    SentencesResources
    Fitting was performed using KaleidaGraph v4.1 non-linear least squares routine.
    KaleidaGraph
    suggested: (KaleidaGraph, RRID:SCR_014980)

    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.

  2. Version published to 10.1101/2021.03.31.21254634 on medRxiv