High-density amplicon sequencing identifies community spread and ongoing evolution of SARS-CoV-2 in the Southern United States

  1. Ryan P. McNamara
  2. Carolina Caro-Vegas
  3. Justin T. Landis
  4. Razia Moorad
  5. Linda J. Pluta
  6. Anthony B. Eason
  7. Cecilia Thompson
  8. Aubrey Bailey
  9. Femi Cleola S. Villamor
  10. Philip T. Lange
  11. Jason P. Wong
  12. Tischan Seltzer
  13. Jedediah Seltzer
  14. Yijun Zhou
  15. Wolfgang Vahrson
  16. Angelica Juarez
  17. James O. Meyo
  18. Tiphaine Calabre
  19. Grant Broussard
  20. Ricardo Rivera-Soto
  21. Danielle L. Chappell
  22. Ralph S. Baric
  23. Blossom Damania
  24. Melissa B. Miller
  25. Dirk P. Dittmer

Reviewed by

Read the full article See related articles

Discuss this preprint

Start a discussion What are Sciety discussions?

Listed in

Log in to save this article

Abstract

SARS-CoV-2 is constantly evolving. Prior studies have focused on high case-density locations, such as the Northern and Western metropolitan areas in the U.S. This study demonstrates continued SARS-CoV-2 evolution in a suburban Southern U.S. region by high-density amplicon sequencing of symptomatic cases. 57% of strains carried the spike D614G variant. The presence of D614G was associated with a higher genome copy number and its prevalence expanded with time. Four strains carried a deletion in a predicted stem loop of the 3’ untranslated region. The data are consistent with community spread within the local population and the larger continental U.S. No strain had mutations in the target sites used in common diagnostic assays. The data instill confidence in the sensitivity of current tests and validate “testing by sequencing” as a new option to uncover cases, particularly those not conforming to the standard clinical presentation of COVID-19. This study contributes to the understanding of COVID-19 by providing an extensive set of genomes from a non-urban setting and further informs vaccine design by defining D614G as a dominant and emergent SARS-CoV-2 isolate in the U.S.

Article activity feed

  1. ScreenIT

    SciScore for 10.1101/2020.06.19.161141: (What is this?)

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

    Table 1: Rigor

    NIH rigor criteria are not applicable to paper type.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    Amplicon sizes ranged between 68 and 232 nucleotides after trimming of low-quality sequences and all primer sequences (125-275 before trimming).
    Amplicon
    suggested: (Amplicon, RRID:SCR_003294)
    Phylogenetic reconstruction: Full-length, viral genome consensus sequences were aligned using MAFFT (Katoh and Standley, 2013) with a PAM200 / k =2 scoring matrix, gap open penalty of 1.53 and offset value of 0.123 as implemented in Genious (Genious Ltd) using n = 92 sequences.
    MAFFT
    suggested: (MAFFT, RRID:SCR_011811)
    S protein sequences were analyzed using MEGA X version 10.1.7 (Kumar et al., 2018).
    MEGA
    suggested: (Mega BLAST, RRID:SCR_011920)

    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:
    Within the limitations presented by measuring viral loads within samples collected at unknown times past infection and with presumably differing clinical sampling efficiency, patients with the D614G SNV presented with higher SARS-CoV-2 genome loads. Whereas the association of the D614G SNV with specific clinical presentations and high peak titers remains the subject of debate (Zhang et al., 2020)(doi.org/10.1101/2020.04.29.069054), it is clear that this variant signifies spread within Europe and the continental U.S. Given the increasing abundance of D614G SNVs, further research into its role in pathogenicity and clinical outcomes is warranted. Four samples had the same 14 nt deletion in the 3’ UTR, and no samples had deletions within the coding region. This deletion is 71 nt away from the stop codon of orf10 (N protein) and eliminates a predicted stem-loop structure. An analogous bulged stem-loop at about the same location, right after the stop codon, is important for the replication of mouse hepatitis virus. In bovine coronaviruses an analogous RNA structure attenuates viral replication (Williams et al., 1999; Zust et al., 2008). There seems to be partial overlap between the bulged stem-loop and the pseudoknot, suggesting that these two structures are mutually exclusive and may serve as a switch to regulate the ratio of full length RNA and defective RNA (Goebel et al., 2004). These two structures are also present in SARS-CoV. These isolates represent full-length genomes from...

    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 10.1101/2020.06.19.161141 on bioRxiv