A Single-Cell Atlas of Lymphocyte Adaptive Immune Repertoires and Transcriptomes Reveals Age-Related Differences in Convalescent COVID-19 Patients

  1. Florian Bieberich
  2. Rodrigo Vazquez-Lombardi
  3. Alexander Yermanos
  4. Roy A. Ehling
  5. Derek M. Mason
  6. Bastian Wagner
  7. Edo Kapetanovic
  8. Raphael Brisset Di Roberto
  9. Cédric R. Weber
  10. Miodrag Savic
  11. Fabian Rudolf
  12. Sai T. Reddy

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Abstract

COVID-19 disease outcome is highly dependent on adaptive immunity from T and B lymphocytes, which play a critical role in the control, clearance and long-term protection against SARS-CoV-2. To date, there is limited knowledge on the composition of the T and B cell immune receptor repertoires [T cell receptors (TCRs) and B cell receptors (BCRs)] and transcriptomes in convalescent COVID-19 patients of different age groups. Here, we utilize single-cell sequencing (scSeq) of lymphocyte immune repertoires and transcriptomes to quantitatively profile the adaptive immune response in COVID-19 patients of varying age. We discovered highly expanded T and B cells in multiple patients, with the most expanded clonotypes coming from the effector CD8 + T cell population. Highly expanded CD8 + and CD4 + T cell clones show elevated markers of cytotoxicity (CD8: PRF1, GZMH, GNLY; CD4: GZMA), whereas clonally expanded B cells show markers of transition into the plasma cell state and activation across patients. By comparing young and old convalescent COVID-19 patients (mean ages = 31 and 66.8 years, respectively), we found that clonally expanded B cells in young patients were predominantly of the IgA isotype and their BCRs had incurred higher levels of somatic hypermutation than elderly patients. In conclusion, our scSeq analysis defines the adaptive immune repertoire and transcriptome in convalescent COVID-19 patients and shows important age-related differences implicated in immunity against SARS-CoV-2.

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  1. Version published to 10.3389/fimmu.2021.701085
  2. ScreenIT

    SciScore for 10.1101/2021.02.12.430907: (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

    Antibodies
    SentencesResources
    After separation, the upper plasma layer was collected for ELISA detection of IgG and IgA SARS-CoV-2-specific antibodies (Euroimmun Medizinische Labordiagnostika, #EI2668-9601G, #EI2606-9601A).
    IgA SARS-CoV-2-specific
    suggested: None
    Point-of-care lateral flow immunoassays assessing the presence of IgG and IgM SARS-CoV-2-specific antibodies (Qingdao Hightop Biotech, #H100) were performed at the time of blood collection.
    IgM SARS-CoV-2-specific
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    Patient samples: Patients were participants of the SERO-BL-COVID-19 study sponsored by the Department of Health, Canton Basel-Landschaft, Switzerland.
    Canton Basel-Landschaft
    suggested: None
    Software and Algorithms
    SentencesResources
    Unique sequences originating from specific patients were identified from their respective DNA barcodes and aligned using the ClustalOmega tool to cluster sequences arising from the same allele.
    ClustalOmega
    suggested: None
    Sequences with the highest amount of reads in each cluster were used as input for the basic local alignment search tool (BLAST;
    BLAST
    suggested: (BLASTX, RRID:SCR_001653)
    Sequences returning matching or highly similar alleles across PCR 1 and PCR 2 in each patient were then assembled and queried against the IMGT/HLA database for final validation.
    IMGT/HLA
    suggested: (IMGT/HLA, RRID:SCR_002971)
    Subsequent data QC and analysis was performed using R (version 3.6.2) and the Seurat package (version 3.1.5).
    Seurat
    suggested: (SEURAT, RRID:SCR_007322)
    Germline identity was used as a proxy for somatic hypermutation levels and was calculated from alignments of BCR clonotypes with their corresponding VH and VL germline sequences.
    Germline
    suggested: (GERMLINE, RRID:SCR_001720)
    Pseudotime analysis: Pseudotime and trajectory inference was applied to scSeq transcriptome data using the slingshot function with default parameters from the Slingshot package in R70.
    Slingshot
    suggested: (Slingshot, RRID:SCR_017012)
    List of utilized R packages: Biobase (2.46.0), BiocGenerics (0.32.0), BiocParallel (1.20.1)
    BiocParallel
    suggested: None
    , Cell Ranger (3.1.0), Change-O (1.0.0), circlize (0.4.10), data.table (1.12.8), DelayedArray (0.12.3), dplyr (0.8.5), GenomeInfoDb (1.22.1), GenomicRanges (1.38.0), ggplot2 (3.3.2.9000), harmony (1.0), pheatmap (1.0.12), princurve (2.1.5), RColorBrewer (1.1-2), matrixStats (0.56.0), sctransform (0.2.1), Seurat (3.1.5), slingshot (1.4.0), stringdist (0.9.5.5), stringr (1.4.0), tibble (3.0.3), tidyr (1.1.0), tidyverse (1.3.0).
    circlize
    suggested: (circlize, RRID:SCR_002141)
    GenomicRanges
    suggested: (GenomicRanges, RRID:SCR_000025)
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)
    pheatmap
    suggested: (pheatmap, RRID:SCR_016418)

    Results from OddPub: Thank you for sharing your data.

    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

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  3. Version published to 10.1101/2021.02.12.430907v2 on bioRxiv
  4. Version published to 10.1101/2021.02.12.430907v1 on bioRxiv