Repeated mRNA vaccines supercharge immune response against COVID-19, study finds

In a recent study published in the journal Nature Immunology, researchers investigated how repeated mRNA vaccinations improve COVID-19 immunity in SARS-CoV-2-naïve and priorly infected individuals. Focusing on the latter cohort, the study evaluated the diversity and concertation in tandem with multiple sequencing analyses of immune cells isolated from the patient’s peripheral blood mononuclear cells. Study findings reveal that sequential vaccination promotes heterogeneous immune cell clonal expansions, with the third mRNA vaccination resulting in almost two times the number of clones as the first vaccination dose. Parallelly, populations of CD8+ T cells substantially increase, thereby better preparing an individual’s immune system to cope with multiple COVID-19 strains. Surprisingly, the presence and severity of COVID-19 infection were directly associated with post-vaccination immunity.

Study: Repeated mRNA vaccination sequentially boosts SARS-CoV-2-specific CD8+ T cells in persons with previous COVID-19. Image Credit: CI Photos / ShutterstockStudy: Repeated mRNA vaccination sequentially boosts SARS-CoV-2-specific CD8+ T cells in persons with previous COVID-19. Image Credit: CI Photos / Shutterstock

Does infection prevent infection?

First, a disclaimer – under no circumstances are the authors of the publication or the author of this article recommending that you allow yourself to become infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as a means of improving your future immunity against the Coronavirus disease 2019 (COVID-19) pandemic. However, a number of previous studies have established the improved anti-COVID-19 capabilities of ‘hybrid immunity.’ Hybrid immunity arises from the combined effects of both a previous COVID-19 infection and vaccination, which have been consistently found to confer better protection than either alone.

Two main mechanisms for this improved observed immunity have been proposed – an increase in the abundance and diversity of virus-specific memory T (Tm) cells and an increased diversity of spike (S) proteins in the Tm cell pool. Hitherto, however, these hypotheses have never been scientifically tested. Understanding the mechanisms underpinning observed immune responses to COVID-19 in hybrid individuals may allow for improved and personalized vaccination regimes, thereby improving global resistance against the pandemic, which has hitherto infected more than 770 million individuals and cost humanity almost 7 million lives.

About the study

In the present study, researchers used peripheral blood mononuclear cells (PBMCs) to identify and quantify the variations in T and B cell populations across individuals exposed to multiple vaccination doses in tandem with COVID-19 infections of varying severity. They then focused their efforts on evaluating the kinetics and diversity of T spike (Ts) cells, thereby verifying both proposed hypothesized mechanisms underpinning previously observed immunity improvements.

The study cohort comprised 54 (28 female) self-reported COVID-19 survivors recruited from Seattle, USA, between April and August 2020. All participants had their demographic and medical data recorded and were subjected to convalescent plasma donation for PBMC isolation. The cohort consisted of 35 individuals with mild to moderate COVID-19, 19 with severe illness (hospital and oxygen support required), and eight with critical COVID-19 (intensive care unit [ICU] admission required).

Participants ranged from 31 to 74 years of age, with 60.3 presenting the median value. Participants presented a diverse array of comorbidities, including cancer (n = 8), renal disease (n = 4), heart ailments (n = 9), hypertension (n = 9), and diabetes (n = 9).

From preserved PCMBs, selective staining was used to identify and isolate live CD3+ single cells, single positive CD4CD8+, and CD4+CD8, excluding all double positive and negative CD cells from further experiments. Gating and Boolean analyses with PBMC stimulation were used to identify and study AIM assay combinations of antigen-specific (in this case, SARS-CoV-2 Wu-1 strain) T-cell frequency.

Intracellular cytokine staining (ICS) was used to identify and quantify the production of cytokines following SARS-CoV-2 stimulation. Genomic DNA isolated from PBMCs was used for T-cell Receptor Sequencing (TCR-Seq), a method used to identify and track specific T cells and their clones. TCR-Seq was carried out independently for bulk repertoire analyses (bulk TCR-Seq) and antibody feature barcode library generation (single-cell TCR-Seq).

The library generated above was used for CD4+ and CD8+ assignments using unique molecular identifier (UMI) counts. SARS-CoV-2 Wu-1 strain-transfected Cos-7 cells were used to evaluate the specificity of assigned CD8+ T cells.

“To test the CD4+ T cell-origin candidate TCRs, engineered reporter T cells were generated by transduction of autologous CD4+ T cells using lentiviruses with candidate-paired TCR expression cassettes.”

Finally, next-generation Human Leukocyte Antigen (HLA) Typing was carried out to identify class 1 and class 2 allotypes.

Study findings

Results from the above tests revealed divergent vaccine response kinetics, both between CD4+ and CD8+ TS cells and between individuals with differing COVID-19 disease severity. “In persons with previous SARS-CoV-2 infection, mRNA vaccines induced profound, albeit variable, expansion of preexisting circulating TM cell clones.” These results were amplified based on the number of mRNA vaccines received following COVID-19 disease, with the first two vaccine doses observed to augment S-reactive clonotypic diversity in the blood, resulting in substantial expansion in CD8+ TS clonotypes.

A similar expansion in CD8+ TS clonotypes has been reported in COVID-19-naïve individuals following their first vaccination dose, albeit to a much lower extent. This study further revealed that while not as substantial as CD8+ TS clonotype expansion, CD4+ clonotypes also expanded following the second vaccination dose.

“Our work used AIM with statistical filters to assign S specificity to individual TCRs. We recovered thousands of paired-chain TCRαβ sequences with S reactivity to augment public database for characterizing T cell responses to vaccines and reveal both α-chain and β-chain sequence features contributing to epitope specificity. Further research is required to determine how the phenotype, durability and distribution of CD4+ and CD8+ T cells elicited by hybrid exposure compare to responses elicited by infection or vaccination alone.”

Journal reference:
  • Ford, E. S., Jing, L., Laing, K. J., Sholukh, A. M., Bossard, E. L., Xie, H., Pulliam, T. H., Jani, S., Selke, S., Burrow, C. J., McClurkan, C. L., Wald, A., Greninger, A. L., Holbrook, M. R., Eaton, B., Eudy, E., Murphy, M., Postnikova, E., Robins, H. S., . . . Koelle, D. M. (2023). Repeated mRNA vaccination sequentially boosts SARS-CoV-2-specific CD8+ T cells in persons with previous COVID-19. Nature Immunology, 1-12. DOI –,
Hugo Francisco de Souza

Written by

Hugo Francisco de Souza

Hugo Francisco de Souza is a scientific writer based in Bangalore, Karnataka, India. His academic passions lie in biogeography, evolutionary biology, and herpetology. He is currently pursuing his Ph.D. from the Centre for Ecological Sciences, Indian Institute of Science, where he studies the origins, dispersal, and speciation of wetland-associated snakes. Hugo has received, amongst others, the DST-INSPIRE fellowship for his doctoral research and the Gold Medal from Pondicherry University for academic excellence during his Masters. His research has been published in high-impact peer-reviewed journals, including PLOS Neglected Tropical Diseases and Systematic Biology. When not working or writing, Hugo can be found consuming copious amounts of anime and manga, composing and making music with his bass guitar, shredding trails on his MTB, playing video games (he prefers the term ‘gaming’), or tinkering with all things tech.


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  1. Doug Foley Doug Foley Canada says:

    A European Journal of Clinical Investigation Peer Reviewed Paper titled 'Effectiveness of a fourth SARS-CoV-2 vaccine dose in previously infected individuals from Austria'. A very readable paper.

  2. Anne Barschall Anne Barschall United States says:

    I keep hearing that immunity lasts only 6 months from these vaccines.  That's why I've had 7 of them.  Is this article saying something different?  If I've had 7 does that mean that i don't have to worry about getting COVID?

The opinions expressed here are the views of the writer and do not necessarily reflect the views and opinions of News Medical.
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