In a recent study posted to the bioRxiv* preprint server, a team of researchers examined single clones of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific memory B cells (MBCs) to understand their adaptability and fate.

Background

Memory B cells are formed when B cells encounter antigens and undergo somatic hypermutation in lymphoid germinal centers to give rise to MBCs and antibody-secreting plasma cells. MBCs can persist in the body for long periods and differentiate into plasma cells when they reencounter antigens from the original pathogen or its variants.

Memory B cells express high levels of complement receptor 2 (CD21) and tumor necrosis factor superfamily member CD27 on their surface. Class-switched subsets of B cells include CD21^–CD27^– atypical and CD21^–CD27⁺ activated MBCs. CD21^–CD27⁺ activated MBCs are thought to be predisposed to plasma cell differentiation, while atypical MBCs are thought to have an extrafollicular origin, express transcription factor T-bet and several inhibitory coreceptors, and are associated with autoimmune diseases and chronic infections.

These two class-switched memory B cell subsets have been observed after some coronavirus disease 2019 (COVID-19) vaccinations and SARS-CoV-2 infections. Examining the fates and plasticity of the various memory B cell subsets would help understand their role in protective immune responses upon antigen re-exposure.

About the study

The present study used a longitudinal cohort to analyze antigen-specific MBCs from patients with acute COVID-19. Participants with SARS-CoV-2 infection confirmed by a positive reverse transcriptase polymerase chain reaction (RT-PCR) test and exhibited symptoms were included in the study. Blood and serum samples were collected from the participants at the time of acute infection and six and 12 weeks after infection.

The study also included a tonsil cohort to understand the variations in SARS-CoV-2-specific MBCs in a peripheral lymphoid organ. Tonsil and blood samples were procured from individuals undergoing a tonsillectomy. The tonsil cohort only included samples from participants who were vaccinated and had no SARS-CoV-2 infections or had recovered from the infection.

The researchers used biotinylated SARS-CoV-2 spike and receptor binding domain proteins to create multimer stains to analyze antigen-specific MBCs using spectral flow cytometry. Fluorescence-activated cell-sorting was used to sort SARS-CoV-2-specific and non-SARS-CoV-2-specific MBCs, which were then used for single-cell RNA sequencing (scRNA-seq), barcoding, and B cell receptor sequencing.

Flow cytometry was also used to characterize the immunoglobulin isotypes of the SARS-CoV-2-specific MBCs. Uniform manifold approximation and projection (UMAP) plots and unsupervised phenograph clustering were used to sort the MBCs into IgM+, IgG+, and IgA+ cells.

Enzyme-linked immunosorbent assays (ELISA) specific for the SARS-CoV-2 S1 subunit protein were used to measure antibodies against SARS-CoV-2 in all the patients. A bead-based multiplexed immunoassay was also used to measure the immunoglobulin responses against various SARS-CoV-2 proteins for the patients in the tonsil cohort.

The acute COVID-19 patients were studied between April and September 2020. The 12-month follow-up phase occurred between April and September 2021. The availability of COVID-19 vaccines resulted in 35 of the 65 participants getting vaccinated during the study, which allowed the researchers to observe MBC response to antigen re-exposure.

Results

The results reported that single MBC clones exhibited diverse phenotypic and functional characteristics in COVID-19 patients with immune memory and after SARS-CoV-2 vaccinations. Single MBC clones differentiated into activated (CD21^–CD27⁺), atypical (CD21^–CD27^–), or resting (CD21⁺CD27⁺) MBCs upon vaccination.

Following SARS-CoV-2 infections or COVID-19 vaccinations, the number of activated MBCs specific for the SARS-CoV-2 spike protein increased and then declined rapidly, while the number of atypical MBCs remained stable, and resting MBCs increased. Activated and atypical MBCs were predominant in the early period of SARS-CoV-2 infection or vaccination, but the number of resting MBCs was higher at the six and 12-month follow-ups.

The study found that SARS-CoV-2-specific atypical MBCs were transcriptionally similar to atypical MBCs from autoimmune diseases. Additionally, studying the tonsil cohort revealed that peripheral lymphoid organs were rich in SARS-CoV-2-specific resting MBCs, and had low levels of transcription factor T-bet. The resting MBCs in the tonsils also exhibited markers of tissue residency.

Conclusions

To summarize, the study investigated the plasticity and fate of single clones of MBCs specific to SARS-CoV-2 to understand the variation in phenotype and function of the MBC subsets. The results demonstrated that single MBC clones from COVID-19 patients gave rise to activated, atypical, and resting MBCs.

During acute infection and immediately after COVID-19 vaccinations, the number of activated MBCs and, to a lesser degree, the number of atypical MBCs peaked. The activated MBCs declined rapidly with time, while the atypical MBCs remained stable, and the resting MBCs increased in number. Peripheral lymphoid organs such as the tonsils were rich in resting MBCs exhibiting tissue-residing markers.

The study indicated that SARS-CoV-2 infections and vaccines induced stable, long-lived immune memory capable of mounting an immune response upon antigen re-exposure.

*Important notice

bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.

Written by

Dr. Chinta Sidharthan

Chinta Sidharthan is a writer based in Bangalore, India. Her academic background is in evolutionary biology and genetics, and she has extensive experience in scientific research, teaching, science writing, and herpetology. Chinta holds a Ph.D. in evolutionary biology from the Indian Institute of Science and is passionate about science education, writing, animals, wildlife, and conservation. For her doctoral research, she explored the origins and diversification of blindsnakes in India, as a part of which she did extensive fieldwork in the jungles of southern India. She has received the Canadian Governor General’s bronze medal and Bangalore University gold medal for academic excellence and published her research in high-impact journals.