B-Cell Receptor Features and Database Establishment in Recovered COVID-19 Patients by Combining 5'-RACE with PacBio Sequencing.

BACKGROUND: Antibodies induced by viral infection can not only prevent subsequent virus infection, but can also mediate pathological injury following infection. Therefore, understanding the B-cell receptor (BCR) repertoire of either specific neutralizing or pathological antibodies from patients convalescing from Coronavirus disease 2019 (COVID-19) infection is of benefit for the preparation of therapeutic or preventive antibodies, and may provide insight into the mechanisms of COVID-19 pathological injury. METHODS: In this study, we used a molecular approach of combining 5' Rapid Amplification of cDNA Ends (5'-RACE) with PacBio sequencing to analyze the BCR repertoire of all 5 IgH and 2 IgL genes in B-cells harvested from 35 convalescent patients after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. RESULTS: We observed numerous BCR clonotypes within most COVID-19 patients, but not in healthy controls, which validates the association of the disease with a prototypical immune response. In addition, many clonotypes were found to be frequently shared between different patients or different classes of antibodies. CONCLUSIONS: These convergent clonotypes provide a resource to identify potential therapeutic/prophylactic antibodies, or identify antibodies associated with pathological effects following infection with SARS-CoV-2.

A Rapid Method for Detection of Antigen-Specific B Cells.

The global SARS-CoV-2 pandemic has united the efforts of many scientists all over the world to develop wet-lab techniques and computational approaches aimed at the identification of antigen-specific T and B cells. The latter provide specific humoral immunity that is essential for the survival of COVID-19 patients, and vaccine development has essentially been based on these cells. Here, we implemented an approach that integrates the sorting of antigen-specific B cells and B-cell receptor mRNA sequencing (BCR-seq), followed by computational analysis. This rapid and cost-efficient method allowed us to identify antigen-specific B cells in the peripheral blood of patients with severe COVID-19 disease. Subsequently, specific BCRs were extracted, cloned, and produced as full antibodies. We confirmed their reactivity toward the spike RBD domain. Such an approach can be effective for the monitoring and identification of B cells participating in an individual immune response.

Proof-of-Concept Analysis of B Cell Receptor Repertoire in COVID-19 Patients Undergoing ECMO by Single-Cell V(D)J and Gene Expression Sequencing.

SARS-CoV-2, which causes COVID-19, has altered human activities all over the world and has become a global hazard to public health. Despite considerable advancements in pandemic containment techniques, in which vaccination played a key role, COVID-19 remains a global threat, particularly for frail patients and unvaccinated individuals, who may be more susceptible to developing ARDS. Several studies reported that patients with COVID-19-related ARDS who were treated with ECMO had a similar survival rate to those with COVID-19-unrelated ARDS. In order to shed light on the potential mechanisms underlying the COVID-19 infection, we conducted this proof-of-concept study using single-cell V(D)J and gene expression sequencing of B cells to examine the dynamic changes in the transcriptomic BCR repertoire present in patients with COVID-19 at various stages. We compared a recovered and a deceased COVID-19 patient supported by ECMO with one COVID-19-recovered patient who did not receive ECMO treatment and one healthy subject who had never been infected previously. Our analysis revealed a downregulation of FXYD, HLA-DRB1, and RPS20 in memory B cells; MTATP8 and HLA-DQA1 in naïve cells; RPS4Y1 in activated B cells; and IGHV3-73 in plasma cells in COVID-19 patients. We further described an increased ratio of IgA + IgG to IgD + IgM, suggestive of an intensive memory antibody response, in the COVID ECMO D patient. Finally, we assessed a V(D)J rearrangement of heavy chain IgHV3, IGHJ4, and IGHD3/IGHD2 families in COVID-19 patients regardless of the severity of the disease.

Profiling the B cell immune response elicited by vaccination against the respiratory virus SARS-CoV-2.

B cells play a fundamental role in host defenses against viral infections. Profiling the B cell response elicited by SARS-CoV-2 vaccination, including the generation and persistence of antigen-specific memory B cells, is essential for improving the knowledge of vaccine immune responsiveness, beyond the antibody response. mRNA-based vaccines have shown to induce a robust class-switched memory B cell response that persists overtime and is boosted by further vaccine administration, suggesting that memory B cells are critical in driving a recall response upon re-exposure to SARS-CoV-2 antigens. Here, we focus on the role of the B cell response in the context of SARS-CoV-2 vaccination, offering an overview of the different technologies that can be used to identify spike-specific B cells, characterize their phenotype using machine learning approaches, measure their capacity to reactivate following antigen encounter, and tracking the maturation of the B cell receptor antigenic affinity.

Comparative global B cell receptor repertoire difference induced by SARS-CoV-2 infection or vaccination via single-cell V(D)J sequencing.

Dynamic changes of the paired heavy and light chain B cell receptor (BCR) repertoire provide an essential insight into understanding the humoral immune response post-SARS-CoV-2 infection and vaccination. However, differences between the endogenous paired BCR repertoire kinetics in SARS-CoV-2 infection and previously recovered/naïve subjects treated with the inactivated vaccine remain largely unknown. We performed single-cell V(D)J sequencing of B cells from six healthy donors with three shots of inactivated SARS-CoV-2 vaccine (BBIBP-CorV), five people who received the BBIBP-CorV vaccine after having recovered from COVID-19, five unvaccinated COVID-19 recovered patients and then integrated with public data of B cells from four SARS-CoV-2-infected subjects. We discovered that BCR variable (V) genes were more prominently used in the SARS-CoV-2 exposed groups (both in the group with active infection and in the group that had recovered) than in the vaccinated groups. The VH gene that expanded the most after SARS-CoV-2 infection was IGHV3-33, while IGHV3-23 in the vaccinated groups. SARS-CoV-2-infected group enhanced more BCR clonal expansion and somatic hypermutation than the vaccinated healthy group. A small proportion of public clonotypes were shared between the SARS-CoV-2 infected, vaccinated healthy, and recovered groups. Moreover, several public antibodies had been identified against SARS-CoV-2 spike protein. We comprehensively characterize the paired heavy and light chain BCR repertoire from SARS-CoV-2 infection to vaccination, providing further guidance for the development of the next-generation precision vaccine.

Global characterization of B cell receptor repertoire in COVID-19 patients by single-cell V(D)J sequencing.

The world is facing a pandemic of Corona Virus Disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Adaptive immune responses are essential for SARS-CoV-2 virus clearance. Although a large body of studies have been conducted to investigate the immune mechanism in COVID-19 patients, we still lack a comprehensive understanding of the BCR repertoire in patients. In this study, we used the single-cell V(D)J sequencing to characterize the BCR repertoire across convalescent COVID-19 patients. We observed that the BCR diversity was significantly reduced in disease compared with healthy controls. And BCRs tend to skew toward different V gene segments in COVID-19 and healthy controls. The CDR3 sequences of heavy chain in clonal BCRs in patients were more convergent than that in healthy controls. In addition, we discovered increased IgG and IgA isotypes in the disease, including IgG1, IgG3 and IgA1. In all clonal BCRs, IgG isotypes had the most frequent class switch recombination events and the highest somatic hypermutation rate, especially IgG3. Moreover, we found that an IgG3 cluster from different clonal groups had the same IGHV, IGHJ and CDR3 sequences (IGHV4-4-CARLANTNQFYDSSSYLNAMDVW-IGHJ6). Overall, our study provides a comprehensive characterization of the BCR repertoire in COVID-19 patients, which contributes to the understanding of the mechanism for the immune response to SARS-CoV-2 infection.