Introduction
The COVID-19 pandemic has led to devastating health outcomes with the death toll exceeding one million cases by the end of 2020. Furthermore, the global lockdowns impeded social and commercial interactions, with psychological and economic impacts on the public1. Vaccines are currently in the final stages of approval or have already been authorized under emergency use authorization2. However, there is still limited data on long-term immunity against SARS-CoV-23. Studying long term immunity is essential to aid decision making for reopening and easing social distancing, assess the need for booster doses of vaccines, and estimate the potential of reinfections4,5. Focusing mainly on antibody-based assays as readouts for long-term immunity may be strongly impeded due to lack of consistency in circulating antibody levels regardless of disease severity. Indeed, there are conflicting results about the longevity of antibody response in SARS-CoV-2 infected individuals6–10, suggesting that long-lived plasma cells may not have been efficiently established in a proportion of COVID-19 patients. Another study found that about 20 % of German SARS-CoV-2 PCR-positive patients with mild to moderate symptoms did not have detectable spike-specific antibodies when tested 3 weeks or longer after infection11. In line with these findings, Long et al. reported that 40 % of non-symptomatic patients, as well as 12.9% with symptoms, became seronegative with no detectable antibodies in the blood eight weeks after discharge from hospital12. Therefore, it is of uttermost importance to develop and test other diagnostic assays to study the long-term immunity against SARS-CoV-2.
For diagnostic assays, the right target, the test accuracy, and minimizing false-negative and false-positive outcomes are the major challenges13. One potential approach that can help to improve the detection of long-term immunity is the analysis of SARS-CoV-2-specific memory T and B cell formation. We and others have reported on the detection of a strong SARS-CoV-2 specific memory T cell response in immune-competent and immunosuppressed patients14–18. Similar studies of B cell-related immunity have so far focused mainly on characterizing antibody specificities19–24. Other studies identified and isolated SARS-COV-2 B cells to generate neutralizing antibodies or to characterize the B cell receptor usage20–22. BMEMORY cells can remain for decades or potentially lifelong, located within lymph nodes, spleen, bone marrow, and the lung, or circulate in the blood25–28. Upon reinfection, they become re-activated by encountering the same antigen and immediately start to proliferate and differentiate into plasma cells secreting neutralizing antibodies. Thus, quantifying BMEMORY cell levels by a reliable assay may be used as an indicator of long-term immunity in convalescent patients. Although the formation of BMEMORY cells was described, controlled studies comparing SARS-COV-2 specific memory B cell formation in unexposed individuals and COVID 19 patients are limited. To our knowledge, only studies with Australian and US-American cohorts assessed the long-term immunity through characterization of the BMEMORY cells for more than 6 months29,30. Since the reports on duration of S1-specific antibody detection upon SARS-CoV-2 resolution are conflicting, several studies on antibody-producing BMEMORY cells performed within different ethnic origin populations are required. In this work, we focus on detecting SARS-CoV2 spike (S)-protein specific memory B cells (BMEMORY) in a well characterized cohort of central European COVID-19 patients as compared to unexposed controls. Furthermore, we analysed the sustainability for several months after infection as a specific long-term marker of adaptive immunity.