RESUMO
BACKGROUND: SARS-CoV-2 has triggered a pandemic and contributes to long-lasting morbidity. Several studies have investigated immediate cellular and humoral immune responses during acute infection. However, little is known about long-term effects of COVID-19 on the immune system. METHODS: We performed a longitudinal investigation of cellular and humoral immune parameters in 106 non-vaccinated subjects ten weeks (10 w) and ten months (10 m) after their first SARS-CoV-2 infection. Peripheral blood immune cells were analyzed by multiparametric flow cytometry, serum cytokines were examined by multiplex technology. Antibodies specific for the Spike protein (S), the receptor-binding domain (RBD) and the nucleocapsid protein (NC) were determined. All parameters measured 10 w and 10 m after infection were compared with those of a matched, noninfected control group (n = 98). RESULTS: Whole blood flow cytometric analyses revealed that 10 m after COVID-19, convalescent patients compared to controls had reduced absolute granulocyte, monocyte, and lymphocyte counts, involving T, B, and NK cells, in particular CD3+CD45RA+CD62L+CD31+ recent thymic emigrant T cells and non-class-switched CD19+IgD+CD27+ memory B cells. Cellular changes were associated with a reversal from Th1- to Th2-dominated serum cytokine patterns. Strong declines of NC- and S-specific antibody levels were associated with younger age (by 10.3 years, p < .01) and fewer CD3-CD56+ NK and CD19+CD27+ B memory cells. Changes of T-cell subsets at 10 m such as normalization of effector and Treg numbers, decline of RTE, and increase of central memory T cell numbers were independent of antibody decline pattern. CONCLUSIONS: COVID-19 causes long-term reduction of innate and adaptive immune cells which is associated with a Th2 serum cytokine profile. This may provide an immunological mechanism for long-term sequelae after COVID-19.
RESUMO
BACKGROUND: COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has now become endemic and is currently one of the important respiratory virus infections regularly affecting mankind. The assessment of immunity against SARS-CoV-2 and its variants is important for guiding active and passive immunization and SARS-CoV-2-specific treatment strategies. METHODS: We here devised a novel flow cytometry-based diagnostic platform for the assessment of immunity against cell-bound virus antigens. This platform is based on a collection of HEK-293T cell lines which, as exemplified in our study, stably express the receptor-binding domains (RBDs) of the SARS-CoV-2 S-proteins of eight major SARS-CoV-2 variants, ranging from Wuhan-Hu-1 to Omicron. RESULTS: RBD-expressing cell lines stably display comparable levels of RBD on the surface of HEK-293T cells, as shown with anti-FLAG-tag antibodies directed against a N-terminally introduced 3x-FLAG sequence while the functionality of RBD was proven by ACE2 binding. We exemplify the usefulness and specificity of the cell-based test by direct binding of IgG and IgA antibodies of SARS-CoV-2-exposed and/or vaccinated individuals in which the assay shows a wide linear performance range both at very low and very high serum antibody concentrations. In another application, i.e., antibody adsorption studies, the test proved to be a powerful tool for measuring the ratios of individual variant-specific antibodies. CONCLUSION: We have established a toolbox for measuring SARS-CoV-2-specific immunity against cell-bound virus antigens, which may be considered as an important addition to the armamentarium of SARS-CoV-2-specific diagnostic tests, allowing flexible and quick adaptation to new variants of concern.