ABSTRACT
The majority of infections with SARS-CoV-2 (SCoV2) are asymptomatic or mild without the necessity of hospitalization. It is of outmost importance to reveal if these patients develop an antibody response against SCoV2 and to define which antibodies confer virus neutralization. We hence conducted a comprehensive serological survey of 49 patients with a mild course of disease and quantified neutralizing antibody responses against authentic SCoV2 employing human cells as targets. Four patients (8%), even though symptomatic, did not develop antibodies against SCoV2 and two other sera (4%) were only positive in one of the serological assays employed. For the remainder, antibody response against the S-protein correlated with serum neutralization whereas antibodies against the nucleocapsid were poor predictors of virus neutralization. Only six sera (12%) could be classified as highly neutralizing. Furthermore, sera from several individuals with fairly high antibody levels had only poor neutralizing activity. In addition, our data suggest that antibodies against the seasonal coronavirus 229E contribute to SCoV2 neutralization. Altogether, we show that there is a wide breadth of antibody responses against SCoV2 in patients that differentially correlate with virus neutralization. This highlights the difficulty to define reliable surrogate markers for immunity against SCoV2.
Subject(s)
Severe Acute Respiratory SyndromeABSTRACT
BO_SCPLOWACKGROUNDC_SCPLOWSeroreactivity against human endemic coronaviruses has been linked to disease severity after SARS-CoV-2 infection. Assays that are capable of concomitantly detecting antibodies against endemic coronaviridae such as OC43, 229E, NL63, and SARS-CoV-2 may help to elucidate this question. We set up a platform for serum-screening and developed a bead-based Western blot system, namely DigiWest, capable of running hundreds of assays using microgram amounts of protein prepared directly from different viruses. MO_SCPLOWETHODSC_SCPLOWThe parallelized and miniaturised DigiWest assay was adapted for detecting antibodies using whole protein extract prepared from isolated SARS-CoV-2 virus particles. After characterisation and optimization of the newly established test, whole virus lysates of OC43, 229E, and NL63 were integrated into the system. RO_SCPLOWESULTSC_SCPLOWThe DigiWest-based immunoassay system for detection of SARS-CoV-2 specific antibodies shows a sensitivity of 87.2 % and diagnostic specificity of 100 %. Concordance analysis with the SARS-CoV-2 immunoassays available by Roche, Siemens, and Euroimmun indicates a comparable assay performance (Cohens Kappa ranging from 0.8799-0.9429). In the multiplexed assay, antibodies against the endemic coronaviruses OC43, 229E, and NL63 were detected, displaying a high incidence of seroreactivity against these coronaviruses. CO_SCPLOWONCLUSIONC_SCPLOWThe DigiWest-based immunoassay, which uses authentic antigens from isolated virus particles, is capable of detecting individual serum responses against SARS-CoV-2 with high specificity and sensitivity in one multiplexed assay. It shows high concordance with other commercially available serologic assays. The DigiWest approach enables a concomitant detection of antibodies against different endemic coronaviruses and will help to elucidate the role of these possibly cross-reactive antibodies.
Subject(s)
COVID-19ABSTRACT
Given the importance of the humoral immune response to SARS-CoV-2 as a global benchmark for immunity, a detailed analysis is needed to (i) monitor seroconversion in the general population, (ii) understand manifestation and progression of the disease, and (iii) predict the outcome of vaccine development. Currently available serological assays utilize single analyte technologies such as ELISA to measure antibodies against SARS-CoV-2 antigens including spike (S) or nucleocapsid (N) protein. To measure individual antibody (IgG and IgA) responses against SARS-CoV-2 and the endemic human coronaviruses (hCoVs) NL63, 229E, OC43, and HKU1, we developed a multiplexed immunoassay (CoVi-plex), for which we included S and N proteins of these coronaviruses in an expanded antigen panel. Compared to commercial in vitro diagnostic (IVD) tests our CoVi-plex achieved the highest sensitivity and specificity when analyzing 310 SARS-CoV-2 infected and 866 uninfected individuals. Simultaneously we see high IgG responses against hCoVs throughout all samples, whereas no consistent cross reactive IgG response patterns can be defined. In summary, our CoVi-plex is highly suited to monitor vaccination studies and will facilitate epidemiologic screenings for the humoral immunity toward pandemic as well as endemic coronaviruses.
Subject(s)
Severe Acute Respiratory SyndromeABSTRACT
The SARS-CoV-2 pandemic calls for the rapid development of diagnostic, preventive, and therapeutic approaches. CD4+ and CD8+ T cell-mediated immunity is central for control of and protection from viral infections[1-3]. A prerequisite to characterize T-cell immunity, but also for the development of vaccines and immunotherapies, is the identification of the exact viral T-cell epitopes presented on human leukocyte antigens (HLA)[2-8]. This is the first work identifying and characterizing SARS-CoV-2-specific and cross-reactive HLA class I and HLA-DR T-cell epitopes in SARS-CoV-2 convalescents (n = 180) as well as unexposed individuals (n = 185) and confirming their relevance for immunity and COVID-19 disease course. SARS-CoV-2-specific T-cell epitopes enabled detection of post-infectious T-cell immunity, even in seronegative convalescents. Cross-reactive SARS-CoV-2 T-cell epitopes revealed preexisting T-cell responses in 81% of unexposed individuals, and validation of similarity to common cold human coronaviruses provided a functional basis for postulated heterologous immunity[9] in SARS-CoV-2 infection[10,11]. Intensity of T-cell responses and recognition rate of T-cell epitopes was significantly higher in the convalescent donors compared to unexposed individuals, suggesting that not only expansion, but also diversity spread of SARS-CoV-2 T-cell responses occur upon active infection. Whereas anti-SARS-CoV-2 antibody levels were associated with severity of symptoms in our SARS-CoV-2 donors, intensity of T-cell responses did not negatively affect COVID-19 severity. Rather, diversity of SARS-CoV-2 T-cell responses was increased in case of mild symptoms of COVID-19, providing evidence that development of immunity requires recognition of multiple SARS-CoV-2 epitopes. Together, the specific and cross-reactive SARS-CoV-2 T-cell epitopes identified in this work enable the identification of heterologous and post-infectious T-cell immunity and facilitate the development of diagnostic, preventive, and therapeutic measures for COVID-19.