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The world is entering a new era of the COVID-19 pandemic in which there is an increasing call for reliable antibody testing. To support decision making on the deployment of serology for either population screening or diagnostics, we present a detailed comparison of serological COVID-19 assays. We show that among the selected assays there is a wide diversity in assay performance in different scenarios and when correlated to virus neutralizing antibodies. The Wantai ELISA detecting total immunoglobulins against the receptor binding domain of SARS CoV-2, has the best overall characteristics to detect functional antibodies in different stages and severity of disease, including the potential to set a cut-off indicating the presence of protective antibodies. The large variety of available serological assays requires proper assay validation before deciding on deployment of assays for specific applications., SARS-CoV-2 is causing a global pandemic in which the implementation of serology can support decision making in different scenarios. Here, the authors compare the outcome of eight commercially available assays to virus neutralization and discuss their use in diagnostics and exposure assessment of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of coronavirus disease 2019 (COVID-19). Little is known about the interplay between preexisting immunity to endemic seasonal coronaviruses and the development of a SARS-CoV-2–specific IgG response. We investigated the kinetics, breadth, magnitude, and level of cross-reactivity of IgG antibodies against SARS-CoV-2 and heterologous seasonal and epidemic coronaviruses at the clonal level in patients with mild or severe COVID-19 as well as in disease control patients. We assessed antibody reactivity to nucleocapsid and spike antigens and correlated this IgG response to SARS-CoV-2 neutralization. Patients with COVID-19 mounted a mostly type-specific SARS-CoV-2 response. Additionally, IgG clones directed against a seasonal coronavirus were boosted in patients with severe COVID-19. These boosted clones showed limited cross-reactivity and did not neutralize SARS-CoV-2. These findings indicate a boost of poorly protective CoV-specific antibodies in patients with COVID-19 that correlated with disease severity, revealing “original antigenic sin.”

A new coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has recently emerged to cause a human pandemic. Although molecular diagnostic tests were rapidly developed, serologic assays are still lacking, yet urgently needed. Validated serologic assays are needed for contact tracing, identifying the viral reservoir, and epidemiologic studies. We developed serologic assays for detection of SARS-CoV-2 neutralizing, spike protein–specific, and nucleocapsid-specific antibodies. Using serum samples from patients with PCR-confirmed SARS-CoV-2 infections, other coronaviruses, or other respiratory pathogenic infections, we validated and tested various antigens in different in-house and commercial ELISAs. We demonstrated that most PCR-confirmed SARS-CoV-2–infected persons seroconverted by 2 weeks after disease onset. We found that commercial S1 IgG or IgA ELISAs were of lower specificity, and sensitivity varied between the 2 assays; the IgA ELISA showed higher sensitivity. Overall, the validated assays described can be instrumental for detection of SARS-CoV-2–specific antibodies for diagnostic, seroepidemiologic, and vaccine evaluation studies.

Middle East respiratory syndrome coronavirus (MERS-CoV) infections in humans can cause asymptomatic to fatal lower respiratory lung disease. Despite posing a probable risk for virus transmission, asymptomatic to mild infections can go unnoticed; a lack of seroconversion among some PCR-confirmed cases has been reported. We found that a MERS-CoV spike S1 protein-based ELISA, routinely used in surveillance studies, showed low sensitivity in detecting infections among PCR-confirmed patients with mild clinical symptoms and cross-reactivity of human coronavirus OC43-positive serum samples. Using in-house S1 ELISA and protein microarray, we demonstrate that most PCR-confirmed MERS-CoV case-patients with mild infections seroconverted; nonetheless, some of these samples did not have detectable levels of virus-neutralizing antibodies. The use of a sensitive and specific serologic S1-based assay can be instrumental in the accurate estimation of MERS-CoV prevalence.

Optimal vaccines are needed for sustained suppression of SARS‐CoV‐2 and other novel coronaviruses. Here, we developed a recombinant type 5 adenovirus vector encoding the gene for the SARS‐CoV‐2 S1 subunit antigen (Ad5.SARS‐CoV‐2‐S1) for COVID‐19 immunization and evaluated its immunogenicity in mice. A single immunization with Ad5.SARS‐CoV‐2‐S1 via S.C. injection or I.N delivery induced robust antibody and cellular immune responses. Vaccination elicited significant S1‐specific IgG, IgG1, and IgG2a endpoint titers as early as 2 weeks, and the induced antibodies were long lasting. I.N. and S.C. administration of Ad5.SARS‐CoV‐2‐S1 produced S1‐specific GC B cells in cervical and axillary LNs, respectively. Moreover, I.N. and S.C. immunization evoked significantly greater antigen‐specific T‐cell responses compared to unimmunized control groups with indications that S.C. injection was more effective than I.N. delivery in eliciting cellular immune responses. Mice vaccinated by either route demonstrated significantly increased virus‐specific neutralization antibodies on weeks 8 and 12 compared to control groups, as well as BM antibody forming cells (AFC), indicative of long‐term immunity. Thus, this Ad5‐vectored SARS‐CoV‐2 vaccine candidate showed promising immunogenicity following delivery to mice by S.C. and I.N. routes of administration, supporting the further development of Ad‐based vaccines against COVID‐19 and other infectious diseases for sustainable global immunization programs., Vaccine development against SARS‐CoV‐2 addresses the ongoing challenges posed by this pathogen and ensures preparedness for future outbreaks of coronaviruses. Subcutaneous or intranasal administration of Ad5.SARS‐CoV‐2 S1 results in multifaceted immune responses in mice, including GCs and long‐lived antibody forming cells, representing an attractive COVID‐19 vaccination strategy.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic is continuing to disrupt personal lives, global healthcare systems, and economies. Hence, there is an urgent need for a vaccine that prevents viral infection, transmission, and disease. Here, we present a two-component protein-based nanoparticle vaccine that displays multiple copies of the SARS-CoV-2 spike protein. Immunization studies show that this vaccine induces potent neutralizing antibody responses in mice, rabbits, and cynomolgus macaques. The vaccine-induced immunity protects macaques against a high-dose challenge, resulting in strongly reduced viral infection and replication in the upper and lower airways. These nanoparticles are a promising vaccine candidate to curtail the SARS-CoV-2 pandemic., Graphical Abstract, Brouwer et al. present preclinical evidence in support of a COVID-19 vaccine candidate, designed as a self-assembling two-component protein nanoparticle displaying multiple copies of the SARS-CoV-2 spike protein, which induces strong neutralizing antibody responses and protects from high-dose SARS-CoV-2 challenge.

Severe acquired respiratory syndrome coronavirus-2 (SARS-CoV-2) is the cause of coronavirus disease (COVID-19). In severe COVID-19 cases, higher antibody titers against seasonal coronaviruses have been observed than in mild cases. To investigate antibody cross-reactivity as potential explanation for severe disease, we determined the kinetics, breadth, magnitude and level of cross-reactivity of IgG against SARS-CoV-2 and seasonal CoV nucleocapsid and spike from 17 severe COVID-19 cases at the clonal level. Although patients mounted a mostly type-specific SARS-CoV-2 response, B-cell clones directed against seasonal CoV dominated and strongly increased over time. Seasonal CoV IgG responses that did not neutralize SARS-CoV-2 were boosted well beyond detectable cross-reactivity, particularly for HCoV-OC43 spike. These findings support a back-boost of poorly protective coronavirus-specific antibodies in severe COVID-19 patients that may negatively impact de novo SARS-CoV-2 immunity, reminiscent of original antigenic sin.

Effective clinical intervention strategies for COVID-19 are urgently needed. Although several clinical trials have evaluated the use of convalescent plasma containing virus-neutralizing antibodies, the effectiveness has not been proven. We show that hamsters treated with a high dose of human convalescent plasma or a monoclonal antibody were protected against weight loss showing reduced pneumonia and pulmonary virus replication compared to control animals. However, a ten-fold lower dose of convalescent plasma showed no protective effect. Thus, variable and relatively low levels of virus neutralizing antibodies in convalescent plasma may limit their use for effective antiviral therapy, favouring concentrated, purified (monoclonal) antibodies.

Understanding the coronavirus (CoV) antibody landscape in relation to disease and susceptibility is critical for modelling of steps in the next phase during the current covid-19 pandemic. In March 2020, during the first month of the epidemic in The Netherlands, we performed cross sectional studies at two time points amongst patients of the Erasmus Medical Centre in Rotterdam, to assess the presence of antibodies against seasonal human coronaviruses (OC43, 229E, NL63, HKU1), emerging zoonotic coronaviruses (SARS, MERS) and SARS-CoV-2 in nine different age groups. We observed minimal SARS-CoV-2 reactivity early March (0.7% of sera), increasing to 3.0%, four weeks later, suggesting probably undetected cases during this early phase of the epidemic. Antibody responses were mostly coronavirus species specific at young age, but possible cross-reactivity between human seasonal CoVs was observed with increasing age.

Structured abstract for full paperBackgroundAfter recovery from COVID-19, most patients have anti-SARS-CoV-2 neutralizing antibodies. Their convalescent plasma could be an inexpensive and widely available treatment for COVID-19.MethodsThe Convalescent-plasma-for-COVID (ConCOVID) study was a randomized trial comparing convalescent plasma with standard of care therapy in patients hospitalized for COVID-19 in the Netherlands. Patients were randomized 1:1 and received 300ml of plasma with anti-SARS-CoV-2 neutralizing antibody titers of at least 1:80. The primary endpoint was day-60 mortality and key secondary endpoints were hospital stay and WHO 8-point disease severity scale improvement on day 15.ResultsThe trial was halted prematurely after 86 patients were enrolled. Although symptomatic for only 10 days (IQR 6-15) at the time of inclusion, 53 of 66 patients tested had anti-SARS-CoV-2 antibodies at baseline. A SARS-CoV-2 plaque reduction neutralization test showed neutralizing antibodies in 44 of the 56 (79%) patients tested with median titers comparable to the 115 donors (1:160 vs 1:160, p=0.40). These observations caused concerns about the potential benefit of convalescent plasma in the study population and after discussion with the data safety monitoring board, the study was discontinued. No difference in mortality (p=0.95), hospital stay (p=0.68) or day-15 disease severity (p=0.58) was observed between plasma treated patients and patients on standard of care.ConclusionMost COVID-19 patients already have high neutralizing antibody titers at hospital admission. Screening for antibodies and prioritizing convalescent plasma to risk groups with recent symptom onset will be key to identify patients that may benefit from convalescent plasma. Clinicaltrials.gov:NCT04342182

Sites of vulnerability in SARS-CoV-2 Antibodies that neutralize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) could be an important tool in treating coronavirus disease 2019 (COVID-19). Brouwer et al. isolated 403 monoclonal antibodies from three convalescent COVID-19 patients. They show that the patients had strong immune responses against the viral spike protein, a complex that binds to receptors on the host cell. A subset of antibodies was able to neutralize the virus. Competition and electron microscopy studies showed that these antibodies target diverse epitopes on the spike, with the two most potent targeting the domain that binds the host receptor. Science , this issue p. 643

The world is entering a new era of the COVID-19 pandemic in which there is an increasing call for reliable antibody testing. To support decision making on the deployment of serology for either population screening or diagnostics, we present a comprehensive comparison of serological COVID-19 assays. We show that the assay detecting total immunoglobulins against the receptor binding domain of SARS CoV-2, had optimal characteristics for antibody detection in different stages of disease.

Thus far, no human MERS-CoV infections have been reported from Israel. Evidence for the circulation of MERS-CoV in dromedaries has been reported from almost all the countries of the Middle East, except Israel. Therefore, we aimed to analyze MERS-CoV infection in Israeli camelids, sampled between 2012 and 2017. A total of 411 camels, 102 alpacas and 19 llamas' sera were tested for the presence of antibodies to MERS-CoV. Our findings indicate a lower MERS-CoV seropositivity among Israeli dromedaries than in the surrounding countries, and for the first time naturally infected llamas were identified. In addition, nasal swabs of 661 camels, alpacas and lamas, obtained from January 2015 to December 2017, were tested for the presence of MERS-CoV RNA. All nasal swabs were negative, indicating no evidence for MERS-CoV active circulation in these camelids during that time period. Keywords: MERS coronavirus, Antibodies, Israel, Dromedary camels, Llamas, Alpacas

We developed and validated 2 species-independent protein-based assays to detect Middle East respiratory syndrome coronavirus functional antibodies that can block virus receptor-binding or sialic acid-attachment. Antibody levels measured in both assays correlated strongly with virus-neutralizing antibody titers, proving their use for serologic confirmatory diagnosis of Middle East respiratory syndrome.