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1.
Nat Commun ; 13(1): 1614, 2022 03 28.
Article in English | MEDLINE | ID: covidwho-1764178

ABSTRACT

SARS-CoV-2 vaccines are crucial in controlling COVID-19, but knowledge of which factors determine waning immunity is limited. We examined antibody levels and T-cell gamma-interferon release after two doses of BNT162b2 vaccine or a combination of ChAdOx1-nCoV19 and BNT162b2 vaccines for up to 230 days after the first dose. Generalized mixed models with and without natural cubic splines were used to determine immunity over time. Antibody responses were influenced by natural infection, sex, and age. IgA only became significant in naturally infected. A one-year IgG projection suggested an initial two-phase response in those given the second dose delayed (ChAdOx1/BNT162b2) followed by a more rapid decrease of antibody levels. T-cell responses correlated significantly with IgG antibody responses. Our results indicate that IgG levels will drop at different rates depending on prior infection, age, sex, T-cell response, and the interval between vaccine injections. Only natural infection mounted a significant and lasting IgA response.


Subject(s)
COVID-19 , Viral Vaccines , Antibodies, Viral , COVID-19/prevention & control , COVID-19 Vaccines , Humans , SARS-CoV-2 , Vaccination , Vaccines, Inactivated
2.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-306522

ABSTRACT

Effective tools to monitor SARS-CoV-2 transmission and humoral immune responses are highly needed. Protective humoral immunity involves neutralizing antibodies and will be a hallmark for the evaluation of a vaccine response efficacy. Here we present a sensitive, fast and simple neutralization ELISA method to determine the levels of antibody-mediated virus neutralization. We can show that it is strongly correlated with the more elaborate plaque reduction neutralization test (PRNT) (ρ = 0.9231, p < 0.0001). Furthermore, we present pre-clinical vaccine models using recombinant receptor binding domain (RBD) and full-length spike antigen as immunogens showing a profound antibody neutralization capacity that exceeds the highest neutralization titers from convalescent individuals. Using a panel of novel high-affinity murine monoclonal antibodies (mAbs) we also show that majority of the RBD-raised mAbs have inhibitory properties while only a few of the spike-raised mAbs do. In conclusion, the ELISA-based viral neutralization test offers a time- and cost-effective alternative to the PRNT. The immunization results indicate that vaccine strategies focused only on the RBD region may have major advantages over those based on the full spike sequence.

3.
Nat Immunol ; 23(2): 275-286, 2022 02.
Article in English | MEDLINE | ID: covidwho-1661973

ABSTRACT

The humoral arm of innate immunity includes diverse molecules with antibody-like functions, some of which serve as disease severity biomarkers in coronavirus disease 2019 (COVID-19). The present study was designed to conduct a systematic investigation of the interaction of human humoral fluid-phase pattern recognition molecules (PRMs) with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Of 12 PRMs tested, the long pentraxin 3 (PTX3) and mannose-binding lectin (MBL) bound the viral nucleocapsid and spike proteins, respectively. MBL bound trimeric spike protein, including that of variants of concern (VoC), in a glycan-dependent manner and inhibited SARS-CoV-2 in three in vitro models. Moreover, after binding to spike protein, MBL activated the lectin pathway of complement activation. Based on retention of glycosylation sites and modeling, MBL was predicted to recognize the Omicron VoC. Genetic polymorphisms at the MBL2 locus were associated with disease severity. These results suggest that selected humoral fluid-phase PRMs can play an important role in resistance to, and pathogenesis of, COVID-19, a finding with translational implications.


Subject(s)
COVID-19/immunology , Immunity, Humoral , Receptors, Pattern Recognition/immunology , SARS-CoV-2/immunology , Animals , C-Reactive Protein/immunology , C-Reactive Protein/metabolism , COVID-19/metabolism , COVID-19/virology , Case-Control Studies , Chlorocebus aethiops , Complement Activation , Coronavirus Nucleocapsid Proteins/genetics , Coronavirus Nucleocapsid Proteins/immunology , Coronavirus Nucleocapsid Proteins/metabolism , Female , Glycosylation , HEK293 Cells , Host-Pathogen Interactions , Humans , Male , Mannose-Binding Lectin/genetics , Mannose-Binding Lectin/immunology , Mannose-Binding Lectin/metabolism , Phosphoproteins/genetics , Phosphoproteins/immunology , Phosphoproteins/metabolism , Polymorphism, Genetic , Protein Binding , Receptors, Pattern Recognition/genetics , Receptors, Pattern Recognition/metabolism , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , SARS-CoV-2/pathogenicity , Serum Amyloid P-Component/immunology , Serum Amyloid P-Component/metabolism , Signal Transduction , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vero Cells
5.
Elife ; 102021 11 25.
Article in English | MEDLINE | ID: covidwho-1534520

ABSTRACT

The alpha/B.1.1.7 SARS-CoV-2 lineage emerged in autumn 2020 in the United Kingdom and transmitted rapidly until winter 2021 when it was responsible for most new COVID-19 cases in many European countries. The incidence domination was likely due to a fitness advantage that could be driven by the receptor-binding domain (RBD) residue change (N501Y), which also emerged independently in other variants of concern such as the beta/B.1.351 and gamma/P.1 strains. Here, we present a functional characterization of the alpha/B.1.1.7 variant and show an eightfold affinity increase towards human angiotensin-converting enzyme-2 (ACE-2). In accordance with this, transgenic hACE2 mice showed a faster disease progression and severity after infection with a low dose of B.1.1.7, compared to an early 2020 SARS-CoV-2 isolate. When challenged with sera from convalescent individuals or anti-RBD monoclonal antibodies, the N501Y variant showed a minor, but significant elevated evasion potential of ACE-2/RBD antibody neutralization. The data suggest that the single asparagine to tyrosine substitution remarkable rise in affinity may be responsible for the higher transmission rate and severity of the B.1.1.7 variant.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2/immunology , Animals , Antibodies, Monoclonal/metabolism , Antibodies, Neutralizing/metabolism , Antibodies, Viral/metabolism , COVID-19/genetics , COVID-19/metabolism , Disease Progression , Female , Humans , Male , Mice , Mutation, Missense , Protein Binding , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Severity of Illness Index , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , United Kingdom
6.
Front Immunol ; 12: 767981, 2021.
Article in English | MEDLINE | ID: covidwho-1528824

ABSTRACT

The ongoing pandemic of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to constitute a serious public health threat worldwide. Protective antibody-mediated viral neutralization in response to SARS-CoV-2 infection has been firmly characterized. Where the effects of the antibody response are generally considered to be beneficial, an important biological question regarding potential negative outcomes of a SARS-CoV-2 antibody response has yet to be answered. We determined the distribution of IgG subclasses and complement activation levels in plasma from convalescent individuals using in-house developed ELISAs. The IgG response towards SARS-CoV-2 receptor-binding domain (RBD) after natural infection appeared to be mainly driven by IgG1 and IgG3 subclasses, which are the main ligands for C1q mediated classical complement pathway activation. The deposition of the complement components C4b, C3bc, and TCC as a consequence of SARS-CoV-2 specific antibodies were depending primarily on the SARS-CoV-2 RBD and significantly correlated with both IgG levels and disease severity, indicating that individuals with high levels of IgG and/or severe disease, might have a more prominent complement activation during viral infection. Finally, freshly isolated monocytes and a monocyte cell line (THP-1) were used to address the cellular mediated inflammatory response as a consequence of Fc-gamma receptor engagement by SARS-CoV-2 specific antibodies. Monocytic Fc gamma receptor charging resulted in a significant rise in the secretion of the pro-inflammatory cytokine TNF-α. Our results indicate that SARS-CoV-2 antibodies might drive significant inflammatory responses through the classical complement pathway and via cellular immune-complex activation that could have negative consequences during COVID-19 disease. We found that increased classical complement activation was highly associated to COVID-19 disease severity. The combination of antibody-mediated complement activation and subsequent cellular priming could constitute a significant risk of exacerbating COVID-19 severity.


Subject(s)
Antibodies, Viral/blood , COVID-19/immunology , Complement System Proteins/immunology , Immunoglobulin G/blood , SARS-CoV-2/immunology , COVID-19/blood , Complement Activation , Cytokines/immunology , Humans , Inflammation/immunology , Receptors, IgG/immunology , THP-1 Cells
7.
Front Immunol ; 12: 757197, 2021.
Article in English | MEDLINE | ID: covidwho-1485060

ABSTRACT

The recent identification and rise to dominance of the P.1 and B.1.351 SARS-CoV-2 variants have brought international concern because they may confer fitness advantages. The same three positions in the receptor-binding domain (RBD) are affected in both variants, but where the 417 substitution differs, the E484K/N501Y have co-evolved by convergent evolution. Here we characterize the functional and immune evasive consequences of the P.1 and B.1.351 RBD mutations. E484K and N501Y result in gain-of-function with two different outcomes: The N501Y confers a ten-fold affinity increase towards ACE-2, but a modest antibody evasion potential of plasma from convalescent or vaccinated individuals, whereas the E484K displays a significant antibody evasion capacity without a major impact on affinity. On the other hand, the two different 417 substitutions severely impair the RBD/ACE-2 affinity, but in the combined P.1 and B.1.351 RBD variants, this effect is partly counterbalanced by the effect of the E484K and N501Y. Our results suggest that the combination of these three mutations is a two-step forward and one step back in terms of viral fitness.


Subject(s)
Angiotensin-Converting Enzyme 2/immunology , Antibodies, Viral/immunology , COVID-19 Vaccines , Mutation, Missense , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Vaccination , Adult , Amino Acid Substitution , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/immunology , Female , Humans , Male , Protein Domains , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
8.
J Immunol ; 207(3): 878-887, 2021 08 01.
Article in English | MEDLINE | ID: covidwho-1323338

ABSTRACT

Tools to monitor SARS-CoV-2 transmission and immune responses are needed. We present a neutralization ELISA to determine the levels of Ab-mediated virus neutralization and a preclinical model of focused immunization strategy. The ELISA is strongly correlated with the elaborate plaque reduction neutralization test (ρ = 0.9231, p < 0.0001). The neutralization potency of convalescent sera strongly correlates to IgG titers against SARS-CoV-2 receptor-binding domain (RBD) and spike (ρ = 0.8291 and 0.8297, respectively; p < 0.0001) and to a lesser extent with the IgG titers against protein N (ρ = 0.6471, p < 0.0001). The preclinical vaccine NMRI mice models using RBD and full-length spike Ag as immunogens show a profound Ab neutralization capacity (IC50 = 1.9 × 104 to 2.6 × 104 and 3.9 × 103 to 5.2 × 103, respectively). Using a panel of novel high-affinity murine mAbs, we also show that a majority of the RBD-raised mAbs have inhibitory properties, whereas only a few of the spike-raised mAbs do. The ELISA-based viral neutralization test offers a time- and cost-effective alternative to the plaque reduction neutralization test. The immunization results indicate that vaccine strategies focused only on the RBD region may have advantages compared with the full spike.


Subject(s)
Antibodies, Neutralizing/blood , Coronavirus Nucleocapsid Proteins/immunology , Enzyme-Linked Immunosorbent Assay/methods , Neutralization Tests/methods , Receptors, Virus/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/immunology , Animals , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Antigens, Viral/immunology , COVID-19/immunology , COVID-19/therapy , COVID-19 Vaccines/immunology , Humans , Immunization , Immunization, Passive , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , Mice , Protein Domains/immunology
10.
J Biol Chem ; 296: 100536, 2021.
Article in English | MEDLINE | ID: covidwho-1126904

ABSTRACT

Transmission of Severe Acute Respiratory Syndrome Coronavirus 2 from humans to animals has been reported for many domesticated species, including farmed minks. The identification of novel spike gene mutations appearing in minks has raised major concerns about potential immune evasion and challenges for the global vaccine strategy. One genetic variant, known as "cluster five," arose among farmed minks in Denmark and resulted in a complete shutdown of the world's largest mink production. However, the functional properties of this new variant are not established. Here we present functional data on the cluster-five variant, which contains a mutation resulting in a Y453F residue change in the receptor-binding domain (RBD) of the spike protein. Using an ELISA-based angiotensin-converting enzyme-2/RBD inhibition assay, we show that the Y453F variant does not decrease established humoral immunity from previously infected individuals or affect the neutralizing antibody response in a vaccine mouse model based on the original Wuhan strain RBD or spike as antigens. However, biolayer interferometry analysis demonstrates that it binds the human angiotensin-converting enzyme-2 receptor with a 4-fold higher affinity than the original strain, suggesting an enhanced transmission capacity and a possible challenge for viral control. These results also indicate that the rise in the frequency of the cluster-five variant in mink farms might be a result of the fitness advantage conferred by the receptor adaptation rather than evading immune responses.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , COVID-19/epidemiology , COVID-19/transmission , Mink/virology , Pandemics , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Amino Acid Substitution , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/immunology , Animals , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/metabolism , Antibodies, Viral/chemistry , Antibodies, Viral/metabolism , COVID-19/immunology , Convalescence , Denmark/epidemiology , Gene Expression , HEK293 Cells , Host-Pathogen Interactions/genetics , Host-Pathogen Interactions/immunology , Humans , Immune Sera/chemistry , Immunity, Innate , Models, Molecular , Mutation , Protein Binding , Protein Structure, Secondary , Recombinant Proteins/genetics , Recombinant Proteins/immunology , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Virus Internalization
11.
J Immunol ; 206(1): 109-117, 2021 01 01.
Article in English | MEDLINE | ID: covidwho-1067832

ABSTRACT

Globally, the COVID-19 pandemic has had extreme consequences for the healthcare system and has led to calls for diagnostic tools to monitor and understand the transmission, pathogenesis, and epidemiology, as well as to evaluate future vaccination strategies. In this study, we have developed novel, to our knowledge, flexible ELISA-based assays for specific detection of human SARS-CoV-2 Abs against the receptor-binding domain, including an Ag sandwich ELISA relevant for large population screening and three isotype-specific assays for in-depth diagnostics. Their performance was evaluated in a cohort of 350 convalescent participants with previous COVID-19 infection, ranging from asymptomatic to critical cases. We mapped the Ab responses to different areas on protein N and S and showed that the IgM, A, and G Ab responses against receptor-binding domain are significantly correlated to the disease severity. These assays and the data generated from them are highly relevant for diagnostics and prognostics and contribute to the understanding of long-term COVID-19 immunity.


Subject(s)
Antibodies, Viral/immunology , COVID-19/diagnosis , COVID-19/immunology , Convalescence , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Antibodies, Neutralizing/immunology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Male , Middle Aged , Reverse Transcriptase Polymerase Chain Reaction , SARS-CoV-2/genetics , Severity of Illness Index , Young Adult
12.
J Immunol ; 206(1): 109-117, 2021 01 01.
Article in English | MEDLINE | ID: covidwho-934538

ABSTRACT

Globally, the COVID-19 pandemic has had extreme consequences for the healthcare system and has led to calls for diagnostic tools to monitor and understand the transmission, pathogenesis, and epidemiology, as well as to evaluate future vaccination strategies. In this study, we have developed novel, to our knowledge, flexible ELISA-based assays for specific detection of human SARS-CoV-2 Abs against the receptor-binding domain, including an Ag sandwich ELISA relevant for large population screening and three isotype-specific assays for in-depth diagnostics. Their performance was evaluated in a cohort of 350 convalescent participants with previous COVID-19 infection, ranging from asymptomatic to critical cases. We mapped the Ab responses to different areas on protein N and S and showed that the IgM, A, and G Ab responses against receptor-binding domain are significantly correlated to the disease severity. These assays and the data generated from them are highly relevant for diagnostics and prognostics and contribute to the understanding of long-term COVID-19 immunity.


Subject(s)
Antibodies, Viral/immunology , COVID-19/diagnosis , COVID-19/immunology , Convalescence , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Antibodies, Neutralizing/immunology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Male , Middle Aged , Reverse Transcriptase Polymerase Chain Reaction , SARS-CoV-2/genetics , Severity of Illness Index , Young Adult
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