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1.
Front Immunol ; 12: 781280, 2021.
Article in English | MEDLINE | ID: covidwho-1608387

ABSTRACT

The development of more effective, accessible, and easy to administer COVID-19 vaccines next to the currently marketed mRNA, viral vector, and whole inactivated virus vaccines is essential to curtailing the SARS-CoV-2 pandemic. A major concern is reduced vaccine-induced immune protection to emerging variants, and therefore booster vaccinations to broaden and strengthen the immune response might be required. Currently, all registered COVID-19 vaccines and the majority of COVID-19 vaccines in development are intramuscularly administered, targeting the induction of systemic immunity. Intranasal vaccines have the capacity to induce local mucosal immunity as well, thereby targeting the primary route of viral entry of SARS-CoV-2 with the potential of blocking transmission. Furthermore, intranasal vaccines offer greater practicality in terms of cost and ease of administration. Currently, only eight out of 112 vaccines in clinical development are administered intranasally. We developed an intranasal COVID-19 subunit vaccine, based on a recombinant, six-proline-stabilized, D614G spike protein (mC-Spike) of SARS-CoV-2 linked via the LPS-binding peptide sequence mCramp (mC) to outer membrane vesicles (OMVs) from Neisseria meningitidis. The spike protein was produced in CHO cells, and after linking to the OMVs, the OMV-mC-Spike vaccine was administered to mice and Syrian hamsters via intranasal or intramuscular prime-boost vaccinations. In all animals that received OMV-mC-Spike, serum-neutralizing antibodies were induced upon vaccination. Importantly, high levels of spike-binding immunoglobulin G (IgG) and A (IgA) antibodies in the nose and lungs were only detected in intranasally vaccinated animals, whereas intramuscular vaccination only induced an IgG response in the serum. Two weeks after their second vaccination, hamsters challenged with SARS-CoV-2 were protected from weight loss and viral replication in the lungs compared to the control groups vaccinated with OMV or spike alone. Histopathology showed no lesions in lungs 7 days after challenge in OMV-mC-Spike-vaccinated hamsters, whereas the control groups did show pathological lesions in the lung. The OMV-mC-Spike candidate vaccine data are very promising and support further development of this novel non-replicating, needle-free, subunit vaccine concept for clinical testing.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/immunology , Immunity, Mucosal/immunology , SARS-CoV-2/immunology , Administration, Intranasal , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/epidemiology , COVID-19/virology , COVID-19 Vaccines/administration & dosage , Cytoplasmic Vesicles/immunology , Female , Humans , Immunoglobulin A/immunology , Mesocricetus , Mice, Inbred BALB C , Neisseria meningitidis/immunology , Pandemics/prevention & control , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vaccination/methods , Vaccines, Subunit/administration & dosage , Vaccines, Subunit/immunology
2.
Front Immunol ; 12: 793191, 2021.
Article in English | MEDLINE | ID: covidwho-1608200

ABSTRACT

Purpose: To compare SARS-CoV-2 antigen-specific antibody production and plasma neutralizing capacity against B.1 wild-type-like strain, and Gamma/P.1 and Delta/B.1.617.2 variants-of-concern, in subjects with different Covid-19 disease and vaccination histories. Methods: Adult subjects were: 1) Unvaccinated/hospitalized for Covid-19; 2) Covid-19-recovered followed by one BNT162b2 vaccine dose; and 3) Covid-19-naïve/2-dose BNT162b2 vaccinated. Multiplex Luminex® immunoassays measured IgG, IgA, and IgM plasma levels against SARS-CoV-2 receptor-binding domain (RBD), spike-1 (S), and nucleocapsid proteins. Neutralizing activity was determined in Vero E6 cytopathic assays. Results: Maximum anti-RBD IgG levels were similar in Covid-19­recovered individuals 8‒10 days after single-dose vaccination and in Covid-19-naïve subjects 7 days after 2nd vaccine dosing; both groups had ≈2­fold higher anti-RBD IgG levels than Unvaccinated/Covid-19 subjects tracked through 2 weeks post-symptom onset. Anti-S IgG expression patterns were similar to RBD within each group, but with lower signal strengths. Viral antigen-specific IgA and IgM levels were more variable than IgG patterns. Anti-nucleocapsid immunoglobulins were not detected in Covid-19-naïve subjects. Neutralizing activity against the B.1 strain, and Gamma/P.1 and Delta/B.1.617.2 variants, was highest in Covid­19-recovered/single-dose vaccinated subjects; although neutralization against the Delta variant in this group was only 26% compared to B.1 neutralization, absolute anti-Delta titers suggested maintained protection. Neutralizing titers against the Gamma and Delta variants were 33‒77% and 26‒67%, respectively, versus neutralization against the B.1 strain (100%) in the three groups. Conclusion: These findings support SARS-CoV-2 mRNA vaccine usefulness regardless of Covid-19 history, and confirm remarkable protection provided by a single vaccine dose in people who have recovered from Covid-19.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , Immunoglobulin Isotypes/immunology , SARS-CoV-2/immunology , Adult , Aged , Aged, 80 and over , Animals , COVID-19/virology , Chlorocebus aethiops , Female , Humans , Immunoassay/methods , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin Isotypes/blood , Immunoglobulin M/blood , Immunoglobulin M/immunology , Male , Middle Aged , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vaccination/methods , Vero Cells
3.
Int Immunopharmacol ; 103: 108491, 2022 Feb.
Article in English | MEDLINE | ID: covidwho-1587489

ABSTRACT

To better understand the immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in individuals with COVID-19, it is important to investigate the kinetics of the antibody responses and their associations with the clinical course in different populations, since there seem to be considerable differences between Western and Asian populations in the clinical features and spread of COVID-19. In this study, we serially measured the serum titers of IgM, IgG and IgA antibodies generated against the nucleocapsid protein (NCP), S1 subunit of the spike protein (S1), and receptor-binding domain in the S1 subunit (RBD) of SARS-CoV-2 in Japanese individuals with COVID-19. Among the IgM, IgG, and IgA antibodies, IgA antibodies against all of the aforementioned viral proteins were the first to appear after the infection, and IgG and/or IgA seroconversion often preceded IgM seroconversion. In regard to the timeline of the antibody responses to the different viral proteins (NCP, S1 and RBD), IgA against NCP appeared than IgA against S1 or RBD, while IgM and IgG against S1 appeared earlier than IgM/IgG against NCP or RBD. The IgG responses to all three viral proteins and responses of all three antibody classes to S1 and RBD were sustained for longer durations than the IgA/IgM responses to all three viral proteins and responses of all three antibody classes to NCP, respectively. The seroconversion of IgA against NCP occurred later and less frequently in patients with mild COVID-19. These results suggest possible differences in the antibody responses to SARS-CoV-2 antigens between the Japanese and Western populations.


Subject(s)
COVID-19/epidemiology , COVID-19/immunology , Immunoglobulin A/blood , Immunoglobulin G/blood , Immunoglobulin M/blood , SARS-CoV-2 , Antibody Formation , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Japan/epidemiology , Japan/ethnology , Seroconversion , Viral Proteins/immunology
4.
Front Immunol ; 12: 777103, 2021.
Article in English | MEDLINE | ID: covidwho-1528826

ABSTRACT

Background: Data regarding symptoms in the lactating mother-infant dyad and their immune response to COVID-19 mRNA vaccination during lactation are needed to inform vaccination guidelines. Methods: From a prospective cohort of 50 lactating individuals who received mRNA-based vaccines for COVID-19 (mRNA-1273 and BNT162b2), blood and milk samples were collected prior to first vaccination dose, immediately prior to 2nd dose, and 4-10 weeks after 2nd dose. Symptoms in mother and infant were assessed by detailed questionnaires. Anti-SARS-CoV-2 antibody levels in blood and milk were measured by Pylon 3D automated immunoassay and ELISA. In addition, vaccine-related PEGylated proteins in milk were measured by ELISA. Blood samples were collected from a subset of infants whose mothers received the vaccine during lactation (4-15 weeks after mothers' 2nd dose). Results: No severe maternal or infant adverse events were reported in this cohort. Two mothers and two infants were diagnosed with COVID-19 during the study period before achieving full immune response. PEGylated proteins were not found at significant levels in milk after vaccination. After vaccination, levels of anti-SARS-CoV-2 IgG and IgM significantly increased in maternal plasma and there was significant transfer of anti-SARS-CoV-2-Receptor Binding Domain (anti-RBD) IgA and IgG antibodies to milk. Milk IgA levels after the 2nd dose were negatively associated with infant age. Anti-SARS-CoV-2 IgG antibodies were not detected in the plasma of infants whose mothers were vaccinated during lactation. Conclusions: COVID-19 mRNA vaccines generate robust immune responses in plasma and milk of lactating individuals without severe adverse events reported.


Subject(s)
Antibodies, Viral/immunology , COVID-19 Vaccines/administration & dosage , Lactation/immunology , Milk, Human/immunology , SARS-CoV-2/immunology , Adult , Antibodies, Viral/blood , COVID-19/prevention & control , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , Infant , Infant, Newborn , Male , Middle Aged
5.
JAMA Netw Open ; 4(11): e2132563, 2021 11 01.
Article in English | MEDLINE | ID: covidwho-1499193

ABSTRACT

Importance: Although several studies have provided information on short-term clinical outcomes in children with perinatal exposure to SARS-CoV-2, data on the immune response in the first months of life among newborns exposed to the virus in utero are lacking. Objective: To characterize systemic and mucosal antibody production during the first 2 months of life among infants who were born to mothers infected with SARS-CoV-2. Design, Setting, and Participants: This prospective cohort study enrolled 28 pregnant women who tested positive for SARS-CoV-2 infection and who gave birth at Policlinico Umberto I in Rome, Italy, from November 2020 to May 2021, and their newborns. Maternal and neonatal systemic immune responses were investigated by detecting spike-specific antibodies in serum, and the mucosal immune response was assessed by measuring specific antibodies in maternal breastmilk and infant saliva 48 hours after delivery and 2 months later. Exposures: Maternal infection with SARS-CoV-2 in late pregnancy. Main Outcomes and Measures: The systemic immune response was evaluated by the detection of SARS-CoV-2 IgG and IgA antibodies and receptor binding domain-specific IgM antibodies in maternal and neonatal serum. The mucosal immune response was assessed by measuring spike-specific antibodies in breastmilk and in infant saliva, and the presence of antigen-antibody spike IgA immune complexes was investigated in breastmilk samples. All antibodies were detected using an enzyme-linked immunosorbent assay. Results: In total, 28 mother-infant dyads (mean [SD] maternal age, 31.8 [6.4] years; mean [SD] gestational age, 38.1 [2.3] weeks; 18 [60%] male infants) were enrolled at delivery, and 21 dyads completed the study at 2 months' follow-up. Because maternal infection was recent in all cases, transplacental transfer of virus spike-specific IgG antibodies occurred in only 1 infant. One case of potential vertical transmission and 1 case of horizontal infection were observed. Virus spike protein-specific salivary IgA antibodies were significantly increased (P = .01) in infants fed breastmilk (0.99 arbitrary units [AU]; IQR, 0.39-1.68 AU) vs infants fed an exclusive formula diet (0.16 AU; IQR, 0.02-0.83 AU). Maternal milk contained IgA spike immune complexes at 48 hours (0.53 AU; IQR, 0.25-0.39 AU) and at 2 months (0.09 AU; IQR, 0.03-0.17 AU) and may have functioned as specific stimuli for the infant mucosal immune response. Conclusions and Relevance: In this cohort study, SARS-CoV-2 spike-specific IgA antibodies were detected in infant saliva, which may partly explain why newborns are resistant to SARS-CoV-2 infection. Mothers infected in the peripartum period appear to not only passively protect the newborn via breastmilk secretory IgA but also actively stimulate and train the neonatal immune system via breastmilk immune complexes.


Subject(s)
COVID-19/immunology , Immunoglobulin A/immunology , Milk, Human/immunology , Pregnancy Complications, Infectious/immunology , Adult , COVID-19/blood , COVID-19/transmission , COVID-19 Serological Testing , Female , Humans , Immunoglobulin A/isolation & purification , Immunoglobulin G/immunology , Immunoglobulin G/isolation & purification , Infant , Infant, Newborn , Infectious Disease Transmission, Vertical/prevention & control , Male , Pregnancy , Pregnancy Complications, Infectious/blood , Prospective Studies , SARS-CoV-2 , Saliva/immunology , Spike Glycoprotein, Coronavirus/immunology
6.
Microbiol Spectr ; 9(2): e0141621, 2021 10 31.
Article in English | MEDLINE | ID: covidwho-1495015

ABSTRACT

The rapid worldwide spread of SARS-CoV-2 has accelerated research and development for controlling the COVID-19 pandemic. A multi-coronavirus protein microarray was created containing full-length proteins, overlapping protein fragments of various lengths, and peptide libraries from SARS-CoV-2 and four other human coronaviruses. Sera from confirmed COVID-19 patients as well as unexposed individuals were applied to multicoronavirus arrays to identify specific antibody reactivity. High-level IgG, IgM, and IgA reactivity to structural proteins S, M, and N of SARS-CoV-2, as well as accessory proteins such as ORF3a and ORF7a, were observed that were specific to COVID-19 patients. Antibody reactivity against overlapping 100-, 50-, and 30-amino acid fragments of SARS-CoV-2 proteins was used to identify antigenic regions. Numerous proteins of SARS-CoV, Middle East respiratory syndrome coronavirus (MERS-CoV), and the endemic human coronaviruses HCoV-NL63 and HCoV-OC43 were also more reactive with IgG, IgM, and IgA in COVID-19 patient sera than in unexposed control sera, providing further evidence of immunologic cross-reactivity between these viruses. Whereas unexposed individuals had minimal reactivity against SARS-CoV-2 proteins that poorly correlated with reactivity against HCoV-NL63 and HCoV-OC43 S2 and N proteins, COVID-19 patient sera had higher correlation between SARS-CoV-2 and HCoV responses, suggesting that de novo antibodies against SARS-CoV-2 cross-react with HCoV epitopes. Array responses were compared with validated spike protein-specific IgG enzyme-linked immunosorbent assays (ELISAs), showing agreement between orthologous methods. SARS-CoV-2 microneutralization titers were low in the COVID-19 patient sera but correlated with array responses against S and N proteins. The multi-coronavirus protein microarray is a useful tool for mapping antibody reactivity in COVID-19 patients. IMPORTANCE With novel mutant SARS-CoV-2 variants of concern on the rise, knowledge of immune specificities against SARS-CoV-2 proteins is increasingly important for understanding the impact of structural changes in antibody-reactive protein epitopes on naturally acquired and vaccine-induced immunity, as well as broader topics of cross-reactivity and viral evolution. A multi-coronavirus protein microarray used to map the binding of COVID-19 patient antibodies to SARS-CoV-2 proteins and protein fragments as well as to the proteins of four other coronaviruses that infect humans has shown specific regions of SARS-CoV-2 proteins that are highly reactive with patient antibodies and revealed cross-reactivity of these antibodies with other human coronaviruses. These data and the multi-coronavirus protein microarray tool will help guide further studies of the antibody response to COVID-19 and to vaccination against this worldwide pandemic.


Subject(s)
Antibodies, Viral/immunology , Coronavirus NL63, Human/immunology , Coronavirus OC43, Human/immunology , Epitopes/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , SARS-CoV-2/immunology , Antibodies, Viral/blood , Binding Sites, Antibody/immunology , COVID-19/immunology , Coronavirus Nucleocapsid Proteins/immunology , Cross Reactions/immunology , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Phosphoproteins/immunology , Protein Array Analysis , Spike Glycoprotein, Coronavirus/immunology , Viral Proteins/immunology , Viroporin Proteins/immunology
7.
Sci Rep ; 11(1): 21308, 2021 10 29.
Article in English | MEDLINE | ID: covidwho-1493219

ABSTRACT

The aim of this study was to present and evaluate novel oral vaccines, based on self-amplifying RNA lipid nanparticles (saRNA LNPs), saRNA transfected Lactobacillus plantarum LNPs, and saRNA transfected Lactobacillus plantarum, to neutralize severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) variants alpha and delta. After invitro evaluation of the oral vaccines on HEK293T/17 cells, we found that saRNA LNPs, saRNA transfected Lactobacillus plantarum LNPs, and saRNA transfected Lactobacillus plantarum could express S-protein at both mRNA and protein levels. In the next step, BALB/c mice were orally vaccinated with saRNA LNPs, saRNA transfected Lactobacillus plantarum LNPs, and saRNA transfected Lactobacillus plantarum at weeks 1 and 3. Importantly, a high titer of IgG and IgA was observed by all of them, sharply in week 6 (P < 0.05). In all study groups, their ratio of IgG2a/IgG1 was upper 1, indicating Th1-biased responses. Wild-type viral neutralization assay showed that the secreted antibodies in vaccinated mice and recovered COVID-19 patients could neutralize SARS-COV-2 variants alpha and delta. After oral administration of oral vaccines, biodistribution assay was done. It was found that all of them had the same biodistribution pattern. The highest concentration of S-protein was seen in the small intestine, followed by the large intestine and liver.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19 Vaccines/administration & dosage , COVID-19/prevention & control , Lactobacillus plantarum/genetics , Lipids/chemistry , Nanoparticles/chemistry , SARS-CoV-2/immunology , Transfection/methods , Vaccination/methods , Vaccines, Synthetic/administration & dosage , Administration, Oral , Adult , Animals , COVID-19/blood , COVID-19/virology , COVID-19 Vaccines/pharmacokinetics , Female , HEK293 Cells , Humans , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Intestine, Small/metabolism , Lactobacillus plantarum/metabolism , Male , Mice , Mice, Inbred BALB C , Middle Aged , Models, Animal , Neutralization Tests , RNA, Messenger/genetics , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Tissue Distribution
8.
Front Immunol ; 12: 693462, 2021.
Article in English | MEDLINE | ID: covidwho-1485053

ABSTRACT

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first reported in December 2019 in Wuhan, China, and then rapidly spread causing an unprecedented pandemic. A robust serological assay is needed to evaluate vaccine candidates and better understand the epidemiology of coronavirus disease (COVID-19). Methods: We used the full-length spike (S) protein of SARS-CoV-2 for the development of qualitative and quantitative IgG and IgA anti-S enzyme linked immunosorbent assays (ELISA). A total of 320 sera used for assay development were comprised of pandemic sera from SARS-CoV-2 infected adults (n=51) and pre-pandemic sera (n=269) including sera from endemic human coronavirus infected adults. Reverse cumulative curves and diagnostic test statistics were evaluated to define the optimal serum dilution and OD cutoff value for IgG anti-S and IgA anti-S ELISAs. The IgG and IgA anti-S, and three functional antibodies (ACE-2 receptor blocking antibody, lentipseudovirus-S neutralizing antibody, and SARS-CoV-2 neutralizing antibody) were measured using additional SARS-CoV-2 PCR positive sera (n=76) and surveillance sera (n=25). Lastly, the IgG and IgA anti-S levels were compared in different demographic groups. Results: The optimal serum dilution for the qualitative IgG anti-S ELISA was at 1:1024 yielding a 99.6% specificity, 92.2% sensitivity, 92.9% positive predictive value (PPV), and 99.6% negative predictive value (NPV) at a SARS-CoV-2 seroprevalence of 5%. The optimal serum dilution for the qualitative IgA anti-S ELISA was at 1:128 yielding a 98.9% specificity, 76.5% sensitivity, 78.3% PPV, and 98.8% NPV at the same seroprevalence. Significant correlations were demonstrated between the IgG and IgA (r=0.833 for concentrations, r=0.840 for titers) as well as between IgG and three functional antibodies (r=0.811-0.924 for concentrations, r=0.795-0.917 for titers). The IgG and IgA anti-S levels were significantly higher in males than females (p<0.05), and in adults with moderate/severe symptoms than in adults with mild/moderate symptoms (p<0.001). Conclusion: We developed a highly specific and sensitive IgG anti-S ELISA assay to SARS-CoV-2 using full length S protein. The IgG anti-S antibody level was strongly associated with IgA and functional antibody levels in adults with SARS-CoV-2 infection. Gender and disease severity, rather than age, play an important role in antibody levels.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , Immunoglobulin A/immunology , Immunoglobulin G/immunology , SARS-CoV-2/immunology , Adult , COVID-19/diagnosis , COVID-19 Serological Testing , Female , HEK293 Cells , Humans
9.
JCI Insight ; 6(22)2021 11 22.
Article in English | MEDLINE | ID: covidwho-1476916

ABSTRACT

Understanding the presence and durability of antibodies against SARS-CoV-2 in the airways is required to provide insights into the ability of individuals to neutralize the virus locally and prevent viral spread. Here, we longitudinally assessed both systemic and airway immune responses upon SARS-CoV-2 infection in a clinically well-characterized cohort of 147 infected individuals representing the full spectrum of COVID-19 severity, from asymptomatic infection to fatal disease. In addition, we evaluated how SARS-CoV-2 vaccination influenced the antibody responses in a subset of these individuals during convalescence as compared with naive individuals. Not only systemic but also airway antibody responses correlated with the degree of COVID-19 disease severity. However, although systemic IgG levels were durable for up to 8 months, airway IgG and IgA declined significantly within 3 months. After vaccination, there was an increase in both systemic and airway antibodies, in particular IgG, often exceeding the levels found during acute disease. In contrast, naive individuals showed low airway antibodies after vaccination. In the former COVID-19 patients, airway antibody levels were significantly elevated after the boost vaccination, highlighting the importance of prime and boost vaccinations for previously infected individuals to obtain optimal mucosal protection.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19 Vaccines/administration & dosage , COVID-19 , Immunization, Secondary , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Lung/immunology , SARS-CoV-2/immunology , Adult , Aged , COVID-19/immunology , COVID-19/prevention & control , COVID-19 Vaccines/immunology , Female , Follow-Up Studies , Humans , Immunity, Humoral/drug effects , Longitudinal Studies , Male , Middle Aged
10.
Cells ; 10(10)2021 09 26.
Article in English | MEDLINE | ID: covidwho-1438527

ABSTRACT

Specific memory B cells and antibodies are a reliable read-out of vaccine efficacy. We analysed these biomarkers after one and two doses of BNT162b2 vaccine. The second dose significantly increases the level of highly specific memory B cells and antibodies. Two months after the second dose, specific antibody levels decline, but highly specific memory B cells continue to increase, thus predicting a sustained protection from COVID-19. We show that although mucosal IgA is not induced by the vaccination, memory B cells migrate in response to inflammation and secrete IgA at mucosal sites. We show that the first vaccine dose may lead to an insufficient number of highly specific memory B cells and low concentration of serum antibodies, thus leaving vaccinees without the immune robustness needed to ensure viral elimination and herd immunity. We also clarify that the reduction of serum antibodies does not diminish the force and duration of the immune protection induced by vaccination. The vaccine does not induce sterilizing immunity. Infection after vaccination may be caused by the lack of local preventive immunity because of the absence of mucosal IgA.


Subject(s)
Antibodies, Viral/immunology , B-Lymphocytes/cytology , COVID-19 Vaccines/therapeutic use , COVID-19/immunology , COVID-19/prevention & control , Immunoglobulin A/immunology , Immunologic Memory , Adult , Antibodies, Neutralizing/blood , Antigens, Viral/immunology , B-Lymphocytes/immunology , Cryopreservation , Female , Health Personnel , Healthy Volunteers , Hospitals, Pediatric , Humans , Immunoglobulin G , Immunoglobulin M/immunology , Lactation , Male , Middle Aged , Mucous Membrane/immunology , Patient Safety , SARS-CoV-2 , Vaccination
11.
Int J Biol Macromol ; 190: 409-416, 2021 Nov 01.
Article in English | MEDLINE | ID: covidwho-1412931

ABSTRACT

Coronavirus Disease 2019 (COVID-19) caused by a novel betacoronavirus SARS-CoV-2 has been an ongoing global pandemic. Several vaccines have been developed to control the COVID-19, but the potential effectiveness of the mucosal vaccine remains to be documented. In this study, we constructed a recombinant L. plantarum LP18:RBD expressing the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein via the surface anchoring route. The amount of the RBD protein was maximally expressed under the culture condition with 200 ng/mL of inducer at 33 °C for 6 h. Further, we evaluated the immune response in mice via the intranasal administration of LP18:RBD. The results showed that the LP18:RBD significantly elicited RBD-specific mucosal IgA antibodies in respiratory tract and intestinal tract. The percentages of CD3 + CD4+ T cells in spleens of mice administrated with the LP18:RBD were also significantly increased. This indicated that LP18:RBD could induce a humoral immune response at the mucosa, and it could be used as a mucosal vaccine candidate against the SARS-CoV-2 infection. We provided the first experimental evidence that the recombinant L. plantarum LP18:RBD could initiate immune response in vivo, which implies that the mucosal immunization using recombinant LAB system could be a promising vaccination strategy to prevent the COVID-19 pandemic.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , Immunity, Mucosal , Immunoglobulin A/immunology , Lactobacillus plantarum , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Administration, Intranasal , Animals , COVID-19/genetics , COVID-19/prevention & control , Gene Expression , Lactobacillus plantarum/genetics , Lactobacillus plantarum/immunology , Mice , Mice, Inbred BALB C , Protein Domains , Recombinant Proteins/genetics , Recombinant Proteins/immunology , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics
13.
Front Immunol ; 12: 690322, 2021.
Article in English | MEDLINE | ID: covidwho-1403471

ABSTRACT

A convalescent, non-severe, patient with COVID-19 was enrolled as a hyper-immune plasma voluntary donor by the Immuno-Hematology and Transfusion Unit of the Regina Elena National Cancer Institute in Rome, under the TSUNAMI national study criteria. During a nearly 6-month period (May-October 2020), the patient was closely monitored and underwent four hyperimmune plasma collections. Serum SARS-CoV-2 (anti-S + anti-N) IgG and IgM, anti-S1 IgA, and neutralizing titers (NTs) were measured. Anti-SARS-CoV-2 antibody levels steadily decreased. No correlation was found between anti-S/anti-N IgG and IgM levels and viral NT, measured by either a microneutralization test or the surrogate RBD/ACE2-binding inhibition test. Conversely, NTs directly correlated with anti-S1 IgA levels. Hyperimmune donor plasma, administered to five SARS-CoV-2 patients with persistent, severe COVID-19 symptoms, induced short-term clinical and pathological improvement. Reported data suggest that high NTs can persist longer than expected, thus widening hyperimmune plasma source, availability, and potential use. In vitro RBD/ACE2-binding inhibition test is confirmed as a convenient surrogate index for neutralizing activity and patients' follow-up, suitable for clinical settings where biosafety level 3 facilities are not available. IgA levels may correlate with serum neutralizing activity and represent a further independent index for patient evaluation.


Subject(s)
Antibodies, Neutralizing/administration & dosage , Antibodies, Viral/administration & dosage , COVID-19/therapy , SARS-CoV-2/immunology , Aged , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Blood Donors , COVID-19/immunology , COVID-19/virology , Humans , Immunization, Passive , Immunoglobulin A/administration & dosage , Immunoglobulin A/blood , Immunoglobulin A/immunology , Male , Middle Aged , Spike Glycoprotein, Coronavirus/immunology , Time Factors , Treatment Outcome
14.
Front Immunol ; 11: 618685, 2020.
Article in English | MEDLINE | ID: covidwho-1389172

ABSTRACT

Understanding humoral immune responses to SARS-CoV-2 infection will play a critical role in the development of vaccines and antibody-based interventions. We report systemic and mucosal antibody responses in convalescent individuals who experienced varying severity of disease. Whereas assessment of neutralization and antibody-mediated effector functions revealed polyfunctional antibody responses in serum, only robust neutralization and phagocytosis were apparent in nasal wash samples. Serum neutralization and effector functions correlated with systemic SARS-CoV-2-specific IgG response magnitude, while mucosal neutralization was associated with nasal SARS-CoV-2-specific IgA. Antibody depletion experiments support the mechanistic relevance of these correlations. Associations between nasal IgA responses, virus neutralization at the mucosa, and less severe disease suggest the importance of assessing mucosal immunity in larger natural infection cohorts. Further characterization of antibody responses at the portal of entry may define their ability to contribute to protection from infection or reduced risk of hospitalization, informing public health assessment strategies and vaccine development efforts.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , Immunity, Humoral/immunology , Immunity, Mucosal/immunology , Nasal Mucosa/immunology , Adolescent , Adult , Aged , Antibodies, Neutralizing/immunology , Antibody-Dependent Cell Cytotoxicity/immunology , Convalescence , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Male , Middle Aged , SARS-CoV-2/immunology , Young Adult
15.
EBioMedicine ; 71: 103519, 2021 Sep.
Article in English | MEDLINE | ID: covidwho-1363986

ABSTRACT

BACKGROUND: Given the importance of neutralising antibodies in protection against SARS-CoV-2 infection, it is critical to assess neutralisation persistence long-term following recovery. This study investigated neutralisation titres against SARS-CoV-2 up to 6 months post-symptom onset in individuals with mild COVID-19. METHODS: Plasma neutralisation titres in convalescent COVID-19 individuals were determined at baseline and 6 months post-symptom onset using a cell culture infectious SARS-CoV-2 assay. Total SARS-CoV-2 spike-specific IgG and IgA binding was measured using a lectin capture ELISA and compared between timepoints and correlated to neutralising titres. FINDINGS: All 48 convalescent COVID-19 individuals were found to have detectable SARS-CoV-2 50% inhibitory dilution neutralisation titres (ID50) at baseline and 6 months post-symptom onset with mean ID50 of 1/943 and 1/411, respectively. SARS-CoV-2 neutralisation titres peaked within 1-2 months post-symptom onset. However, 50% of individuals showed comparable ID50 at baseline and 6 months post-symptom onset. Both SARS-CoV-2 spike-specific IgG and IgA levels correlated well with neutralising titres. IgG binding was found to be sustained up to 6 months post-symptom onset, whereas IgA levels declined. INTERPRETATION: This study demonstrates durability of SARS-CoV-2 spike-specific IgG and neutralisation responses following recovery from mild COVID-19. Thus, all subjects included in this study might potentially have protective levels of neutralising antibodies 6 months post-symptom onset. This study also demonstrates a relationship between spike-specific IgA and neutralisation decline, with implications for long-term protection against SARS-CoV-2 infection. FUNDING: Novo Nordisk Foundation, Independent Research Fund Denmark and Danish Agency for Science and Higher Education.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/pathogenicity , Adult , COVID-19/epidemiology , COVID-19/virology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Male , Middle Aged
16.
Front Immunol ; 12: 696370, 2021.
Article in English | MEDLINE | ID: covidwho-1357528

ABSTRACT

The COVID-19 pandemic is caused by SARS-CoV-2, a novel zoonotic coronavirus. Emerging evidence indicates that preexisting humoral immunity against other seasonal human coronaviruses (HCoVs) plays a critical role in the specific antibody response to SARS-CoV-2. However, current work to assess the effects of preexisting and cross-reactive anti-HCoVs antibodies has been limited. To address this issue, we have adapted our previously reported multiplex assay to simultaneously and quantitatively measure anti-HCoV antibodies. The full mPlex-CoV panel covers the spike (S) and nucleocapsid (N) proteins of three highly pathogenic HCoVs (SARS-CoV-1, SARS-CoV-2, MERS) and four human seasonal strains (OC43, HKU1, NL63, 229E). Combining this assay with volumetric absorptive microsampling (VAMS), we measured the anti-HCoV IgG, IgA, and IgM antibodies in fingerstick blood samples. The results demonstrate that the mPlex-CoV assay has high specificity and sensitivity. It can detect strain-specific anti-HCoV antibodies down to 0.1 ng/ml with 4 log assay range and with low intra- and inter-assay coefficients of variation (%CV). We also estimate multiple strain HCoVs IgG, IgA and IgM concentration in VAMS samples in three categories of subjects: pre-COVID-19 (n=21), post-COVID-19 convalescents (n=19), and COVID-19 vaccine recipients (n=14). Using metric multidimensional scaling (MDS) analysis, HCoVs IgG concentrations in fingerstick blood samples were well separated between the pre-COVID-19, post-COVID-19 convalescents, and COVID-19 vaccine recipients. In addition, we demonstrate how multi-dimensional scaling analysis can be used to visualize IgG mediated antibody immunity against multiple human coronaviruses. We conclude that the combination of VAMS and the mPlex-Cov assay is well suited to performing remote study sample collection under pandemic conditions to monitor HCoVs antibody responses in population studies.


Subject(s)
Antibodies, Viral/blood , Coronavirus/immunology , Cross Reactions/immunology , Immunoassay/methods , Antibodies, Viral/immunology , Betacoronavirus/immunology , COVID-19/immunology , Coronavirus 229E, Human/immunology , Coronavirus NL63, Human/immunology , Coronavirus Nucleocapsid Proteins/immunology , Coronavirus OC43, Human/immunology , Humans , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , SARS Virus/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology
17.
Front Immunol ; 12: 690534, 2021.
Article in English | MEDLINE | ID: covidwho-1348488

ABSTRACT

High quality medical assistance and preventive strategies, including pursuing a healthy lifestyle, result in a progressively growing percentage of older people. The population and workforce is aging in all countries of the world. It is widely recognized that older individuals show an increased susceptibility to infections and a reduced response to vaccination suggesting that the aged immune system is less able to react and consequently protect the organism. The SARS-CoV-2 pandemic is dramatically showing us that the organism reacts to novel pathogens in an age-dependent manner. The decline of the immune system observed in aging remains unclear. We aimed to understand the role of B cells. We analyzed peripheral blood from children (4-18 years); young people (23-60 years) and elderly people (65-91 years) by flow cytometry. We also measured antibody secretion by ELISA following a T-independent stimulation. Here we show that the elderly have a significant reduction of CD27dull memory B cells, a population that bridges innate and adaptive immune functions. In older people, memory B cells are mostly high specialized antigen-selected CD27bright. Moreover, after in vitro stimulation with CpG, B cells from older individuals produced significantly fewer IgM and IgA antibodies compared to younger individuals. Aging is a complex process characterized by a functional decline in multiple physiological systems. The immune system of older people is well equipped to react to often encountered antigens but has a low ability to respond to new pathogens.


Subject(s)
Aging/immunology , B-Lymphocytes/immunology , COVID-19 , Immunologic Memory , Pandemics , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , COVID-19/epidemiology , COVID-19/immunology , Child , Child, Preschool , Cytokines/immunology , Female , Humans , Immunoglobulin A/immunology , Immunoglobulin M/immunology , Male , Middle Aged
18.
Proc Natl Acad Sci U S A ; 118(34)2021 08 24.
Article in English | MEDLINE | ID: covidwho-1345645

ABSTRACT

Alum, used as an adjuvant in injected vaccines, promotes T helper 2 (Th2) and serum antibody (Ab) responses. However, it fails to induce secretory immunoglobulin (Ig) A (SIgA) in mucosal tissues and is poor in inducing Th1 and cell-mediated immunity. Alum stimulates interleukin 1 (IL-1) and the recruitment of myeloid cells, including neutrophils. We investigated whether neutrophil elastase regulates the adjuvanticity of alum, and whether a strategy targeting neutrophil elastase could improve responses to injected vaccines. Mice coadministered a pharmacological inhibitor of elastase, or lacking elastase, developed high-affinity serum IgG and IgA antibodies after immunization with alum-adsorbed protein vaccines, including the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2). These mice also developed broader antigen-specific CD4+ T cell responses, including high Th1 and T follicular helper (Tfh) responses. Interestingly, in the absence of elastase activity, mucosal SIgA responses were induced after systemic immunization with alum as adjuvant. Importantly, lack or suppression of elastase activity enhanced the magnitude of anti-SARS-CoV-2 spike subunit 1 (S1) antibodies, and these antibodies reacted with the same epitopes of spike 1 protein as sera from COVID-19 patients. Therefore, suppression of neutrophil elastase could represent an attractive strategy for improving the efficacy of alum-based injected vaccines for the induction of broad immunity, including mucosal immunity.


Subject(s)
Adjuvants, Immunologic/pharmacology , Alum Compounds/pharmacology , COVID-19/immunology , COVID-19/therapy , Enzyme Inhibitors/pharmacology , Leukocyte Elastase/antagonists & inhibitors , SARS-CoV-2/immunology , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antibody Formation/drug effects , COVID-19/drug therapy , COVID-19/metabolism , HEK293 Cells , Humans , Immunity, Innate/drug effects , Immunity, Innate/immunology , Immunity, Mucosal/drug effects , Immunity, Mucosal/immunology , Immunoglobulin A/immunology , Leukocyte Elastase/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , SARS-CoV-2/drug effects , Spike Glycoprotein, Coronavirus/immunology , Swine , Th1 Cells/immunology
19.
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
20.
Curr Rheumatol Rep ; 23(8): 65, 2021 07 03.
Article in English | MEDLINE | ID: covidwho-1293441

ABSTRACT

PURPOSE OF REVIEW: COVID-19 patients have a procoagulant state with a high prevalence of thrombotic events. The hypothesis of an involvement of antiphospholipid antibodies (aPL) has been suggested by several reports. Here, we reviewed 48 studies investigating aPL in COVID-19 patients. RECENT FINDINGS: Prevalence of Lupus Anticoagulant (LA) ranged from 35% to 92% in ICU patients. Anti-cardiolipin (aCL) IgG and IgM were found in up to 52% and up to 40% of patients respectively. Anti-ß2-glycoprotein I (aß2-GPI) IgG and IgM were found in up to 39% and up to 34% of patients respectively. Between 1% and 12% of patients had a triple positive aPL profile. There was a high prevalence of aß2-GPI and aCL IgA isotype. Two cohort studies found few persistent LA but more persistent solid phase assay aPL over time. aPL determination and their potential role is a real challenge for the treatment of this disease.


Subject(s)
Antibodies, Antiphospholipid/immunology , COVID-19/immunology , Thrombosis/immunology , Antibodies, Anticardiolipin/immunology , C-Reactive Protein/immunology , COVID-19/blood , COVID-19/complications , Fibrin Fibrinogen Degradation Products/metabolism , Fibrinogen/metabolism , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Lupus Coagulation Inhibitor/immunology , SARS-CoV-2 , Severity of Illness Index , Thrombosis/blood , Thrombosis/etiology , beta 2-Glycoprotein I/immunology
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