Pre-existing cross-reactive antibody responses do not significantly impact inactivated COVID-19 vaccine-induced neutralization
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ABSTRACT: Recent exposure to seasonal coronaviruses (sCoVs) may stimulate cross-reactive antibody responses against SARS-CoV-2. Previous studies have shown divergent results regarding protective or damaging immunity induced by prior exposure to sCoVs. It is still unknown whether pre-existing humoral immunity may play a role in the vaccine-induced neutralization and antibody responses. In this study, we collected 36 paired sera in healthy volunteers before and after immunization with inactivated SARS-CoV-2 vaccines, and analyzed the distribution and intensity of pre-existing antibody responses at the epitope level before vaccine immunization, as well as the relationship between pre-existing sCoVs immunity and vaccine-induced neutralization.
Project description:Recent exposure to seasonal coronaviruses (sCoVs) may stimulate cross-reactive antibody responses against SARS-CoV-2. Previous studies have shown divergent results regarding protective or damaging immunity induced by prior exposure to sCoVs. It is still unknown whether pre-existing humoral immunity may play a role in the vaccine-induced neutralization and antibody responses. In this study, we collected 36 paired sera in healthy volunteers before and after immunization with inactivated SARS-CoV-2 vaccines, and analyzed the distribution and intensity of pre-existing antibody responses at the epitope level before vaccine immunization, as well as the relationship between pre-existing sCoVs immunity and vaccine-induced neutralization.
Project description:This experiment aims to profile polyclonal antibody binding profiles in serum from vaccinated animals relative to antibody function in a virus neutralization assay. Rabbits received three vaccinations with a DNA vaccine encoding the spike protein of the SARS-CoV-2 index strain. Serum samples were selected based on a three-tier (low, intermediate, and high) capacity to cross-neutralize SARS-CoV-2 strains with known neutralization resistance. Following normalization of total anti-spike IgG levels, serum of each animal (n=3) were evaluated for antibody binding to 10mer cyclic constrained peptides spanning the entire spike protein and regions with known SARS-CoV-2 variant of concern spike mutations.
Project description:The on-going COVID-19 pandemic requires a deeper understanding of the long-term antibody responses that persist following SARS-CoV-2 infection. To that end, we determined epitope-specific IgG antibody responses in COVID-19 convalescent sera collected at 5 months post-diagnosis and compared that to sera from naïve individuals. Each serum sample was reacted with a high-density peptide microarray representing the complete proteome of SARS-CoV-2 as 15 mer peptides with 11 amino acid overlap and homologs of spike glycoprotein, nucleoprotein, membrane protein, and envelope small membrane protein from related human coronaviruses. Binding signatures were compared between COVID-19 convalescent patients and naïve individuals using the web service tool EPIphany.
Project description:6 patients with severe COVID-19 were followed longitudinally during hospitalization and up to 1 year after infection, when they also received a vaccine. For each time point and patient, PBMCs were collected and split into 3 pools: 1/3 was sequenced straight; from 1/3 of the samples B cells were enriched (B cell enrichment kit from StemCell) and from 1/3 of the samples, antigen-specific cells were sorted using barcoded N, S and RBD probes. All samples were further stained using a cocktail of 181 barcoded Abs. For all samples we have sequences gene-expression, B cell receptor and Cell surface proteins.
Project description:The aim of the study was to test the hypothesis that peptide array reactivity will reveal patterns that can be associated with poor or effective neutralization response to seasonal influenza vaccine We diluted patient sera 1:80 and incubated dilutions on a streptavidin-coated glass surface that was printed with biotinylated peptides derived from seasonal strains of influenza hemagglutinin included in the 2008/2009 trivalent influenza vaccine In this study, serum from 76 study participants, subsetted into young (?30 years old) and elderly (?60 years old) categories were profiled for IgG/IgM antibody reactivity to influenza hemagglutinin peptudes. Using multivariate linear regression and correcting for multiple hypothesis testing by cross-validation, ee found that patterns of peptide reactivity associated with a poor or effective neutralization response to H1N1 and B strains of influenza.
Project description:We performed a study investigating donors previously infected with SARS-CoV-2 during the first wave of the COVID-19 pandemic to understand how vaccination may reshape T cell populations formed after infection. 10 donors were investigated using single-cell RNA-sequencing consisting of a) 3 mild non-hospitalised donors, b) 3 severe hospitalised donors, c) 1 mild recently vaccinated donor and d) 3 recently convalescent donors. For each of these groups we sampled at the following timepoints a) 6-9 months and 18 months after infection, b) 6-9 months and 18 months after infection, c) 13 months and 15 months after infection and d) 35 days after infection. All donors were unvaccinated at the first time point and vaccinated at the second time point. We stimulated PBMC with an overlapping peptide pool derived from the spike glycoprotein of the SARS-CoV-2 virus and sorted spike-specific CD4+ T cells (CD4+CD69+CD40L+) and spike-specific CD8+ T cells (CD8+CD69+4-1BB+) from every donor and time point using flow cytometry for 10X single-cell sequencing (5' protocol). Each time point from Dnr4868 was sequenced across two 10X reactions/libraries performed on the same day. Multiple samples were hashed and pooled together for sequencing using TotalSeq-C anti-human hashing antibodies from BioLegend. Individual library expression matrix files were generated using the CellRanger pipeline from 10X Genomics. The processed data file named immune.combined220929.rds is a Seurat Object containing all samples and generated using the Seurat package in R.
Project description:Antigen specific humoral immunity can be characterized by the analysis of serum antibodies. While serological assays for the measurement of antibody levels and of neutralization potential against SARS-CoV-2 are available, these are not quantitative in the biochemical sense. Yet, understanding the biology of COVID-19 would need an unambiguous, complete, quantitative, comparable measurement of specific serum antibodies. Here we describe a fluorescent, dual-titration immunoassay, which provides the physico-chemical parameters that are both necessary and sufficient to quantitatively characterize the humoral immune response. We used recombinant Receptor Binding Domain of SARS-CoV-2 as antigen on microspot arrays and varied the concentration of both the antigen and serum antibodies from vaccinated persons to obtain a measurement matrix of binding data. Binding curves were fitted using a novel algorithm to obtain thermodynamic variables of binding. We defined the standard state for a system of serum antibodies and antigen and showed how a normalized generalized logistic function is related to thermodynamic activity, standard concentration and activity coefficient. The utility of the method is demonstrated by defining the composition of tested sera with respect to immunoglobulin classes, affinity, concentration, and thermodynamic activity. The proposed fluorescent dual-titration microspot immunoassay can generate truly quantitative serological data that is suitable for immunological, medical and systems biological analysis.
Project description:To further investigate the underlying mechanisms of severe acute respiratory syndrome (SARS) pathogenesis and evaluate the therapeutic efficacy of potential drugs and vaccines it is necessary to use an animal model that is highly representative of the human condition in terms of respiratory anatomy, physiology and clinical sequelae. The ferret, Mustela putorius furo, supports SARS-CoV replication and displays many of the symptoms and pathological features seen in SARS-CoV-infected humans. We have recently established a SARS-CoV infection-challenge ferret platform for use in evaluating potential therapeutics to treat SARS. The main objective of the current study was to extend our previous results and identify early host immune responses upon infection and determine immune correlates of protection upon challenge with SARS-CoV in ferrets. Keywords: time course This study is a simple time course (58 day) examination of host responses in 35 SARS-CoV (TOR2) infected ferrets with the addition of a challenge inoculation of SARS CoV (TOR2) at day 29 post infection. Three mock-infected ferrets are included as negative controls. Due to the unavailability of ferret microarrays, Affymetrix Canine 2.0 oligonucleotide arrays were chosen following sequence analysis of our ferret cDNA library (~5000 clones) and demonstration of high levels of homology (>80%) between dog and ferret.
Project description:We sought to test whether vaccine-induced immune responses could protect rhesus macaques (RMs) against upfront heterologous challenges with an R5 simian-human immunodeficiency virus, SHIV-2873Nip. We immunized the RMs with recombinant Env proteins heterologous to the challenge virus. For induction of immune responses against Gag, Tat, and Nef, we explored a strategy of immunization with overlapping synthetic peptides (OSP). The immune responses against Gag and Tat were finally boosted with recombinant proteins. The vaccinees and a group of ten control animals were given five low-dose intrarectal (i.r.) challenges with SHIV-2873Nip. All controls and seven out of eight vaccinees became systemically infected; there was no significant difference in viremia levels of vaccinees vs. controls. Prevention of viremia was observed in one vaccinee which showed strong boosting of virus-specific cellular immunity during virus exposures. The protected animal showed no challenge virus-specific neutralizing antibodies in the TZM-bl or A3R5 cell-based assays and had low level ADCC activity after the virus exposures. Microarray data strongly supported a role for cellular immunity in the protected animal. Our study represents a case of protection against heterologous tier 2 SHIV-C by vaccine-induced, virus-specific cellular immune responses. For the protected RM, blood was collected before vaccination, on the day of first virus exposure and six weeks after last virus challenge. Lymph node and rectal pinch biopsies were performed before vaccination and six weeks after last virus challenge. Blood was collected in tempus tubes and processed immediately according to the manufacturer’s instructions and stored at -80C. The biopsy specimens were cut into small pieces and immediately placed into RNAlater solution (Qiagen, Valencia, CA) and also stored at -80C. Total RNA from blood, lymph node and rectal biopsies was extracted using RNAeasy extraction kits (Qiagen, Valencia, CA). cDNA labeling, hybridization, staining and scanning were performed according to the manufacturer’s instructions (Affymetrix, Santa Clara, CA) for rhesus gene expression arrays.