A primary Chlamydia trachomatis genital infection of rhesus macaques identifies new immunodominant B-cell antigens

To identify immunodominant antigens that elicit a humoral immune response following a primary and a secondary genital infection, rhesus monkeys were inoculated cervically with Chlamydia trachomatis serovar D. Serum samples were collected and probed with a protein microarray expressing 864/894 (96.4%) of the open reading frames of the C. trachomatis serovar D genome. The antibody response to the primary infection was analyzed in 72 serum samples from 12 inoculated monkeys. The following criteria were utilized to identify immunodominant antigens: proteins found to be recognized by at least 75% (9/12) of the infected monkeys with at least 15% elevations in signal intensity from week 0 to week 8 post infection. All infected monkeys developed Chlamydia specific serum antibodies. Eight proteins satisfied the selection criteria for immunodominant antigens: CT242 (OmpH-like protein), CT541 (mip), CT681 (ompA), CT381 (artJ), CT443 (omcB), CT119 (incA), CT486 (fliY), and CT110 (groEL). Of these, three antigens, CT119, CT486 and CT381, were not previously identified as immunodominant antigens using non-human primate sera. Following the secondary infection, the antibody responses to the eight immunodominant antigens were analyzed and found to be quite different in intensity and duration to the primary infection. In conclusion, these eight immunodominant antigens can now be tested for their ability to identify individuals with a primary C. trachomatis genital infection and to design vaccine strategies to protect against a primary infection with this pathogen.

Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform

Modified Vaccinia Ankara (MVA) is a highly attenuated poxvirus vector that is widely used to develop vaccines for infectious diseases and cancer. We demonstrate the construction of a vaccine platform based on a unique three-plasmid system to efficiently generate recombinant MVA vectors from chemically synthesized DNA. In response to the ongoing global pandemic caused by SARS coronavirus-2 (SARS-CoV-2), we use this vaccine platform to rapidly produce fully synthetic MVA (sMVA) vectors co-expressing SARS-CoV-2 spike and nucleocapsid antigens, two immunodominant antigens implicated in protective immunity. We show that mice immunized with these sMVA vectors develop robust SARS-CoV-2 antigen-specific humoral and cellular immune responses, including potent neutralizing antibodies. These results demonstrate the potential of a vaccine platform based on synthetic DNA to efficiently generate recombinant MVA vectors and to rapidly develop a multi-antigenic poxvirus-based SARS-CoV-2 vaccine candidate.

Development of a Multi-Antigenic SARS-CoV-2 Vaccine Using a Synthetic Poxvirus Platform

Modified Vaccinia Ankara (MVA) is a highly attenuated poxvirus vector that is widely used to develop vaccines for infectious diseases and cancer. We developed a novel vaccine platform based on a unique three-plasmid system to efficiently generate recombinant MVA vectors from chemically synthesized DNA. In response to the ongoing global pandemic caused by SARS coronavirus-2 (SARS-CoV-2), we used this novel vaccine platform to rapidly produce fully synthetic MVA (sMVA) vectors co-expressing SARS-CoV-2 spike and nucleocapsid antigens, two immunodominant antigens implicated in protective immunity. Mice immunized with these sMVA vectors developed robust SARS-CoV-2 antigen-specific humoral and cellular immune responses, including potent neutralizing antibodies. These results demonstrate the potential of a novel vaccine platform based on synthetic DNA to efficiently generate recombinant MVA vectors and to rapidly develop a multi-antigenic poxvirus-based SARS-CoV-2 vaccine candidate.

Development of a Synthetic Poxvirus-Based SARS-CoV-2 Vaccine

Modified Vaccinia Ankara (MVA) is a highly attenuated poxvirus vector that is widely used to develop vaccines for infectious diseases and cancer. We developed a novel vaccine platform based on a unique three-plasmid system to efficiently generate recombinant MVA vectors from chemically synthesized DNA. In response to the ongoing global pandemic caused by SARS coronavirus-2 (SARS-CoV-2), we used this novel vaccine platform to rapidly produce fully synthetic MVA (sMVA) vectors co-expressing SARS-CoV-2 spike and nucleocapsid antigens, two immunodominant antigens implicated in protective immunity. Mice immunized with these sMVA vectors developed robust SARS-CoV-2 antigen-specific humoral and cellular immune responses, including potent neutralizing antibodies. These results demonstrate the potential of a novel vaccine platform based on synthetic DNA to efficiently generate recombinant MVA vectors and to rapidly develop a multi-antigenic poxvirus-based SARS-CoV-2 vaccine candidate.

Sequence Analysis of Pvama-1 among Plasmodium Vivax Isolates in Sistan-Baluchistan

BACKGROUND: Apical Membrane antigen 1 (AMA-1) is an important membrane protein that presents in all Plasmodium species and participates in critical phases in the attraction of cells. In human, it is one of the most immunodominant antigens with a protective immune response simulation role Apical Membrane antigen 1 (AMA-1) is an important membrane protein which presents in all Plasmodium species and is located on the surface of merozoite and sporozoites that participates in critical phases in attraction of human red blood cells by merozoites and hepatocytes by sporozoites, so in human, it is one of the most immunodominant antigens with a protective immune response simulation role. Since extra information is necessary to lighten of AMA-1 scope, we equaled genetic variation in P.vivax AMA-1 from 40 Iranian isolates with those reported from the other malarious countries.METHODS: Blood samples were collected from 40 patients’ positive of P.vivax, and genomic DNA was extracted from the blood. The nucleotide sequence for 446 amino acid (AA) residues (42–488 of PvAMA-1) of AMA-1 gene was amplified via PCR and then sequenced.RESULTS: A total of 24 different haplotypes were recognized between samples. No new haplotype was determined in this research that was reported previously in other regions of Iran and the world. We detected 37-point mutations at the nucleotide level in their sequences and showed 43 amino acid variations, at 37 positions in which 6 sites demonstrate trimorphic polymorphism, and the others were dimorphic.CONCLUSION: Sequence analysis of the major haplotype showed 95% similarity with P.vivax Sal-1 AMA-1 gene and high level of allelic diversity at the domain I of PvAMA-1 among P. vivaxisolates of Iran. Because PvAMA-1 is noticeable as vaccinecandidate antigen, these documents provide valuable informationfor the development of malaria vaccine.

In Silico Design of Multi-Epitope ESAT-6:Ag85b:Fcγ2a Fusion Protein as a Novel Candidate for Tuberculosis Vaccine

: Mycobacterium tuberculosis (MTB), which is the causative agent of tuberculosis (TB), is among the most important infectious bacteria with high morbidity and mortality rates worldwide. Bacilli Calmette-Guerin (BCG) vaccine has been discovered for about a century, and it is considered as a major vaccine for humans. However, some factors, such as its attenuated nature and its inefficacy against the latent form of the disease, have led to the use of alternative vaccines. Multi-epitope subunit vaccines are new-generation vaccines that are being developed in clinical trial phases. For the production of a subunit vaccine, the selection of immunodominant antigens and targeted delivery systems to antigen presenting cells (APCs) are considered as basic parameters. In the present study, we designed the novel multi-epitope ESAT-6:Ag85B:Fcγ2a, which was evaluated completely by various online tools as an optimum vaccine against TB. The early secreted antigenic target of 6 kDa (ESAT-6) and antigen 85B (Ag85B) are two immunodominant antigens, and Fcγ2a is a targeted delivery system. This vaccine candidate can be used for future preclinical studies.

Development of candidate vaccines against infection caused by shiga-toxin producing Escherichia coli. Part 1

Shiga-toxin producing Escherichia coli (STEC) strains cause serious and life-threatening diseases, hemorrhagic colitis (HC) and associated hemolytic uremic syndrome (HUS). Antibacterial etiotropic therapy against these diseases are not recommended. There are no vaccines against human HA and HUS. The review provides materials on the design of various types of candidate vaccines against STEC strains and assessment of their immunogenic and protective properties in experiments on laboratory and farm animals. The prospects for the use of inactivated corpuscular and live (vector) vaccines, lipopolysaccharide vaccines, DNA vaccines and nanovaccines, vaccines based on bacterial cell membranes (ghost), as well as vaccines created by reverse vaccinology methods in practice are considered. Key words: STEC, hemorrhagic colitis, immunodominant antigens, shiga toxins, EspA, EspB, Tir, intimine, IgG, sIgA

Repertoire of Cytomegalovirus-Specific T Cells Is Focused on the Immunodominant Epitopes in Fixed Hierarchy Dependent on HLA Genotype of the Donor

Background: Cytomegalovirus (HCMV) infection is one of the major complications and causes of mortality in patients receiving immunosuppressive therapy following hematopoietic stem cell transplantation (HSCT). Antiviral drugs have limited efficacy and high toxicity while using CMV-specific T cells from healthy individuals as a cellular vaccine had proven to be safe and efficient. Conventional protocols rely on expanding virus-specific cells ex vivo while more recently it was proposed to transfuse relatively small numbers of minimally manipulated cells and allow them to expand in vivo. To isolate virus-specific cell one could use either the stimulation with overlapping pools of peptides covering the whole immunogenic viral protein followed by isolation of INFγ-producing cells, or direct isolations of cells bound to MHC-multimer loaded with known immunodominant peptides. Both methods have their advantages. The former approach isolates all activated clones irrespective of their MHC-restriction and supplements CD8+ cells with CD4+ fraction, while the latter method is faster and simpler, besides it captures cells irrespective of their ability to produce IFNγ. The efficiency of the two methods was not directly compared and it is currently unknown to what extent T-cell populations isolated by these two approaches are different in terms of absolute quantity and clonal composition. Aims: to compare the quantity and T-cell receptor repertoire composition of T cells specific to pp65 CMV and to pp65-derived immunodominant epitopes NLV restricted by HLA-A*02, TPR and RPH restricted both by HLA-B*07. Methods: PBMC of healthy donors with different combination of HLA-A*02 and HLA-B*07 alleles were stimulated with peptides of immunodominant epitopes or overlapping peptide pool covering the entire pp65 CMV (Miltenyi Biotec, Cat.-130-093-435). Antigen-specific T cells were detected by the intracellular staining for IFNγ and flow cytometry. We obtained fractions of virus-specific T cells from PBMC either by MHC-tetramer staining or IFNγ secretion assay followed by fluorescent activated cell sorting. Then we prepared cDNA libraries of TCR α- and β-chains and sequenced them using NGS. Characterization of TCR repertories was performed by proprietary bioinformatic pipeline. Results: T-cell response to pp65 CMV is strongly focused on immunodominant pp65-derived epitopes. In donors having HLA-A*02 and/or HLA-B*07 absolute majority of pp65-reactive cells are specific to one of the three immunodominant epitopes - NLV, TPR and RPH. Immunodominant antigens are structured in a fixed hierarchy. TPR and RPH are superior to NLV. When HLA-B*07 is present response to the NLV epitope is drastically reduced, most of the pp65-specific cells recognize either TPR or RPH epitopes. TCR repertoire of the NLV-specific T cells is highly skewed and consists of few large dominant clones. TPR and RPH-specific T cells are more clonally diverse. Substantial share of the cells belonging to the large clones specific to the immunodominant antigens do not secrete IFNγ upon antigenic stimulation. Nevertheless, they may be therapeutically valuable, as they might secrete other cytokines. Conclusion: In summary, CMV-specific response in donors positive for HLA-A*02 and/or HLA-B*07 is focused to the combination of three immunodominant epitopes - NLV, TPR and RPH. Substantial share of the cells are not IFNγ-producers. For the patients having these alleles it might be beneficial to use MHC-multimers for isolation of the therapeutic lymphocytes. Figure 1: A - Healthy donors with different combination of HLA-A*02 and HLA-B*07 B - Intracellular staining for IFNγ of CD8+ T cells after stimulation with pp65-derived peptides C - NGS to identify clonally diverse of pp65-sprcific T cells on combination of HLA-A*02 and HLA-B*07 Figure 1 Disclosures No relevant conflicts of interest to declare.