Candidate Antigens Research Articles

Designing of novel chimeric PvpA-pMGA protein of Mycoplasma gallisepticum, applicable for indirect ELISA

BackgroundMycoplasma gallisepticum is the primary agent of chronic respiratory disease in chickens creating important economic losses in poultry industry. pMGA and pvpA genes encode major surface proteins in M. gallisepticum containing pathogenic, antigenic, and immune evasion characteristics. The objective of the present study was to design, express, and purify the recombinant chimeric PvpA-pMGA protein from M.gallisepticum for using in serological diagnostic test. MethodsAntigenic regions of PvpA and pMGA proteins were predicted for designing chimeric pvpA-pMGA gene construct. The codon optimized sequence was cloned into the expression vector pET32a+ and transformed into the Escherichia coli strain BL21 (DE3). The pET32a-PvpA-pMGA recombinant plasmid was expressed and confirmed by SDS-PAGE and immunoblotting. PvpA-pMGA recombinant protein (20μg and 50μg), ts-11 vaccine strain, and S6 strain that formulated by montanide adjuvant and two control groups (PBS and adjuvant) were injected subcutaneously to six groups of chickens. ResultsHigh yield of protein was purified amount 138 mg/L by affinity batch formation method. Indirect ELISA showed the levels of antibodies in rPvpA-pMGA was significantly higher than ts-11 and S6 groups (p<0.05). The results indicated that antigen-specific response was successfully elicited by the rpMGA-PvpA in chickens. The result of the ELISA with sera collected from ts-11 and S6 groups showed that indirect PvpA-pMGA-ELISA is appropriate candidate for detection of specific antibodies against M. gallisepticum with 100% sensitivity and specificity. ConclusionsThe rPvpA-pMGA is a highly candidate immunogenic protein which induced high amount of humoral immune response. Novel rPvpA-pMGA protein could be useful for evaluation of antibody level in vaccinated poultry flocks. Highlights•This is first study to present chimeric protein rPvpA-pMGA as a candidate antigen in ELISA.•High-yield 138 mg/L purification of recombinant protein.•rPvpA-pMGA is a reliable immunogenic protein which induced high amount of humoral immune response.•rPvpA-pMGA showed high performance for evaluation of antibody level in vaccinated poultry flocks.

Longitudinal IgG antibody responses to Plasmodium vivax blood-stage antigens during and after acute vivax malaria in individuals living in the Brazilian Amazon.

To make progress towards malaria elimination, a highly effective vaccine targeting Plasmodium vivax is urgently needed. Evaluating the kinetics of natural antibody responses to vaccine candidate antigens after acute vivax malaria can inform the design of serological markers of exposure and vaccines. The responses of IgG antibodies to 9 P. vivax vaccine candidate antigens were evaluated in longitudinal serum samples from Brazilian individuals collected at the time of acute vivax malaria and 30, 60, and 180 days afterwards. Antigen-specific IgG correlations, seroprevalence, and half-lives were determined for each antigen using the longitudinal data. Antibody reactivities against Pv41 and PVX_081550 strongly correlated with each other at each of the four time points. The analysis identified robust responses in terms of magnitude and seroprevalence against Pv41 and PvGAMA at 30 and 60 days. Among the 8 P. vivax antigens demonstrating >50% seropositivity across all individuals, antibodies specific to PVX_081550 had the longest half-life (100 days; 95% CI, 83-130 days), followed by PvRBP2b (91 days; 95% CI, 76-110 days) and Pv12 (82 days; 95% CI, 64-110 days). This study provides an in-depth assessment of the kinetics of antibody responses to key vaccine candidate antigens in Brazilians with acute vivax malaria. Follow-up studies are needed to determine whether the longer-lived antibody responses induced by natural infection are effective in controlling blood-stage infection and mediating clinical protection.

Guillain-Barré Syndrome Following Zika Virus Infection Is Associated With a Diverse Spectrum of Peripheral Nerve Reactive Antibodies.

Recent outbreaks of Zika virus (ZIKV) in South and Central America have highlighted significant neurologic side effects. Concurrence with the inflammatory neuropathy Guillain-Barré syndrome (GBS) is observed in 1:4,000 ZIKV cases. Whether the neurologic symptoms of ZIKV infection are immune mediated is unclear. We used rodent and human live cellular models to screen for anti-peripheral nerve reactive IgG and IgM autoantibodies in the sera of patients with ZIKV with and without GBS. In this study, 52 patients with ZIKV-GBS were compared with 134 ZIKV-infected patients without GBS and 91 non-ZIKV controls. Positive sera were taken forward for target identification by immunoprecipitation and mass spectrometry, and candidate antigens were validated by ELISA and cell-based assays. Autoantibody reactions against glycolipid antigens were also screened on an array. Overall, IgG antibody reactivities to rat Schwann cells (SCs) (6.5%) and myelinated cocultures (9.6%) were significantly higher, albeit infrequent, in the ZIKV-GBS group compared with all controls. IgM antibody immunoreactivity to dorsal root ganglia neurones (32.3%) and SCs (19.4%) was more frequently observed in the ZIKV-GBS group compared with other controls, whereas IgM reactivity to cocultures was as common in ZIKV and non-ZIKV sera. Strong axonal-binding ZIKV-GBS serum IgG antibodies from 1 patient were confirmed to react with neurofascin 155 and 186. Serum from a ZIKV-infected patient without GBS displayed strong myelin-binding and putative antilipid antigen reaction characteristics. There was, however, no significant association of ZIKV-GBS with any known antiglycolipid antibodies. Autoantibody responses in ZIKV-GBS target heterogeneous peripheral nerve antigens suggesting heterogeneity of the humoral immune response despite a common prodromal infection.

IMMU-28. THE INTRA-TUMORAL SPATIAL HETEROGENEITY OF T CELL ANTIGENS IN GLIOBLASTOMA: AN INTEGRATED MULTI-OMICS APPROACH

Abstract This study investigates the intratumoral heterogeneity of T cell antigen presentation in 15 newly diagnosed glioblastoma patients. Our multi-omics approach includes mass spectrometry-based immunopeptidome analysis, next-generation sequencing (whole-exome and RNA sequencing), and imaging mass cytometry performed on biopsies deriving from the necrotic core (NEC), the gadolinium contrast-enhancing region (T1), and the peritumoral infiltrating zone (INF, 5-ALA positive). A total of 24493 unique HLA class I and 17394 unique HLA class II peptides were identified. Comparative profiling of peptides from our study and a benign tissue database (in-house; n = 429 donors combined with HLA ligand atlas (https://hla-ligand-atlas.org)) revealed that 20% of HLA class I ligands are glioblastoma exclusive. Of these ligands, 21%, 21%, and 15% were exclusively presented in the INF, T1, or NEC zone, respectively. Here, we focused on the INF-specific antigens as this zone is likely to remain after tumor resection and gives rise to glioblastoma recurrence. Interestingly, two INF-specific HLA ligands showed a frequency presentation of 45% in our patient cohort. One ligand originated from BAALC (Brain and acute leukemia cytoplasmic protein) and one from NCAN (Neurocan core protein), which are both known to be glioblastoma-associated proteins. Immunogenicity of pre-selected candidate antigens was assessed with autologous expanded T cells and revealed a set of novel promising targets for immunotherapy. Integrated RNA/DNA sequencing also enabled the identification of neoepitopes deriving from tumor- and region-specific mutations. Furthermore, imaging mass cytometry contributed to investigating the immune compartment of the tumor microenvironment in high dimensionality and spatial resolution. In conclusion, our approach characterized the intra-tumoral regional heterogeneity of both infiltrating immune cells and tumor antigens. This multi-omics spatial atlas of the immune landscape can be used for the informed design of immunotherapy strategies against glioblastoma.

Immunopeptidomic Analysis of BoLA-I and BoLA-DR Presented Peptides from Theileria parva Infected Cells.

The apicomplexan parasite Theileria parva is the causative agent of East Coast fever, usually a fatal disease for cattle, which is prevalent in large areas of eastern, central, and southern Africa. Protective immunity against T. parva is mediated by CD8+ T cells, with CD4+ T-cells thought to be important in facilitating the full maturation and development of the CD8+ T-cell response. T. parva has a large proteome, with >4000 protein-coding genes, making T-cell antigen identification using conventional screening approaches laborious and expensive. To date, only a limited number of T-cell antigens have been described. Novel approaches for identifying candidate antigens for T. parva are required to replace and/or complement those currently employed. In this study, we report on the use of immunopeptidomics to study the repertoire of T. parva peptides presented by both BoLA-I and BoLA-DR molecules on infected cells. The study reports on peptides identified from the analysis of 13 BoLA-I and 6 BoLA-DR datasets covering a range of different BoLA genotypes. This represents the most comprehensive immunopeptidomic dataset available for any eukaryotic pathogen to date. Examination of the immunopeptidome data suggested the presence of a large number of coprecipitated and non-MHC-binding peptides. As part of the work, a pipeline to curate the datasets to remove these peptides was developed and used to generate a final list of 74 BoLA-I and 15 BoLA-DR-presented peptides. Together, the data demonstrated the utility of immunopeptidomics as a method to identify novel T-cell antigens for T. parva and the importance of careful curation and the application of high-quality immunoinformatics to parse the data generated.

A Vaccine of SARS-CoV-2 S Protein RBD Induces Protective Immunity.

The pandemic of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has posed great threat to the world in many aspects. There is an urgent requirement for an effective preventive vaccine. The receptor binding domain (RBD), located on the spike (S) gene, is responsible for binding to the angiotensin-converting enzyme 2 (ACE2) receptor of host cells. The RBD protein is an effective and safe antigen candidate. The six-helix bundle (6HB) "molecular clamp" is a novel thermally-stable trimerization domain derived from a human immunodeficiency virus (HIV) gp41 protein segment. We selected the baculovirus system to fuse and express the RBD protein and 6HB for imitating the natural trimeric structure of RBD, named RBD-6HB. Recombinant RBD-6HB was successfully obtained from the cell culture supernatant and purified to high homogeneity. The purity of the final protein preparation was more than 97%. The results showed that the protein was identified as a homogeneous polymer. Further studies showed that the RBD-6HB protein combined with AL/CpG adjuvant could stimulate animals to produce sustained high-level antibodies and establish an effective protective barrier to protect mice from challenges. Our findings highlight the importance of trimerized SARS-CoV-2 S protein RBD in designing SARS-CoV-2 vaccines and provide a rationale for developing a protective vaccine through the induction of antibodies against the RBD domain.

Dual RNA-Seq Analysis Reveals Transcriptome Effects during the Salmon-Louse Interaction in Fish Immunized with Three Lice Vaccines.

Due to the reduced efficacy of delousing drugs used for sea lice control in salmon aquaculture, fish vaccines have emerged as one of the most sustainable strategies in animal health. Herein, the availability of C. rogercresseyi and Salmo salar genomes increases the capability of identifying new candidate antigens for lice vaccines using RNA sequencing and computational tools. This study aimed to evaluate the effects of two recombinant antigens characterized as peritrophin and cathepsin proteins on the transcriptome profiling of Atlantic salmon during a sea lice infestation. Four experimental groups were used: Peritrophin, cathepsin, and peritrophin/cathepsin (P/C), and PBS as the control. C. rogercresseyi female, S. salar head kidney, and skin tissue samples were sampled at 25 days post-infestation (dpi) for Illumina sequencing and RNA-seq analysis. Differential gene expression, gene ontology, and chromosomal expression analyses were performed. Furthermore, the dual RNA-seq analysis approach was performed to simultaneously explore host and pathogen transcriptomes, identifying functional associations for vaccine design. The morphometry of female sea lice exposed to immunized fish was also evaluated. The RNA-Seq analysis exhibited prototype-dependent transcriptome modulation, showing a conspicuous competition for metal ions during the infestation. Moreover, Dual RNA-seq analysis revealed vaccine-dependent gene patterns in both the host and the pathogen. Notably, significant morphometric differences between lice collected from immunized and control fish were observed, where cathepsin and P/C showed 57% efficacy. This study showed the potential of two proteins as lice vaccines for the salmon industry, suggesting novel molecular mechanisms between host-parasite interactions.

Optimization of purification conditions and study of antigenic properties of recombinant nucleocapsid protein of different SARS-CoV-2 strains

BACKGROUND: In the context of the constant manifestation of new SARS-CoV-2 strains and the need to determine the immunogenicity of new variants of antiviral vaccines, it is necessary to create diagnostic test systems based on conservative viral proteins. The SARS-CoV-2 nucleocapsid protein can be considered as a candidate antigen. However, the relevance of existing test systems based on it for determining the titer of antibodies produced in response to infection by recently emerging strains is unknown.&#x0D; AIM: The goal is to optimize the conditions for obtaining recombinant N proteins of various SARS-CoV-2 strains and to analyze the possibility of creating ELISA test systems based on them.&#x0D; MATERIALS AND METHODS: Bacterial strains producing N proteins were obtained by amplifying the corresponding genes and ligating them into the pETDuet-1 expression vector. Expression was induced at 20 or 37C for 1, 2, 4, or 20 h using inducer (IPTG) concentrations of 0.1 mM or 0.5 mM with or without the addition of 3% ethanol. Proteins were purified from biomass by metal affinity chromatography and used as antigens for the detection of antiviral antibodies by ELISA.&#x0D; RESULTS: It was found that the concentration of the inductor sufficient for the expression of recombinant proteins is 0.1 mM, the induction time is 1 h, and the required temperature is 37 C. The influence of the presence of ethanol as an expression-stimulating reagent was not revealed. When determining the titers of antiviral antibodies using the obtained proteins, cross-reactivity of serums of COVID-19 convalescents was established regarding to antigens of various SARS-CoV-2 strains.&#x0D; CONCLUSIONS: The possibility of effective induction of protein synthesis at a minimum concentration of the inducer and cultivation time indicates the economy of its production, and antigen recognition by antiviral antibodies indicates a native structure. Cross-reactivity of the blood sera of convalescents indicates the slow character of the evolution of the antigenic properties of the SARS-CoV-2 N protein. Thus, the purified proteins can be used as a basis for development of diagnostic test systems.

Recombinant Vaccinia Virus Expressing Plasmodium berghei Apical Membrane Antigen 1 or Microneme Protein Enhances Protection against P. berghei Infection in Mice.

Recombinant vaccinia viruses (rVV) are effective antigen delivery vectors and are researched widely as vaccine platforms against numerous diseases. Apical membrane antigen 1 (AMA1) is one of the candidate antigens for malaria vaccines but rising concerns regarding its genetic diversity and polymorphism have necessitated the need to search for an alternative antigen. Here, we compare the efficacies of the rVV vaccines expressing either AMA1 or microneme protein (MIC) of Plasmodium berghei in mice. Mice (BALB/c) were immunized with either rVV-AMA1 or rVV-MIC and subsequently challenge-infected with P. berghei. Compared to the control group, both antigens elicited elevated levels of parasite-specific antibody responses. Immunization with either one of the two vaccines induced high levels of T cells and germinal center B cell responses. Interestingly, rVV-MIC immunization elicited higher levels of cellular immune response compared to rVV-AMA1 immunization, and significantly reduced pro-inflammatory cytokine productions were observed from the former vaccine. While differences in parasitemia and bodyweight changes were negligible between rVV-AMA1 and rVV-MIC immunization groups, prolonged survival was observed for the latter of the two. Based on these results, our findings suggest that the rVV expressing the P. berghei MIC could be a vaccine-candidate antigen.

MAKING A STAT DIAGNOSIS IN A PATIENT PRESENTING WITH VILLOUS ATROPHY

<h3>Introduction</h3> Signal transducer and activator of transcription 1 (STAT1) gain-of-function (GOF) gene mutations are highly penetrant, resulting in enhanced STAT1-dependent responses to interferon-alpha/beta and interferon-gamma signaling, and impaired Th17 development. This manifests as recurrent infections, cytopenias, and autoimmune disease. Herein, we present a case of a child diagnosed with a STAT1 mutation. <h3>Case Description</h3> A 3-year-old female presented for secondary evaluation of persistent gastrointestinal and respiratory symptoms, recurrent sinopulmonary and skin infections, failure to thrive, and developmental delay. An esophagogastroduodenoscopy performed elsewhere showed increased duodenal intraepithelial lymphocytes with crypt hyperplasia and villous atrophy. Testing for Celiac disease and autoimmune enteritis were negative. Chest computed tomography showed bilateral bronchiectasis with mucous plugging. Bronchoalveolar lavage culture was positive for Streptococcus pneumoniae. Laboratory workup was notable for anemia and T-cell lymphopenia. Quantitative lymphocytes showed a low CD3+ count of 446 cells/mcL, low CD4+ count of 185 cells/mcL, and low CD8+ count of 194 cells/mcL. Lymphocyte proliferation to antigens showed essentially absent total CD45+ lymphocytes and CD3+ T cell responses to Candida and Tetanus toxoid antigens. Genetic testing later revealed a heterozygous, pathogenic c.1154C>T (p.Thr385Met) variant in the STAT1 gene. Additional testing for Th17 enumeration showed a decreased IL17 and Th17 cell percentage. She was treated with supportive care, antibiotics, and referred to Medical Genetics for additional education and counseling. <h3>Discussion</h3> STAT1-GOF gene mutations are rare and associated with variable phenotypes and disease severity. Genetic testing can help determine the cause of complex, multisystem diseases. Stem cell transplant, immunoglobulin replacement therapy, and JAK inhibitors are potential treatment considerations.

Evaluation of Protective Immune Responses Induced in BALB/c Mice and Goats by the Neospora caninum Surface SRS Proteins and Interleukin-18

Neosporosis is caused by Neospora caninum (N. caninum), which mainly infects cattle and goats and severely threatens the animal industry. In this study, the inhibitory effects of polyclonal antiserum anti-NcSRS17, NcSRS2 and NcSRS52 were explored. Cytokines in mice or goat serum were detected after immunization. After infection, the survival of mice was recorded. The pathological changes and parasite loads were observed and detected in tissues. The results showed that anti-NcSRS2, NcSRS17 and NcSRS52 antibodies all inhibit the invasion and proliferation of N. caninum. The IFN-γ level in the NcSRS17 group was higher than that in the NcSRS2 and NcSRS52 groups, and higher in the NcSRS2-mIL-18 group than in the NcSRS2 group. The survival rates of mice were 16% in the positive control group, 67% in the SRS52 group, 83% in the SRS2 and mIL-18 groups and 100% in the SRS17 and SRS2-mIL-18 groups. Goats immunized with NcSRS17-gIL-18 developed high levels of IL-4, IL-12 and IFN-γ compared with those immunized with NcSRS-17. Parasite loads in the brains of animals in the NcSRS17 and NcSRS17-gIL-18 groups were significantly reduced, and were significantly lower in the NcSRS17-gIL-18 group (p ≤ 0.01). This study indicates that SRS17 may be an antigen candidate for vaccine development against neosporosis, and IL-18 can enhance the immune protective efficiency of antigen candidates.

Immunoinformatics-Aided Design of a Peptide Based Multiepitope Vaccine Targeting Glycoproteins and Membrane Proteins against Monkeypox Virus.

Monkeypox is a self-limiting zoonotic viral disease and causes smallpox-like symptoms. The disease has a case fatality ratio of 3-6% and, recently, a multi-country outbreak of the disease has occurred. The currently available vaccines that have provided immunization against monkeypox are classified as live attenuated vaccinia virus-based vaccines, which pose challenges of safety and efficacy in chronic infections. In this study, we have used an immunoinformatics-aided design of a multi-epitope vaccine (MEV) candidate by targeting monkeypox virus (MPXV) glycoproteins and membrane proteins. From these proteins, seven epitopes (two T-helper cell epitopes, four T-cytotoxic cell epitopes and one linear B cell epitopes) were finally selected and predicted as antigenic, non-allergic, interferon-γ activating and non-toxic. These epitopes were linked to adjuvants to design a non-allergic and antigenic candidate MPXV-MEV. Further, molecular docking and molecular dynamics simulations predicted stable interactions between predicted MEV and human receptor TLR5. Finally, the immune-simulation analysis showed that the candidate MPXV-MEV could elicit a human immune response. The results obtained from these in silico experiments are promising but require further validation through additional in vivo experiments.

Potential Antigenic Candidates for the Development of Peptide-Based Vaccines to Induce Immunization against Helicobacter pylori Infection in BALB/c Mice.

Helicobacter pylori (H. pylori) has been identified as a group-1 definite carcinogen. As of yet, there is no available vaccine for this microorganism. Our study aimed to identify antigenic peptides in H. pylori using an in silico proteomic approach, and to evaluate their effectiveness as potential vaccine candidates. Four different peptide sequences were prioritized using the reverse vaccinology, namely, CagA1, CagA2, VacA, and SabA. Peptides emulsified with Freunde's adjuvant were used to immunize BALB/C mice. Subcutaneously immunized mice were challenged by oral administration of H. pylori. IgG, IgA, IL4, and IL17 were detected in mice sera. Histopathology of the dissected stomach of vaccinated and control mice were assessed using H&E stain. IgG was significantly higher in mice vaccinated with SabA. IL-4 was significantly increased in CagA1, CagA2, VacA, and SabA vaccinated mice compared to the adjuvant group. Additionally, histopathological examination of gastric tissue showed a protective effect in the vaccinated groups compared to adjuvant and PBS groups. Our findings indicate a promising effect of the tested epitopes, particularly the SabA antigen, to induce an immune response against H. pylori.

Lineage-mosaic and mutation-patched spike proteins for broad-spectrum COVID-19 vaccine

SARS-CoV-2 spread in humans results in continuous emergence of new variants, highlighting the need for vaccines with broad-spectrum antigenic coverage. Using inter-lineage chimera and mutation patch strategies, we engineered a recombinant monomeric spike variant (STFK1628x) that contains key regions and residues across multiple SAR-CoV-2 variants. STFK1628x demonstrated high immunogenicity and mutually complementary antigenicity to its prototypic form (STFK). In hamsters, a bivalent vaccine composed of STFK and STFK1628x elicited high titers of broad-spectrum neutralizing antibodies to 19 circulating SARS-CoV-2 variants, including Omicron sublineages BA.1, BA.1.1, BA.2, BA.2.12.1, BA.2.75 and BA.4/5. Furthermore, this vaccine conferred robust protection against intranasal challenges by either SARS-CoV-2 ancestral strain or immune-evasive Beta and Omicron BA.1. Strikingly, vaccination with the bivalent vaccine in hamsters effectively blocked within-cage virus transmission of ancestral SARS-CoV-2, Beta variant and Omicron BA.1 to unvaccinated sentinels. Thus, our study provided insight and antigen candidates for the development of next-generation COVID-19 vaccines.

The efficacy and safety of intralesional Candida vaccine versus topical diphencyproprobenone in immunotherapy of verruca vulgaris: A randomized comparative study

Various therapeutic options are available for verruca. While physical destruction may be associated with scarring, immunotherapy may be effective in treating warts through stimulating body immune response. The objective of the study was to compare the efficacy, safety, and outcome of Candida antigen vs diphencyprone (DPCP) in the treatment of warts. Fifty patients were randomly assigned to receive either intralesional Candida antigen every 3 weeks or weekly DPCP application. Both treatments were applied only to the mother wart. Lesions’ clearance and associated side effects were observed up to 4 weeks after treatment. Two blinded physicians evaluated photos of warts before and 4 weeks after the end of treatment. Both modalities granted wart clearance and/or improvement with no statistically significant difference; however, Candida antigen was significantly better in clearing adjacent untreated warts (p = 0.046). Fewer side effects were observed among the Candida antigen group. The response was duration associated in the Candida groups only. Intralesional Candida antigen injection and DPCP treatments for warts yielded improvement with superiority of Candida injection in eradicating distant lesions and fewer side effects. A shorter wart duration may be associated with a better therapeutic response with Candida antigen.

Level of maternal antibodies against respiratory syncytial virus (RSV) nucleoprotein at birth and risk of RSV very severe lower respiratory tract infection.

The nucleoprotein (N protein) of respiratory syncytial virus (RSV) is a candidate antigen for new RSV vaccine development. The aim of the present study was to investigate the association between maternal antibody titers against the RSV N protein at birth and the newborns' risk of developing very severe lower respiratory tract infection (VS-LRTI). In this single-center prospective cohort study, 578 infants born during the RSV epidemic season in France were included. Among these, 36 were hospitalized for RSV VS-LRTI. A generalized linear model was used to test the occurrence of a VS-LRTI in function of sex, mode of delivery, parity of the mother, type of pregnancy, date of birth in relation to the peak of the epidemic, and antibody titer against N protein. All cord blood samples had detectable antibodies against N protein. The mean titers were significantly lower in newborns with risk factors for RSV severe LRTI (preterm infants, birth before the peak epidemic, multiparous mother). There was no association between antibody titer against the N protein and a protection against VS-LRTI. Further studies are needed to support the hypothesis that transfer of maternal antibodies against the RSV N protein can provide a significant immune protection early in infancy and that N protein candidate vaccine may be a suitable target for maternal vaccine.

Molecular Assessment of Domain I of Apical Membrane Antigen I Gene in Plasmodium falciparum: Implications in Plasmodium Invasion, Taxonomy, Vaccine Development, and Drug Discovery.

Given its global morbidity and mortality rates, malaria continues to be a major public health concern. Despite significant progress in the fight against malaria, efforts to control and eradicate the disease globally are in jeopardy due to lack of a universal vaccine. The conserved short peptide sequences found in Domain I of Plasmodium falciparum apical membrane antigen 1 (PfAMA1), which are exposed on the parasite cell surface and in charge of Plasmodium falciparum invasion of host cells, make PfAMA1 a promising vaccine candidate antigen. The precise amino acids that make up these conserved short peptides are still unknown, and it is still difficult to pinpoint the molecular processes by which PfAMA1 interacts with the human host cell during invasion. The creation of a universal malaria vaccine based on the AMA1 antigen is challenging due to these knowledge limitations. This study used genome mining techniques to look for these particular short peptides in PfAMA1. Thirty individuals with Plasmodium falciparum malaria had blood samples taken using Whatman's filter papers. DNA from the parasite was taken out using the Chelex technique. Domain I of the Plasmodium falciparum AMA1 gene was amplified using nested polymerase chain reactions, and the amplified products were removed, purified, and sequenced. The DNA sequence generated was converted into the matching amino acid sequence using bioinformatic techniques. These amino acid sequences were utilized to search for antigenic epitopes, therapeutic targets, and conserved short peptides in Domain I of PfAMA1. The results of this investigation shed important light on the molecular mechanisms behind Plasmodium invasion of host cells, a potential PfAMA1 vaccine antigen sequence, and prospective malaria treatment options in the future. Our work offers fresh information on malaria medication and vaccine research that has not been previously discussed.

Recognition of mRNA Splice Variant and Secretory Granule Epitopes by CD4+ T Cells in Type 1 Diabetes.

A recent discovery effort resulted in identification of novel splice variant and secretory granule antigens within the HLA class I peptidome of human islets and documentation of their recognition by CD8+ T cells from peripheral blood and human islets. In the current study, we applied a systematic discovery process to identify novel CD4+ T cell epitopes derived from these candidate antigens. We predicted 145 potential epitopes spanning unique splice junctions and within conventional secretory granule antigens and measured their invitro binding to DRB1*04:01. We generated HLA class II tetramers for the 35 peptides with detectable binding and used these to assess immunogenicity and isolate T cell clones. Tetramers corresponding to peptides with verified immunogenicity were then used to label T cells specific for these putative epitopes in peripheral blood. T cells that recognize distinct epitopes derived from a cyclin I splice variant, neuroendocrine convertase 2, and urocortin-3 were detected at frequencies that were similar to those of an immunodominant proinsulin epitope. Cells specific for these novel epitopes predominantly exhibited a Th1-like surface phenotype. Among the three epitopes, responses to the cyclin I peptide exhibited a distinct memory profile. Responses to neuroendocrine convertase 2 were detected among pancreatic infiltrating T cells. These results further establish the contribution of unconventional antigens to the loss of tolerance in autoimmune diabetes.

Evaluation of new Toxocara canis chimeric antigens as an alternative to conventional TES-Ag for anti-Toxocara antibodies detection

Human toxocariasis is one of the neglected helminthiases and it is caused by the zoonotic roundworm species Toxocara canis and Toxocara cati. Diagnosis of human toxocariasis is based on the combination of clinical, parasitological, and epidemiological criteria, as well as serology tests that detect anti-Toxocara antibodies. Notwithstanding, due to the absence of pathognomonic symptoms and signs of the disease, serology is the key evidence to support a conclusive diagnosis. TES-ELISA is the most widely used serological test for diagnosis. However, cross-reaction of TES antigens with antibodies produced to other helminth antigens is a major drawback for its application in countries with high parasitic prevalence. T. canis recombinant antigens have been described as an alternative to native TES for diagnosis. Nevertheless, the selection of antigenic proteins is a complex process that requires validation. In this paper, we developed an eGFP carrier-based system to express and purify blocks of recombinant polypeptides of T. canis antigenic proteins. Intense cross-reaction polypeptides were detected by Immunoblot and avoided to finally produce a chimeric prototype protein. Additionally, a control chimeric protein that harbors the complete tested proteins was produced. Purified chimeric antigens were tested in ELISA and Immunoblot assays with 310 sera samples of negative and positive control individuals. Our results showed that chimeric rCHITC0 and rCHITC1 antigens (with sensitivities of 62% 58%, 38% and 16% in IB-rCHITC0, ELISA-rCHITC0, ELISA-rCHITC1 and IB-rCHITC1 respectively for OLMS) can perform better in terms of specificity (being 91%, 89%, 87% and 76% for ELISA-rCHITC1, IB-rCHITC1, ELISA-rCHITC0 and IB-rCHITC0 respectively for OLMS) than T. canis TES-ELISA (with 61% specificity), giving a higher signal with serum samples of infected individuals as well the possibility to discriminate false positive cases with other parasitic infections. Our data suggest that T. canis chimeric proteins, represent candidate antigens for phase II studies.

Bioinformatics-based SARS-CoV-2 epitopes design and the impact of spike protein mutants on epitope humoral immunities

BackgroundEpitope selection is the key to peptide vaccines development. Bioinformatics tools can efficiently improve the screening of antigenic epitopes and help to choose the right ones. ObjectiveTo predict, synthesize and testify peptide epitopes at spike protein, assess the effect of mutations on epitope humoral immunity, thus provide clues for the design and development of epitope peptide vaccines against SARS-CoV-2. MethodsBioinformatics servers and immunological tools were used to identify the helper T lymphocyte, cytotoxic T lymphocyte, and linear B lymphocyte epitopes on the S protein of SARS-CoV-2. Physicochemical properties of candidate epitopes were analyzed using IEDB, VaxiJen, and AllerTOP online software. Three candidate epitopes were synthesized and their antigenic responses were evaluated by binding antibody detection. ResultsA total of 20 antigenic, non-toxic and non-allergenic candidate epitopes were identified from 1502 epitopes, including 6 helper T-cell epitopes, 13 cytotoxic T-cell epitopes, and 1 linear B cell epitope. After immunization with antigen containing candidate epitopes S206-221, S403-425, and S1157-1170 in rabbits, the binding titers of serum antibody to the corresponding peptide, S protein, receptor-binding domain protein were (415044, 2582, 209.3), (852819, 45238, 457767) and (357897, 10528, 13.79), respectively. The binding titers to Omicron S protein were 642, 12,878 and 7750, respectively, showing that N211L, DEL212 and K417N mutations cause the reduction of the antibody binding activity. ConclusionsBioinformatic methods are effective in peptide epitopes design. Certain mutations of the Omicron would lead to the loss of antibody affinity to Omicron S protein.