Search Results (398)

Ancient wheat materials are increasingly used in cereal-based products because of their perceived nutritional advantages and distinctive protein composition. However, their protein quality and IgE-binding properties after processing remain insufficiently characterized. This study evaluated total protein content, essential amino acid composition, ELISA-detectable gliadin content, and IgE-binding patterns in flours and corresponding pasta products prepared from eight industry-sourced ancient and modern wheat materials, including einkorn, emmer, Kamut, spelt, round grain wheat, wheat 2Ab, common wheat, and durum wheat. Protein content was determined by the Kjeldahl method, amino acid composition by UHPLC, gliadin content by direct ELISA, and IgE-binding properties by slot blot analysis using sera from allergic individuals. The analysed wheat materials differed markedly in protein content, amino acid profile, gliadin detectability, and IgE-binding capacity. Spelt and Kamut showed the highest total protein contents, while selected ancient wheat materials, particularly emmer, Kamut, and einkorn, exhibited more favourable scores for some essential amino acids than common wheat and durum wheat. Nevertheless, none of the analysed materials fully met the FAO/WHO reference pattern for all evaluated essential amino acids, with lysine and histidine remaining the main limiting amino acids. Einkorn flour showed the highest ELISA-detectable gliadin content, consistent with its distinctive gluten protein composition. Processing into pasta reduced ELISA-detectable gliadin levels in all analysed materials, indicating processing-related changes in protein extractability and epitope accessibility. Despite reduced gliadin detectability after processing, IgE-reactive components remained detectable in both flour and pasta extracts. IgE-binding patterns were strongly dependent on wheat material, product form, and individual serum profile. Kamut pasta repeatedly showed one of the strongest IgE-binding responses across different sera, indicating that processing did not eliminate IgE-reactive components in this material. These findings demonstrate that selected ancient wheat materials may offer favourable protein-related nutritional characteristics, but these features should not be interpreted as evidence of reduced IgE-binding capacity. Integrated assessment of protein quality, gliadin detectability, and IgE-binding properties is therefore necessary for a more complete evaluation of ancient and modern wheat-based products. Full article

With the increasing consumption of plant-based foods, allergies to these foods are becoming more widespread and seriously affect human health. This review focuses on the properties of major plant-based food allergens, including those from soybeans, peanuts, wheat, sesame, and tree nuts. The composition, stability, structure, and IgE-binding epitopes of allergens from these major plant allergenic foods are summarized. For peanuts, the major allergens include Ara h 1, Ara h 2, Ara h 3, and Ara h 6, with Ara h 2 and Ara h 6 being notably heat- and protease-resistant. Gly m 4, Gly m 5, and Gly m 6 are the predominant allergens in soybeans. The primary sesame allergens are Ses i 1, Ses i 2, Ses i 3, Ses i 6, and Ses i 7, with Ses i 1 serving as a stable and highly specific diagnostic marker. Wheat allergens are categorized by exposure route, with ingestion of allergens such as gliadins and glutenins being particularly relevant for food-induced reactions. Major storage proteins and lipid transfer proteins are central to the allergenicity of nut varieties (such as almonds, hazelnuts, and walnuts). Understanding these molecular characteristics is crucial for developing hypoallergenic food products using targeted processing methods. Further studies are needed to comprehensively investigate these allergenic characteristics. Full article

Persistent infection with high-risk human papillomavirus (HPV), particularly HPV16, is a major driver of HPV-associated cancers; however, strategies for treating established HPV-induced tumors remain scarce. Here, we developed a DNA-based vaccine linking the SARS-CoV-2 spike (S) protein with an HPV16 E7 epitope (aa 49-57) to simultaneously induce antiviral humoral immunity and antitumor cellular responses. We generated 2 constructs, S-E7 and S-RE7, with the latter incorporating a furin cleavage site (R) to enhance antigen processing. In vitro, S-RE7 significantly enhanced E7-specific CD8⁺ T cell activation compared to S-E7, highlighting the importance of the furin sequence. In vivo, both S-linked vaccines elicited robust E7-specific CD8⁺ T cell responses and provided complete protection against TC-1 tumor challenge in a prophylactic murine model, with long-lasting immunity upon tumor rechallenge. In therapeutic settings, vaccination with S-E7 or S-RE7 significantly suppressed tumor growth, extended survival, and reduced circulating myeloid-derived suppressor cells (MDSCs), indicating alleviation of systemic immunosuppression. Notably, S-RE7 demonstrated faster antitumor effects overall in early tumor progression. In addition to cellular immunity, both constructs induced high levels of anti-spike antibodies, with S-RE7 eliciting approximately fourfold higher responses than S-E7. Furthermore, S-RE7 effectively boosted pre-existing anti-spike immunity in mice that were previously vaccinated. This “two-in-one” strategy represents a promising and versatile platform for the prevention and treatment of HPV-associated cancers while maintaining preparedness against potential SARS-CoV-2. Full article

Identifying antigens that elicit protective immunity is the key step for vaccine development. Here, we describe the use of the MS2 VLP platform to identify epitopes of SARS-CoV-2 structural proteins recognized by antibodies from COVID-19 patients, and to present those epitopes to the immune system as vaccines. We constructed an MS2 virus-like particle (VLP) library covering all four structural proteins of SARS-CoV-2 and affinity-selected vaccine candidates by biopanning on antibodies from infected humans. We focused on the structural proteins, reasoning that they are the most likely targets of a protective antibody response. The epitopes we found map almost entirely to the spike and nucleocapsid proteins. The VLPs displaying such epitopes were produced individually in E. coli and then tested for their potential as vaccines. While none of the affinity-selected spike-specific VLPs elicited neutralizing antibodies, VLPs displaying nucleocapsid epitopes induced protective immunity in a hamster model. This work illustrates the MS2 VLP platform’s capacity for the identification of new vaccine candidates and raises the possibility that VLPs displaying nucleocapsid epitopes could provide long-lasting protection against a range of virus variants. Full article

A single vaccine cannot prevent or treat all cancers; however, recurrent tumor-specific epitopes may facilitate the development of pre-manufactured, HLA-matched mRNA vaccines tailored for specific molecular subgroups. We define the shared tumor-specific epitope as a recurring peptide derived from a viral oncoprotein, a driver mutation, a frameshift, an altered protein C-terminus, or a fusion junction, and we employ a rigorous cancer-cell-only criterion: a target must be recurrent within a defined subgroup, absent from essential normal tissues at the peptide–HLA level, naturally presented on tumor cells, and sufficiently clonal to minimize immune escape. Under this criterion, we present fifteen sequence-anchored reference designs alongside one conceptual placeholder across thirteen candidates divided into four superclasses: viral oncoproteins (such as HPV16/18 E6 and E7 as attenuated antigenic reference designs; Merkel cell polyomavirus serving as a design-specific placeholder), recurrent driver neoepitopes (including KRAS G12/G13, IDH1 R132H, and H3 K27M), hematologic neoantigens (such as NPM1 Type A C-terminus; and a single CALR exon 9 construct encoding the shared novel C-terminus of types 1 and 2 mutations), and fusion junctions (notably EWS-FLI1 and BCR-ABL). Each open reading frame is anchored to a canonical accession with its documented event; representative ORFs are provided as reference designs, with the intended residue-level verification records. These sequence designs are intended as reference constructs and are not suitable as clinical-grade or manufacturing-ready products; they require independent residue-level validation and comprehensive safety assessments prior to laboratory or clinical application. The historical record of non-personalized vaccination—including HPV and hepatitis B prophylaxis, intravesical BCG, and unsuccessful tumor-associated antigen trials—frames both the potential and limitations of such approaches. The practical product is not a universal vaccine but rather a governed library aligned with specific genotype, viral etiology, HLA context, and clinical setting. Currently, none of these designs have established proof-of-benefit-tier evidence. Full article

(1) Background: As one of the world’s four major nuts, walnuts are rich in nutritional value; however, concerns regarding their allergenicity are becoming increasingly prominent. (2) Scope and Approach: This article provides a systematic review of the nutritional value and allergenicity of walnuts, the composition of major allergenic proteins, and their detection techniques. A particular focus is placed on elucidating the mechanisms by which different processing methods—including heat treatment, ultra-high pressure, ultrasound, low-temperature plasma, enzymatic treatment, and polyphenol modification—affect the structure and allergenicity of walnut allergenic proteins. (3) Key Findings and Conclusions: Current evidence suggests that processing techniques can alter the secondary and tertiary structures of walnut proteins, change the accessibility of linear or conformational epitopes, and reduce their Immunoglobulin E/Immunoglobulin G (IgE/IgG) binding capacity under certain in vitro conditions. Among these, high-temperature and high-pressure treatment, enzymatic hydrolysis, polyphenol modification, and combined processing strategies demonstrate promising potential for reducing walnut protein immunoreactivity. However, structural modifications, reduced antibody-binding capacity, or increased digestibility should not be directly interpreted as definitive evidence of reduced clinical sensitization. This paper summarizes the current status of the development and application of hypoallergenic foods, analyzes the technical challenges and future development directions, and aims to provide a theoretical basis and technical reference for the development of allergenicity-reduced walnut products. Full article

This review describes a novel method for increasing the effectiveness of inactivated enveloped whole-virus vaccines by targeting them for extensive uptake by antigen-presenting cells (APCs). Several inactivated whole-virus vaccines with dense glycan shields display suboptimal effectiveness because the multiple carbohydrate chains (glycans) on the virus mask immunogenic peptides and surround the virus with a negative electrostatic charge that decreases uptake by APCs. It is postulated that engineering such vaccinating viruses to present the carbohydrate antigen “α-gal epitope” on the glycan shields will immunocomplex them with the anti-Gal antibody; thus, it will target them for robust uptake by APCs. Anti-Gal is an abundant natural antibody in humans, constituting ~1% of human circulating immunoglobulins. The ligand of anti-Gal is the α-gal epitope, which is naturally synthesized in non-primate mammals and New World monkeys by the glycosylation enzyme α1,3galactosyltransferase. This enzyme is encoded by the GGTA1-gene. Viral vaccines presenting multiple α-gal epitopes on their glycan shield bind anti-Gal and activate the complement system to produce complement chemotactic cleavage peptides C5a and C3a that induce extensive recruitment of APCs to vaccine injection sites. The virion-bound anti-Gal further targets the viral vaccine for robust uptake by APCs, following binding of its Fc “tail” to Fcγ-receptors on APCs. The efficacy of this method was studied in anti-Gal-producing mice with α-gal presenting inactivated influenza virus vaccine and with gp120 of HIV presenting this epitope. These studies indicated that virus vaccines engineered to present α-gal epitopes increase anti-virus antibody production and virus-specific T-cell activation by 15- to 100-fold in comparison to the same vaccines lacking α-gal epitopes. It is suggested that α-gal presenting inactivated SARS-CoV-2 virus vaccines can induce a similar protective long-term immune memory against S- M-, E-, and N-viral proteins. Furthermore, immune-escaping variants of the mutated S-protein may be destroyed by antibodies to M and E proteins, and cells infected with such variants may be killed by cytotoxic T cells specific to peptides of the N-protein. Such an anti-M-, E-, and N-protein immune protection may prevent expansion of these variants and thus may avoid the need for immunization with COVID-19 vaccines every 6 months or following the appearance of new variants. A similar potent immunization may be achieved with an inactivated Ebolavirus vaccine engineered to present α-gal epitopes on the glycan shield. The resulting immune response to the various Ebolavirus proteins also may contribute to cross-reactive protection against other Ebolavirus species containing proteins with evolutionarily conserved structures. An effective method for the preparation of a whole-virus vaccine presenting α-gal epitopes is by arming it with the GGTA1-gene inserted into the viral genome. Such virions will present multiple α-gal epitopes on their glycan shield, which will amplify their immunogenicity instead of reducing it in the wild-type virus. Full article

Human papillomavirus type 53 (HPV53) is one of the most prevalent HPV genotypes in China, frequently detected in cervical intraepithelial neoplasia and cervical cancer, yet remains outside the coverage of all currently available prophylactic vaccines and is relatively understudied. This study performed a comprehensive analysis of HPV53 clinical infection profiles, genomic diversity, and T-cell epitopes to inform therapeutic vaccine development. Clinical analysis of 158 HPV53-positive patients showed that infections were most prevalent in women aged 40–59 years, with persistent infection identified in 13.3% participants and a subset of cases associated with cervical lesions. Genomic analysis of 134 HPV53 isolates identified four lineages (A-D, with lineage D further subdivided into four sublineages, and an overall nucleotide variability of 4.4%. E2 was the most variable protein while E7 was the most conserved. Immunoinformatic prediction identified 176 HLA class I-restricted T-cell epitopes across E6, E7, E1, and E2, from which 20 candidates were selected for experimental validation. Ten demonstrated strong HLA binding affinity in vitro, and murine immunization identified a E6 peptide VYNFAYTDL as an immunodominant epitope. Three validated epitopes exhibited sequence overlap with 12 to 13 of other 13 high-risk HPV genotypes, suggesting their potential as broadly cross-reactive targets. These findings clarify the genomic diversity and immunogenic epitope landscape of HPV53, providing a foundation for the rational design of therapeutic vaccines. Full article

Background: The high incidence and severe health threat of Toxoplasma gondii (T. gondii) infection, particularly in immunocompromised patients, underscore the urgent need for the development of a safe and effective vaccine. The aim of this study was to develop a novel multi-epitope vaccine (USM.TOXOII) incorporating the T. gondii GRA14, SAG1, and GRA1 antigens, and to assess its immunogenicity in BALB/c mice. Methods: Using bioinformatics approach, the USM.TOXOII was designed and evaluated. The encoding gene (471 bp) was then constructed and cloned into the pET-30a (+) plasmid before being transformed into E. coli expression system. The recombinant USM.TOXOII protein was subsequently expressed and purified. Finally, an animal study was performed to assess the vaccine’s immunogenicity. Results: The USM.TOXOII protein (17.27 kDa) was soluble and contained a His tag protein. Immunization of BALB/c mice with USM.TOXOII significantly elevated serum levels of total IgG, IgG1, and IgG2a (p < 0.05). Cytokine analysis revealed a significant increase in IFN-γ production, whereas IL-4 levels remained unchanged, suggesting a Th1-biased immune response. Conclusions: Collectively, these findings indicate that USM.TOXOII possesses immunogenic potential and is capable of inducing both humoral and cellular immune responses in BALB/c mice. Future challenge studies with live T. gondii tachyzoites are warranted to evaluate its protective efficacy in vivo. Full article

Dengue is an arboviral disease of global significance caused by Orthoflavivirus denguei (DENV), which has four antigenically distinct serotypes. The envelope (E) protein plays a critical role in viral entry and eliciting immune responses. This study aimed to compare the biochemical and immunological properties of the E protein across the four DENV serotypes using in silico approaches. E protein reference sequences were retrieved from RefSeq and analyzed with various bioinformatics tools. Sequence alignment revealed identities ranging from 63.08% to 77.69%. Biochemical analysis showed minimal variation in molecular weight and isoelectric point; however, the net charge of DENV-3 E protein was notably lower. Secondary structure predictions indicated a predominance of alpha-helices in DENVs-1/2, while DENVs-3/4 featured more beta-sheets. Post-translational modification analysis revealed mostly casein kinase II phosphorylation sites across all serotypes, with DENV-4 uniquely presenting also tyrosine kinase sites. Amino acids W231/D341 in DENV-1, Q86 in DENVs-2/4, and D87/D339 in DENV-3 showed maximum antigenicity scores in B cell recognition, while the human leukocyte antigen (HLA) alleles B*08:01/B*39:01 and DRB4*01:01, recognized by T cells, presented the highest number of predicted epitopes for the different DENV serotypes. Conservation analysis showed that the major antigenic regions highlighted in this study are highly conserved among contemporary DENV isolates despite the genetic variability observed within each serotype. These findings suggest that subtle structural differences in the E protein may contribute to distinct immunogenic profiles, highlighting candidate regions for future investigation. Full article

Background/Objectives: Canine papillomavirus (CPV) is an important viral pathogen associated with papillomatosis in dogs, with canine papillomavirus type 1 (CPV1) and type 2 (CPV2) among the most prevalent and clinically relevant genotypes. The L1 capsid protein is a major immunogenic antigen of papillomaviruses; however, conserved linear B-cell epitopes shared between CPV genotypes remain poorly defined. This study aimed to identify conserved cross-reactive B-cell epitopes within CPV1 and CPV2 L1 proteins and to evaluate their preliminary immunoreactivity. Methods: Conserved linear B-cell epitopes were predicted through integrated bioinformatic and structural analyses based on sequence conservation and surface accessibility. Three candidate epitopes were selected. Recombinant CPV1 and CPV2 L1 proteins were expressed in Escherichia coli (E. coli), purified, used as recombinant L1 antigens, together with BSA-conjugated synthetic epitope peptides for mouse immunization. Antigen-specific IgG responses were assessed by ELISA, antigen-associated IFN-γ responses were evaluated by ELISpot, and cross-reactive antibody recognition was assessed by Western blot. Results: Recombinant L1 proteins induced strong antigen-specific IgG responses in mice. The selected peptides induced detectable but weaker humoral responses compared with the recombinant L1 proteins. Among the three epitopes, TPSGSLV and TVVDNTR elicited antibodies that recognized both CPV1 and CPV2 L1 proteins, while the epitope VIVPKVS showed minimal or no detectable immunoreactivity. ELISpot analysis showed only modest antigen-associated IFN-γ responses, particularly in peptide-immunized groups. Conclusions: This study identified conserved cross-reactive linear B-cell epitope candidates within CPV1 and CPV2 L1 proteins and provided preliminary immunological evidence supporting their potential relevance for CPV antigen design. However, peptide-induced responses were weaker than those induced by recombinant L1 proteins, and VLP formation, antibody neutralizing activity, and protective efficacy were not evaluated. Further studies in dogs, including optimized antigen-display platforms, neutralization assays, and protection studies, are required to determine the practical value of these epitopes for CPV vaccine development. Full article

Background/Objectives: Respiratory syncytial virus (RSV), human metapneumovirus (hMPV), and parainfluenza virus type 3 (PIV3) are leading causes of acute respiratory infections in children and the elderly, yet no licensed T-cell vaccines are available. This study aimed to develop multivalent T-cell vaccine candidates against these pathogens using a live attenuated influenza virus (LAIV) vector platform. Methods: Conserved F, N, and M proteins of RSV, hMPV, and PIV3 were identified through multiple sequence alignments. Fragments enriched with experimentally confirmed and predicted T-cell epitopes were selected using the IEDB and NetMHCpan servers. These fragments were assembled into polyepitope immunogenic cassettes, and their selected order was determined by thermodynamic analysis of mRNA secondary structures using the RNAfold Web Server. The selected cassettes were cloned into the neuraminidase (NA) gene of a cold-adapted LAIV vector. Recombinant viruses were rescued by reverse genetics and assessed for replicative fitness in embryonated chicken eggs and MDCK cells, NA enzymatic activity and genetic stability upon serial passaging. Results: Four cassettes were designed for RSV, three for hMPV, and one for PIV3, all containing fragments with multiple T-cell epitopes. Three recombinant viruses of LAIV/RSV type and three of LAIV/hMPV type were successfully rescued, while attempts to recover the remaining recombinant viruses, i.e., LAIV/RSV and LAIV/PIV3, were not successful. All rescued recombinant viruses replicated to titers comparable to the parental LAIV strain and retained the full-length insert for at least eight passages in eggs. Importantly, NA enzymatic activity of the LAIV vector was not compromised by the insertion of the polyepitope T-cell cassettes. Conclusions: We developed a panel of recombinant T cell-based vaccine candidates against RSV and hMPV using the LAIV vector platform. These recombinant viruses encode conserved T-cell epitopes of the target viruses while retaining the biological properties of LAIV strains. Taken together, these characteristics warrant further evaluation of these recombinant viruses in appropriate relevant in vitro models to directly assess their immunogenicity in terms of stimulating a T-cell response against target pathogens. Full article

Background: The translocation of diet-derived antigens from the maternal intestine to breast milk represents a primary gateway for neonatal immune priming, yet the structural basis for why certain proteins survive this transit while others do not remains poorly understood. This review introduces the “Survivor Peptide” hypothesis, proposing that specific food allergens possess intrinsic “stability architectures” that enable them to resist maternal digestion and navigate the gut–mammary axis to reach the infant in an immunologically active form. Methods: We analyzed the current literature regarding the detection and structural characteristics of food allergens in human milk. Integrating evidence from 26 major sources, we performed an in silico structural analysis of five representative “survivor” proteins: Gal d 1 (egg white), Bos d 5 (cow’s milk), Gal d 6 (egg yolk), Tri a 19 (wheat), and tropomyosin (Der p 10-mite/shellfish). High-resolution 3D models were retrieved from the Protein Data Bank and AlphaFold2, and then visualized in UCSF ChimeraX to map stability anchors, including disulfide bonds and hydrophobic clusters, against solvent-accessible IgE-binding epitopes. Results: We identified and categorized allergens into distinct Molecular Resilience Architectures: the “Covalent Cage” (Gal d 1), defined by dense disulfide stapling, the “Glycoprotein Shield” (Gal d 6), utilizing yolk-matrix structural anchors, the “Topological Shield” (Bos d 5), characterized by a stable β-barrel, and “Coiled-Coil Rigidity” (Der p 10). These frameworks protect large, immunogenic fragments that maintain the spatial arrangement required for IgE cross-linking. Conclusions: Allergen persistence in the gut–mammary axis is dictated by a protein’s intrinsic structural architecture. Identifying these stability fingerprints provides a unified theory for allergen persistence and offers a path for refining component-resolved diagnostics and neonatal oral tolerance strategies. Full article

The Getah virus (GETV) is a mosquito-borne pathogen that infects diverse hosts, including pigs, horses, and humans, which can cause swine reproductive disorders such as abortion and stillbirth, posing a potential threat to animal and public health. Therefore, there is an urgent need for efficient and accurate serological diagnostic methods for surveillance and control of GETV. However, commercial diagnostic kits for swine GETV infection remain unavailable. In this study, we developed a novel enzyme-linked immunosorbent assay (ELISA) based on a GETV-specific epitope peptide (E2EP3) for serological detection. The N-terminally biotinylated E2EP3 peptide was synthesized, and the reaction conditions were systematically optimized, resulting in a cut-off value of 0.363. The assay exhibited no cross-reactivity with Japanese encephalitis virus (JEV), porcine circovirus type 2 (PCV2), porcine circovirus type 3 (PCV3), pseudorabies virus (PRV), or classical swine fever virus (CSFV). It demonstrated good reproducibility and high sensitivity, detecting GETV-positive serum diluted up to 1:640. The overall agreement rate reached 95%, consistent with a conventional recombinant GETV E2 protein-based ELISA. Benefiting from the biotin–streptavidin system, this assay achieved strong signal amplification and low background. Moreover, the procedure is simple, cost-effective, and stable, making it suitable for GETV large-scale serological surveillance and vaccine evaluation. Full article

Background/Objectives: Recombinant poliovirus (PV) virus-like particle (VLP) antigens mimic the conformation of the surface proteins in native PVs (i.e., serotype-specific D-antigen epitopes). Since they lack genomes and are non-infectious, PV-VLPs offer the promise of a safer, next-generation polio vaccine compared to traditional inactivated (IPV) or attenuated live (OPV) vaccines. Sandwich D-antigen ELISA formats are commonly used to measure the in vitro potency values (relative D-antigen content, DU/mL) of unadjuvanted trivalent IPV antigens. If IPV is formulated with aluminum-salt adjuvants, however, a pretreatment step (i.e., adjuvant dissolution or antigen desorption) is required, which may compromise antigen integrity during sample handling. Methods: This work describes the development of three competitive ELISAs to measure the relative D-antigen content of aluminum-salt adjuvanted PV-VLPs (Types 1, 2, 3) without the need for pretreatment. Results: First, key assay parameters were established, including specificity, accuracy, precision, linearity, limit of quantification, and stability-indication. Next, preformulation characterization studies were performed with these methods including (1) rank-ordering the inherent thermal stability profiles of the PV-VLPs (Types 1 > 3 > 2) in-solution and adsorbed to an aluminum phosphate adjuvant (AdjuPhos™, AP) and (2) determining the effect of formulation variables on the thermal stability profiles of AP-adsorbed PV-VLPs including antimicrobial preservatives (thimerosal, 2-PE) and five different antigens present in pediatric combination vaccines (D, T, wP, Hib, Hep B). Conclusions: The development and application of three competitive D-antigen ELISAs were demonstrated, and future use in formulation and storage stability studies with the AP-adjuvanted, trivalent PV-VLPs (Types 1, 2, 3) is discussed with the long-term goal to develop a stable, efficacious, multi-dose, hexavalent combination vaccine presentation. Full article