Search Results (259)

Background/Objectives: Streptococcus suis (SS), an important zoonotic pathogen, has caused significant economic losses to the global pig industry. Existing commercial vaccines for SS mainly provide effective protection against a single serotype. Due to the existence of many serotypes and their robust immune evasion capabilities, the development of multi-component subunit vaccines or multi-epitope vaccines that provide effective cross-protection against different strains of SS is a key focus of current research. Methods: We applied two-dimensional electrophoresis (2-DE) and immunoblotting to screen for candidate immunogens among the immunogenic cell wall proteins of SS. BALB/c mice were immunized intradermally with a multi-component, multi-epitope vaccine. The vaccine’s safety and immunogenicity were assessed via clinical monitoring, antibody titer detection, cytokine assays, and survival curve analyses. Results: In this study, eight immunogenic cell wall proteins (GH25, Pk, PdhA, Ldh, ExoA, Pgk, MalX, and Dnak) were successfully identified using MALDI-TOF-MS, all of which could induce high IgG antibody titers. Based on the conservation and immunoprotection demonstrated by these eight protective antigenic proteins, PdhA, Ldh, and MalX were screened to construct a multi-component subunit vaccine as a candidate vaccine for providing cross-protection against SS isolates of multiple serotypes. Challenge studies showed that mice immunized with the multi-component subunit vaccine (PdhA, Ldh, and MalX) were protected against challenges with the SS2 virulent strain ZY05719 (62.5% protection) and the SSChz virulent strain CZ130302 (75% protection). Subsequently, we utilized immunoinformatics techniques to design a novel multi-epitope vaccine (MVPLM) derived from the immunogenic proteins PdhA, Ldh, and MalX. However, challenge tests revealed that the MVPLM offered limited protection against SS. Conclusions: These data demonstrate that a multi-component subunit vaccine composed of PdhA, Ldh, and MalX proteins shows promise as a candidate universal vaccine against multiple SS serotypes. Full article

Background/Objectives: Influenza A viruses—including highly pathogenic H5N1—remain a global threat due to rapid evolution, zoonoses, and pandemic potential. Strain-specific vaccines targeting variable antigens often yield limited, short-lived immunity. The HA receptor-binding domain (RBD), a functionally constrained and immunologically relevant region, is a promising target for broad and subtype-focused vaccines. We aimed to design multiepitope constructs targeting conserved HA-RBD and adjacent domains to elicit robust, durable, cross-protective responses. Methods: Extensive sequence analyses (>20,000 H5N1 and >190,000 influenza A sequences) were used to derive consensus sequences. Three HA-based candidates were developed: (i) EpitoCore-HA-VX, a multi-epitope construct containing CTL, HTL, and B-cell epitopes from the H5N1 HA-RBD; (ii) StructiRBD-HA-VX, incorporating a conformationally preserved RBD segment; and (iii) FusiCon-HA-VX, targeting the conserved HA fusion peptide shared across subtypes. Two external HA comparators—a 400-aa HA fragment and the literature-reported HA-13–263-Fd-His—were analyzed under the same pipeline. The workflow predicted epitopes; evaluated antigenicity, allergenicity, toxicity, conservation, and HLA coverage; generated AlphaFold models; performed TLR2/TLR4 docking with pyDockWEB; and carried out interface analysis with PDBsum; and C-ImmSim simulations. Results: Models suggested stable, energetically favorable TLR2/TLR4 interfaces supported by substantial binding surfaces and complementary electrostatic/desolvation profiles. Distinct docking patterns indicated receptor-binding flexibility. Immune simulations predicted strong humoral responses with modeled memory formation and, for the H5N1-focused designs, cytotoxic T-cell activity. All candidates and comparators were predicted to be antigenic, non-allergenic, and non-toxic, with combined HLA coverage approaching global breadth. Conclusions: This study compares three design strategies within a harmonized framework—epitope collation, structure-preserved RBD, and fusion-peptide targeting—while benchmarking against two HA comparators. EpitoCore-HA-VX and StructiRBD-HA-VX showed promise against diverse H5N1 isolates, whereas FusiCon-HA-VX supported cross-subtype coverage. As these findings are model-based, they should be interpreted qualitatively; nonetheless, the integrated, structure-guided approach provides an adaptable path for advancing targeted H5N1 and broader influenza A vaccine concepts. Full article

Background: The increasing prevalence of antibiotic-resistant Mycoplasma genitalium poses a significant challenge to global public health, necessitating the exploration of alternative therapeutic strategies, including vaccine development. Methods: In this study, we employed an immuno-informatics-based reverse vaccinology approach augmented with artificial intelligence-driven tools, to identify and characterize potential B-cell and T-cell epitopes from the hypothetical proteins (HPs) retrieved from the genome of the MG_G37T strain, a previously uncharacterized yet promising vaccine target. Using multiple softwares, a systematic pipeline was utilized to assess the sub-cellular localization, antigenicity, and allergenicity of the selected proteins. Results: Sub-cellular localization analysis identified the presence of several outer membrane and extracellular proteins in the genome of MG_G37T, indicating their surface association and accessibility to immune surveillance. Antigenicity and allergenicity prediction tools led to the identification of two top-scoring hypothetical proteins (fig|2097.71.peg.1 (UniProt ID: P22747) and fig|2097.70.peg.33 (UniProt ID: Q57081)) that demonstrated strong antigenic potential, non-allergenic properties, and suitability as vaccine candidates. Epitope mapping and structural modeling analyses further validated the immunogenic potential of these epitopes, highlighting their ability to interact with host immune components effectively. Comparative analyses with mouse allelic regions indicated the potential translational relevance of these predicted epitopes for preclinical studies. Conclusions: In particular, this study highlights the potential of these two hypothetical proteins as a promising vaccine candidate and provides a strong reason for experimental validation towards the design and development of effective vaccines to combat M. genitalium infections in the era of antimicrobial resistance. Full article

Canine parvovirus (CPV) is a virus of veterinary health significance and a member of the Parvoviridae family. Despite its clinical significance and global distribution, surveillance is often limited to cases serious enough to result in veterinary visit and/or hospitalization, thereby limiting our understanding of its evolution and diversity. In this study, we coupled wastewater surveillance (WWS), long-range polymerase chain reaction (PCR) and long-read sequencing and demonstrate the utility of this approach for community-level monitoring of parvovirus diversity. We screened archived viral concentrates from wastewater (WW) collected monthly from July 2022 to June 2023 as part of a previous virus surveillance study from a population of ~500,000 people in Maricopa County, Arizona, USA. Using long-range PCR, the coding-complete sequences (~4.5 kb) were amplified as single contigs and sequenced on a long-read sequencer (MinION). Reads were trimmed, assembled, and contigs subjected to a bioinformatics workflow that includes phylogenetics, immuno-informatics and protein structure modelling. The ~4.5 kb amplicons were amplified from all the samples and sequenced. Twelve contigs (length: 4555 nt to 4675 nt: GC%: 35% to 36%) were assembled from 86,858 trimmed and size-selected reads (length 4400 nt–4900 nt) and all typed as parvoviruses. Overall, there were 11 CPV variants (2a, 2b and 2c) and 1 feline panleukopenia virus (FPV) variant. The FPV was 100% similar in the VP2 genomic region to the 1964 Johnson snow leopard strain present in the Felocell vaccine, suggesting recent shedding post-vaccination. For the CPVs, our analysis showed multiple amino acid substitutions in the VP2 and NS1 proteins, suggestive of host immune pressure and viral adaptation, respectively. The CPV variants clustered predominantly with North and South American variants, suggesting transboundary viral movement and multiple CPV-2c transmission chains seem evident. To the best of our knowledge, we here document the first detection of vaccine-origin FPV in WW. We show the presence of CPV-2a, 2b and 2c in the population sampled and provide evidence that suggests transmission of CPVs across the Americas. Our results also show that WWS coupled with long-range PCR and long-read sequencing is a feasible population-level complement to clinical case surveillance that also facilitates detection of vaccine-origin virus variants. The model we demonstrate here for tracking parvoviruses can also be easily extended to other DNA viruses of human and veterinary health significance. Full article

This study investigated the role of molecular mimicry in the context of autoimmunity associated with viral infection, using SARS-CoV-2 as a model system. A bioinformatic analysis was performed to identify sequence homologies between the SARS-CoV-2 Spike (S) protein and the human proteome, with a specific focus on immunogenic regions to assess potential cross-reactivity. The analysis revealed homologous regions between the viral S protein and several human proteins, including DAAM2, CHL1, HAVR2/TIM3, FSTL1, FHOD3, MYO18A, EMILIN3, LAMP1, and αENaC, which are predicted to be recognizable by B-cell receptors. Such recognition could potentially lead to the production of autoreactive antibodies, which can contribute to the development of autoimmune diseases. Furthermore, the study examined potential autoreactive CD4+ T-cell responses to human protein autoepitopes that could be presented by HLA class II molecules. Several HLA class II genetic variants were computationally associated with a higher likelihood of cross-reactive immune reactions following COVID-19, including HLA-DPA1*01:03/DPB1*02:01, HLA-DPA1*02:01/DPB1*01:01, HLA-DPA1*02:01/DPB1*05:01, HLA-DPA1*02:01/DPB1*14:01, HLA-DQA1*01:02/DQB1*06:02, HLA-DRB1*04:01, HLA-DRB1*04:05, HLA-DRB1*07:01, and HLA-DRB1*15:01. Additionally, seven T helper cell autoepitopes (YSEILDKYFKNFDNG, ERTRFQTLLNELDRS, AERTRFQTLLNELDR, RERKVEAEVQAIQEQ, NAINIGLTVLPPPRT, PQSAVYSTGSNGILL, TIRIGIYIGAGICAG) were identified that could be implicated in autoimmune T-cell responses through presentation by class II HLA molecules. These findings highlight the utility of viral B- and T-cell epitope prediction for investigating molecular mimicry as a possible mechanism in virus-associated autoimmunity. Full article

Background: Tuberculosis (TB) remains a global health priority, with current interventions like the Bacille Calmette–Guérin (BCG) vaccine lacking efficacy against latent infection and drug-resistant strains. Novel vaccines targeting both latent and active TB are urgently needed. Objective: This study aims to design a multi-epitope vaccine (MEV) and evaluate its immunogenicity, structural stability, and interactions with toll-like receptor 2/4 (TLR-2/4) via computational biology approaches. Methods: We designed MEV using bioinformatics tools, prioritizing immunodominant epitopes from Mycobacterium tuberculosis antigens. Structural stability was optimized through disulfide engineering, and molecular docking/dynamics simulations were used to analyze interactions and conformational dynamics with TLR-2/4. Antigenicity, immunogenicity, population coverage, and immune responses were computationally assessed. Results: The MEV candidate, CP91110P, exhibited 86.18% predicted global human leukocyte antigen (HLA)-I/II coverage, high antigenicity (VaxiJen: 0.8789), and immunogenicity (IEDB: 4.40091), with favorable stability (instability index: 33.48) and solubility (0.485). Tertiary structure analysis indicated that 98.34% residues were located in favored regions. Molecular docking suggested strong TLR-2 (−1535.9 kcal/mol) and TLR-4 (−1672.5 kcal/mol) binding. Molecular dynamics simulations indicated stable TLR-2 interactions (RMSD: 6–8 Å; Rg: 38.50–39.50 Å) and flexible TLR-4 binding (RMSD: 2–6 Å; Rg: 33–36 Å). Principal component analysis, free energy landscapes, and dynamic cross-correlation matrix analyses highlighted TLR-2’s structural coherence versus TLR-4’s adaptive flexibility. Immune simulations predicted potential robust natural killer cell activation, T helper 1 polarization (interferon-gamma/interleukin-2 dominance), and elevated IgM/IgG levels. Conclusions: CP91110P is predicted to stably bind to TLR-2 and flexibly interact with TLR-4, with prediction of its high antigenicity and broad coverage across immune populations. However, this conclusion requires confirmation through experimental validation. Therefore, it may provide a promising candidate for experimental validation in the development of tuberculosis vaccines. Full article

Background: Helcococcus kunzii, a facultative anaerobe and Gram-positive coccus, has been documented as a cunning pathogen, mainly in immunocompromised individuals, as evidenced by recent clinical and microbiological reports. It has been associated with a variety of polymicrobial infections, comprising diabetic foot ulcers, prosthetic joint infections, osteomyelitis, endocarditis, and bloodstream infections. Despite its emerging clinical relevance, no licensed vaccine or targeted immunotherapy currently exists for H. kunzii, and its rising resistance to conventional antibiotics presents a growing public health concern. Objectives: In this study, we employed an integrated subtractive proteomics and immunoinformatics pipeline to design a multi-epitope subunit vaccine (MEV) candidate against H. kunzii. Initially, pan-proteome analysis identified non-redundant, essential, non-homologous, and virulent proteins suitable for therapeutic targeting. Methods/Results: From these, two highly conserved and surface-accessible proteins, cell division protein FtsZ and peptidoglycan glycosyltransferase FtsW, were selected as promising vaccine targets. Comprehensive epitope prediction identified nine cytotoxic T-lymphocyte (CTL), five helper T-lymphocyte (HTL), and two linear B-cell (LBL) epitopes, which were rationally assembled into a 397-amino-acid-long chimeric construct. The construct was designed using appropriate linkers and adjuvanted with the cholera toxin B (CTB) subunit (NCBI accession: AND74811.1) to enhance immunogenicity. Molecular docking and dynamics simulations revealed persistent and high-affinity ties amongst the MEV and essential immune receptors, indicating a durable ability to elicit an immune reaction. In silico immune dynamic simulations predicted vigorous B- and T-cell-mediated immune responses. Codon optimization and computer-aided cloning into the E. coli K12 host employing the pET-28a(+) vector suggested high translational efficiency and suitability for bacterial expression. Conclusions: Overall, this computationally designed MEV demonstrates favorable immunological and physicochemical properties, and presents a durable candidate for subsequent in vitro and in vivo validation against H. kunzii-associated infections. Full article

Background: While most Escherichia coli strains are harmless members of the gastrointestinal microbiota, certain pathogenic variants can cause severe intestinal and extraintestinal diseases. A notable outbreak of E. coli O104:H4, involving both enteroaggregative (EAEC) and enterohemorrhagic (EHEC) strains, occurred in Europe, resulting in symptoms ranging from bloody diarrhea to life-threatening colitis and hemolytic uremic syndrome (HUS). Since treatment options remain limited and have changed little over the past 40 years, there is an urgent need for an effective vaccine. Such a vaccine would offer major public health and economic benefits by preventing severe infections and reducing outbreak-related costs. A multiepitope vaccine approach, enabled by advances in immunoinformatics, offers a promising strategy for targeting HUS-causing E. coli (O104:H4 and O157:H7 serotypes) with minimal disruption to normal microbiota. This study aimed to design an immunogenic multiepitope vaccine (MEV) construct using bioinformatics and immunoinformatic tools. Methods and Results: Comparative proteomic analysis identified 672 proteins unique to E. coli O104:H4, excluding proteins shared with the nonpathogenic E. coli K-12-MG1655 strain and those shorter than 100 amino acids. Subcellular localization (P-SORTb) identified 17 extracellular or outer membrane proteins. Four proteins were selected as vaccine candidates based on transmembrane domains (TMHMM), antigenicity (VaxiJen), and conservation among EHEC strains. Epitope prediction revealed ten B-cell, four cytotoxic T-cell, and three helper T-cell epitopes. Four MEVs with different adjuvants were designed and assessed for solubility, stability, and antigenicity. Structural refinement (GALAXY) and docking studies confirmed strong interaction with Toll-Like Receptor 4 (TLR4). In silico immune simulations (C-ImmSim) indicated robust humoral and cellular immune responses. In Conclusions, the proposed MEV construct demonstrated promising immunogenicity and warrants further validation in experimental models. Full article

Novel waterfowl reoviruses (nWRVs) have been reported in several countries, but their circulation and genetic characteristics in Vietnam remain poorly understood. In this study, we investigated nWRVs in northern Vietnam through molecular detection, virus isolation, experimental infection in ducklings, and molecular analysis of the sigma C-encoding (sC) gene. We also applied immunoinformatic tools to explore the antigenic and structural features of the sC protein. nWRVs were detected in 15.6% of tested samples across ten provinces. Three isolates were successfully recovered, all showing a characteristic cytopathic effect—syncytium formation—in Vero cells. When tested in ducklings (n = 72), the isolates caused disease of varying severity, but all induced characteristic gross and microscopic lesions, particularly ecchymotic hemorrhages and large necrotic foci in the liver and spleen. Phylogenetic analysis based on sC sequences placed the Vietnamese isolates (n = 14) within the nWRV clade, with evidence of two genetically distinct groups. Our immunoinformatic analysis identified four predicted B-cell epitopes located in the head and body domains of the sC protein, with little variation. Full article

Background: Multi-epitope vaccines have become the preferred strategy for protection against infectious diseases by integrating multiple MHC-restricted T-cell and B-cell epitopes that elicit both humoral and cellular immune responses against pathogens. Computational methods address various aspects independently, yet their orchestration is technically challenging, as most bioinformatics tools are accessible through heterogeneous interfaces and lack interoperability features. The present work proposes a novel framework for rationalized multi-epitope vaccine design that streamlines end-to-end analyses through an integrated web-based environment. Results: VaccineDesigner is a comprehensive web-based framework that streamlines the design of protective epitope-based vaccines by seamlessly integrating computational methods for B-cell, CTL, and HTL epitope prediction. VaccineDesigner incorporates single-epitope prediction and evaluation as well as additional analyses, such as multi-epitope vaccine generation, estimation of population coverage, molecular mimicry, and proteasome cleavage. The functionalities are transparently integrated into a modular architecture, providing a single access point for rationalized, multi-epitope vaccine generation in a time- and cost-effective manner. Conclusions: VaccineDesigner is a web-based tool that identifies and evaluates candidate B-cell, CTL, and HTL epitopes and constructs a library of multi-epitope vaccines that combine strong immunogenic responses, safety, and broad population coverage. The source code is available under the academic license and freely accessible. Full article

Phospholipase A1 (Ves a 1), a major toxin from Vespa affinis venom, poses significant risks to allergic individuals. Nevertheless, the epitope determinants of Ves a 1 have not been characterized. Thus, identifying its linear B-cell epitopes is crucial for understanding envenomation mechanisms. In this study, we predicted and identified B-cell epitopes EP5 and EP6 as potential candidates. EP5 formed an α-helix at the active site of Ves a 1, whereas EP6 adopted an extended loop conformation. Both synthetic peptides were synthesized and evaluated for their inhibitory effects using immune-inhibitory assays with polyclonal antibodies (pAbs) targeting both native (nVes a 1) and recombinant (rVes a 1) forms. The Ves a 1 polyclonal antibodies (pAb-nVes a 1 and pAb-Ves a 1) were produced, and their specificity binding to Ves a 1 was confirmed by Western blot. Next, ELISA inhibition assays showed that EP5 and EP6 significantly blocked pAb binding to both nVes a 1 and rVes a 1. Dot blot and Western blot assays supported these findings, particularly with stronger inhibition toward rVes a 1. Furthermore, enzymatic assays indicated that nVes a 1 and rVes a 1 retained phospholipase activity. Immunoinformatics docking showed that EP5 and EP6 specifically bind to a single-chain variable fragment antibody (scFv) targeting Naja naja PLA2. Molecular analysis revealed similar amino acid interactions to the template, suggesting effective paratope–epitope binding. These results support the potential of EP5 and EP6 for future diagnosis and therapy of V. affinis venom allergy. Full article

Monkeypox virus (MPXV) has caused 148,892 confirmed cases and 341 deaths from 137 countries worldwide, as reported by the World Health Organization (WHO), highlighting the urgent need for effective vaccines to prevent the spread of MPXV. Traditional vaccine development is low-throughput, expensive, time consuming, and susceptible to reversion to virulence. Alternatively, a reverse vaccinology approach offers a rapid, efficient, and safer alternative for MPXV vaccine design. Here, MPXV proteins associated with viral infection were analyzed for immunogenic epitopes to design multi-epitope vaccines based on B-cell, CD4+, and CD8+ epitopes. Epitopes were selected based on allergenicity, antigenicity, and toxicity parameters. The prioritized epitopes were then combined via peptide linkers and N-terminally fused to various protein adjuvants, including PADRE, beta-defensin 3, 50S ribosomal protein L7/12, RS-09, and the cholera toxin B subunit (CTB). All vaccine constructs were computationally validated for physicochemical properties, antigenicity, allergenicity, safety, solubility, and structural stability. The three-dimensional structure of the selected construct was also predicted. Moreover, molecular docking and molecular dynamics (MD) simulations between the vaccine and the TLR-4 immune receptor demonstrated a strong and stable interaction. The vaccine construct was codon-optimized for high expression in the E. coli and was finally cloned in silico into the pET21a (+) vector. Collectively, these results could represent innovative tools for vaccine formulation against MPXV and be transformative for other infectious diseases. Full article

Fasciola gigantica (F. gigantica) is a vital parasite that causes fasciolosis. Liver fluke infections affect livestock animals, and the Fasciola species (Fasciola spp.) vaccine has been tested for many types of these diseases. Currently, computer-based vaccine design represents an attractive alternative for constructing vaccines. Thus, this study aimed to design the epitopes of linear B-cells (BCL) and helper T lymphocytes (HTL) using an immunoinformatic approach and to investigate in silico and the mice’s immune response. A non-conserved host region, overlapping F. gigantica cathepsin B proteins (FgCatB), and the highest conserved residue percentages were the criteria used to construct epitopes. The GPGPG linker was used to link epitopes in the multi-epitope Fasciola gigantica cathepsin B (MeFgCatB) peptide. The MeFgCatB peptide has high antigenicity, non-allergenicity, non-toxicity, good solubility, and a high-quality structure. The molecular docking between the MeFgCatB peptide and Toll-like receptor 2 (TLR-2) was evaluated. The IgM, IgG1, and IgG2 levels were elevated in silico. In mice, the MeFgCatB peptide was synthesized and administered as an injection. The MeFgCatB-specific IgG1 and IgG2a levels were elevated after week 2, showing a predominance of IgG1. The rFgCatB1, rFgCatB2, and rFgCatB3 were detected using the MeFgCatB peptide-immunized sera. The MeFgCatB peptide-immunized sera were detected at approximately 28–34 kDa in the whole body. In addition, the MeFgCatB immunized sera can positively signal at the caecal epithelium in the NEJ, 4WKJ, and adult stages. In summary, the MeFgCatB peptide is able to induce mixed Th1/Th2 immune responses with Th2 dominating and to detect the native protein of F. gigantica. The MeFgCatB peptide should help against F. gigantica in future experiments. Full article

Guillain-Barre syndrome is an autoimmune disease that provokes neural illness causing acute paralysis neuropathy. This syndrome appears after some bacterial infections produced by Campylobacter jejuni, Streptococcus pyogenes, S. pneumoniae, Haemophilus influenciae, E. coli and current studies showed the appears of this syndrome after SARS-CoV-2 infection. In this study, a in silico analysis was carry out in which to determinate bacterial epitopes than produce the molecule mimicry phenomena and that can produce the immune system activation against this epitope. A conserved amino acid sequence has been encountered with the highest probability to activate the immune system against this bacterial epitope, human gliomedin and ryanodine 3 type receptor. More studies needed to demonstrate in vivo the molecular mimicry in Guillain-Barre syndrome patients. Full article

Recent outbreaks caused by hMPXV, especially hMPXV lineages/sub-lineages, represent public health threats necessitating stringent prophylactic measures to ameliorate their colossal impact. The current study annotated the EEV form of hMPXV’s target proteins to formulate a reverse vaccinology-dependent hMPXV multiepitope vaccine. Epitope determination, followed by vaccine formulation, was undertaken. The promising formulation was validated for its potential to trigger immune responses immunoinformatically. The MPXV-1-Beta formulation was characterised as a promising candidate based on antigenicity score, physicochemical properties, solubility score, ProSA Z-score, and Ramachandran plot. Docking, normal mode analysis, and molecular dynamic simulation of MPXV-1-Beta with TLRs and MHCs authenticated rigid docking and its efficacy in enhancing immune receptor activation under physiological conditions. MPXV-1-Beta was discerned to trigger a sustained immune response (IR) with a broader average population coverage of 97.526, SD = 12.44. The proposed MPXV-1-Beta candidate showed significant potential. The findings of this study provide a preliminary framework for developing an efficacious hMPXV vaccine; however, extensive in vitro, in vivo, and clinical evaluations are required to substantiate the computational insights. Full article