Search Results (636)

Aflatoxin B1 (AFB1), a highly carcinogenic mycotoxin, has been the focus of research for the development of efficient detection methods. In this study, a novel magnetic immuno-ratiometric fluorescent sensing system was constructed for the quantitative detection of AFB1. Green-emitting carbon quantum dots were conjugated with AFB1 monoclonal antibody to obtain GCDs@AFB1 mAb, and AFB1 oxime was immobilized on Fe₃O₄ magnetic microspheres to prepare AFB1-Ox@Fe₃O₄ NPs. After the immune-competitive adsorption of GCDs@AFB1 mAb by AFB1-Ox@Fe₃O₄ NPs and free AFB1, magnetic separation was performed. Red fluorescent silver nanoclusters were introduced as an internal reference to construct a GCDs-AgNCs ratiometric fluorescent system. The sensor exhibited a good linear response in the range of 0~240 pg/mL with a low limit of detection of 18 pg/mL and excellent selectivity. The spiked recoveries in real samples ranged from 92.14% to 110.02%, with a relative standard deviation of 0.57% to 4.58%. Combining the specific antigen–antibody recognition with magnetic separation technology, this method addresses the issues of poor stability and high environmental interference of traditional fluorescent sensors, and provides a new strategy for the sensitive and stable detection of AFB1. Full article

Background/Objectives: Cholera remains an important global health problem. Inactivated oral cholera vaccines (OCVs) are essential in the WHO/GTFCC (World Health Organization/Global Task Force on Cholera Control) strategy to end cholera by 2030; however, global supply is insufficient, they require partial cold-chain storage, and their formulation and antigen contents leave room for improvement. We describe here the development and preclinical evaluation of DuoChol OCV, a next-generation thermostable oral vaccine designed to address these gaps. Methods: DuoChol is a lyophilized dry-powder formulation in enteric capsules containing formalin-inactivated Vibrio cholerae O1 El Tor Ogawa and Inaba isogenic bacteria, recombinant cholera toxin B subunit (rCTB), and sucrose as stabilizer. Methods describe the construction of the novel vaccine strains, processes for the preparation and characterization of vaccine components, and the final dry formulation in enteric capsules, and in vitro and in vivo vaccine stability analyses. Results: The newly engineered vaccine strains, together with a high-yield mixed-mode chromatography process for rCTB purification, enabled efficient and cost-effective vaccine production. Stability studies demonstrated complete preservation of O1 LPS and rCTB antigens for at least 21 months across temperatures of 4–40 °C. Moreover, regardless of storage duration or temperature, oral immunization of mice with DuoChol elicited strong serum and mucosal antibacterial and antitoxin responses that were similar to those induced by the licensed Dukoral^® OCV. Conclusions: Its heat stability, practical enteric capsule formulation, and potential for improved efficacy compared to inactivated whole-cell only OCVs support positioning DuoChol as a promising next-generation OCV, suitable for national cholera control programs and particularly advantageous for outbreak response, where rapid deployment and early, robust protection are essential. Full article

Simple and non-invasive transdermal vaccination is an attractive alternative to conventional injection-based immunization. However, the effectiveness of transdermal vaccines is often constrained by the stratum corneum barrier. Although the use of solid-in-oil (S/O) nanodispersion technology has successfully facilitated skin permeation to induce an immunological response, the antibody titers remain suboptimal. Herein, a dectin-1 selective ligand, laminarin, was used as an immunostimulatory adjuvant to enhance the immune response. S/O nanodispersions loaded with laminarin and ovalbumin (OVA) were systematically developed and characterized in terms of particle size, in vitro OVA release behavior, and skin permeation performance using excised mouse skin. In vivo immunization via transcutaneous administration was performed to evaluate biocompatibility and antigen-specific immunoglobulin-G (IgG) responses. Laminarin-loaded S/O nanodispersions demonstrated long-term stability and efficient ex vivo skin permeability. All the prepared laminarin-loaded S/O nanodispersions showed increased OVA-specific IgG responses compared with the laminarin-free S/O formulation. Among the formulations, the S/O nanodispersion containing OVA and laminarin at a 1:4 weight ratio induced 20-fold higher OVA-specific IgG responses than PBS and 7-fold higher responses than laminarin-free S/O formulations. This study clearly demonstrates the potential of laminarin-loaded S/O nanodispersions as a non-invasive vaccine delivery platform for enhancing antigen-specific antibody responses. Full article

Background: Lipopolysaccharide (LPS) functions as a Toll-like receptor 4 (TLR4) agonist that triggers innate immunity; however, structural variations between pathogenic and commensal bacteria distinctly influence its immunostimulatory profile. This study evaluated the immunostimulatory activity of LPS derived from the commensal bacterium Hafnia alvei and explored its potential as an exploratory vaccine adjuvant. Methods: Cytokine induction was evaluated in immune cells across diverse host species, and receptor activation was assessed via reporter assays. To investigate in vivo immunogenicity and preliminary tolerability, H. alvei LPS was formulated into a prototype oil-in-water (O/W) emulsion utilizing ovalbumin (OVA) as a model antigen. Results: LPS from H. alvei strain BA2000346 exhibited immunostimulatory activity comparable to that of Escherichia coli, while inducing greater TNF-α expression than pathogenic Salmonella and Pseudomonas strains. Distinct from E. coli LPS, it demonstrated the capacity to activate both TLR4 and the mannose-recognizing Dectin-2 receptor in reporter systems. This cytokine induction was consistent across various strains and host species. Furthermore, the prototype O/W emulsion formulation enhanced antigen-specific humoral and cellular immune responses while demonstrating preliminary tolerability based on body-weight monitoring and visual clinical observation. Conclusions: H. alvei-derived LPS exhibits TLR4 and Dectin-2 agonist activity in vitro. When synergized with an O/W emulsion delivery system, it provides a preliminary indication of cross-species stimulatory potential and supports further investigation as a hypothesis-generating platform for future vaccine adjuvant development. Full article

Background: The aim of the study was to investigate whether red blood cell antigens (A, B, D, C^w, C, c, E, e, K, k, Jk^a, Jk^b, Fy^a, Fy^b, Le^a, Le^b, M, N, S, s, and P1) from clinically relevant blood group systems were associated with susceptibility to COVID-19. Methods: This exploratory case-control study was carried out on a group of 263 donors from the Regional Center for Transfusion Medicine, Bialystok, Poland (including 121 convalescents and 142 healthy subjects). The microplate method was applied to examine the expression of 21 antigens from eight clinically relevant systems in the donors’ red blood cells. Results: No significant correlation was found between any single blood group and susceptibility to COVID-19. For a more detailed analysis, we adopted an approach involving 3-, 4- and 5-element mutual combinations of antigens. In this exploratory analysis of multi-antigen combinations, nominal statistical significance was found for a number of associations, but none remained statistically significant after adjustment for multiple testing. In the case of three-antigen associations, the strongest association was observed for cc combined with Ee and kk, with the effect size showing approximately 4× higher odds of COVID-19 (OR = 4.49, p = 0.020); in the case of four-antigen combinations, the strongest association was found for RhD(+) combined with kk, Fy^b and P1(−), as well as RhD(+) combined with cc, Ee and kk, also indicating 4× higher odds of COVID-19 (OR = 4.49 p = 0.016). For five-antigen combinations, the strongest association was observed for blood type O combined with kk, Le^b, P1(+) and MM, with the association strength reaching an OR = 4.44, p = 0.025. Conclusions: The findings suggest that analyses based solely on single blood group antigens may have limited value for assessing the susceptibility to COVID-19. In contrast, combinations of red blood cell antigens may provide more reliable correlations with the susceptibility to COVID-19. However, these findings should be interpreted with caution and require confirmation on larger and more representative populations. Full article

Background and Objectives: Blood group antigens are well known for their importance in transfusion medicine and transplant compatibility; however, their biological role extends beyond these functions and includes associations with the risk of several diseases. In this study, we investigated the relationship between ABO blood groups and the circulating levels of 73 different molecules. Patients and Methods: Fifty-six healthy donors were enrolled, including 24 individuals with blood group O, 19 with blood group A, and 13 with blood group B. Blood samples were collected and analyzed in a single laboratory using Luminex fluorescent bead-based assay panels to determine the concentrations of 73 circulating molecules. Depending on data distribution, ANOVA or Kruskal–Wallis tests and Student’s t-test or Kolmogorov–Smirnov tests were applied to identify significant differences among groups. Associations were further assessed by binary logistic regression analysis. Results: Subjects with blood group A showed significantly higher circulating levels of IL-1R1, IL-13, IL-23, PDGF-BB, VEGF-A, VEGF-D, soluble VEGF-R2 (KDR), soluble VEGF-R3 (FLT-4), VLA-4, CD141, MMP-1, syndecan-1 (SDC-1), and mannose-binding lectin (MBL) compared with the other blood groups. In contrast, individuals with blood group B exhibited significantly higher levels of IL-22, IL-23, PDGF-BB, CD62P (P-selectin), and amyloid β1–42. Several significant associations were identified by logistic regression analysis. Conclusions: Our findings indicate that ABO blood groups are associated with distinct circulating molecular profiles, supporting the existence of biological differences that may contribute to variations in disease susceptibility among individuals with different blood types. Nevertheless, given the exploratory’s nature and limited sample size of this study, further investigations are required to validate these findings, confirm the observed associations, and clarify their potential clinical implications. Full article

Background/Objectives: Pseudomonas aeruginosa is a major cause of healthcare-associated infections, and the global rise of multidrug-resistant (MDR) strains has created an urgent need for alternative therapeutics. R- and F-type pyocins are phage tail-like bacteriocins that selectively kill P. aeruginosa by binding to lipopolysaccharide (LPS) receptors. We characterized O-serotype distribution and pyocin susceptibility among clinical isolates from central Taiwan to evaluate their therapeutic potential. Methods: A total of 109 ICU-derived P. aeruginosa isolates were analyzed. O-serotypes were determined by PCR, and pyocin gene carriage was confirmed by sequencing. Purified R1, R2, R5, F1, F2, F4, F7, and F12 pyocins were tested using spot assays. LPS profiles were examined by SDS-PAGE to explore structural correlates of resistance. Synergistic effects of combined R- and F-type pyocins were assessed in MDR isolates. Results: The most prevalent serotypes were O6 (23.9%), O2/O5/O16/O18/O20 (20.2%), O1 (16.5%), and O11/O17 (15.6%). Susceptibility was strongly serotype-dependent: O1 and O6 were highly sensitive to both pyocin types, whereas the O2/O5/O16/O18/O20 group showed marked resistance. SDS-PAGE demonstrated that resistant isolates possessed densely packed long-chain O-antigens, likely shielding LPS core receptors from pyocin binding. F-type pyocins exhibited bactericidal activity comparable to R-types, and R/F pyocin cocktails produced synergistic killing against MDR isolates. Conclusions: These findings provide an updated serotype profile of P. aeruginosa in Taiwan and highlight the importance of LPS structural variability in pyocin susceptibility. These results underscore the potential of pyocin-based cocktails as a promising precision-medicine strategy to inhibit the planktonic growth and biofilm formation of multidrug-resistant P. aeruginosa isolates. Full article

Atypical Salmonella enterica strains that evade conventional detection pose significant challenges to food safety surveillance. A hydrogen sulfide (H₂S)-negative and serologically untypable S. enterica strain (SF1060) was detected by qPCR from cooked farmed mussels in Galicia, Spain, and characterized using phenotypic and genomic approaches. Despite typical biochemical profiles, SF1060 failed to produce black colonies on Xylose Lysine Deoxycholate (XLD) agar and lacked detectable somatic antigens by conventional serotyping. Hybrid genome assembly using nanopore and illumina sequencing yielded a closed chromosome and five plasmids. In silico analyses identified the strain as S. Senftenberg ST14. Comparative genomics revealed a chromosomal inversion at the rfb operon (encoding enzymes needed to synthesize deoxysugars and O antigens) mediated by IS5-family transposase ISEc68, which truncated the rfbD gene and separated the remaining rfb genes at rfbD, disrupting O-antigen biosynthesis, explaining the inconclusive phenotypic serotyping results. The phs operon responsible for H₂S production lacked premature stop codons, suggesting the H₂S-negative phenotype may result from an alternative mechanism. This study demonstrates how whole-genome sequencing resolves identification of atypical strains that fail culture-based detection and emphasizes the critical need for molecular surveillance methods in seafood safety programs, particularly in regions where atypical S. enterica variants may be endemic. Full article

O-linked glycosylation comprises distinct regulatory systems, including secretory-pathway mucin-type O-GalNAc glycosylation and intracellular O-GlcNAcylation. These modifications both target serine/threonine residues but differ in glycan structure, cellular compartment, enzymatic machinery, and biological function. This narrative review was based on targeted searches of PubMed, Web of Science, and related literature using keywords related to O-glycosylation, O-GalNAc glycosylation, O-GlcNAcylation, immune regulation, cell signaling, glycoproteomics, and congenital disorders of glycosylation (CDG). We summarize evidence that mucin-type O-glycosylation regulates receptor behavior, cell adhesion, immune checkpoints, immunoglobulin function, antigen recognition, and pathogen–host interactions, whereas O-GlcNAcylation mainly modulates intracellular signaling, transcriptional control, stress responses, post-translational modification crosstalk, and innate immune pathways. We also discuss how glycosylation defects, including CDG and selected O-linked glycosylation disorders, connect genetic variation with disease phenotypes. Recent advances in site-specific glycoproteomics, O-glycoprotease-assisted workflows, LC–MS/MS-based glycopeptide analysis, and spatial or temporal profiling have improved mechanistic interpretation but still face limitations in site localization, structural resolution, and functional validation. Overall, the evidence supports the hypothesis that distinct O-linked glycosylation systems act through different molecular mechanisms but converge on signaling regulation, immune homeostasis, and disease susceptibility. Full article

Acute respiratory diseases caused by viral pathogens such as Influenza A continue to represent a major global health challenge, emphasizing the need for rapid, sensitive, and accessible diagnostic tools. In this work, a carbon screen-printed electrode (SPCE)-based electrochemical immunosensor for the detection of an Influenza A (H1) antigen is reported, incorporating a comparative electrochemical evaluation of four electrode materials. Fe₃O₄ nanoparticles, Fe₃O₄@C nanoparticles, graphene quantum dots (GQDs), and gold nanoparticles (AuNPs) were systematically assessed by cyclic voltammetry to evaluate their electrocatalytic performance. The highest electrochemical response was selected for biosensor construction. The immunosensor was fabricated by immobilizing antibodies on a modified SPCE and characterized using differential pulse voltammetry (DPV). A concentration-dependent response was observed for H1 antigen concentrations ranging from 0 to 300 ng/mL, with a minimum detectable concentration (MDC) of 1 ng/mL and limit of detection (LOD) of 176 ng/mL and 45 ng/mL for serum and nasopharyngeal swabs, respectively. The biosensor performance was specifically evaluated in complex biological fluids, demonstrating reproducible performance and moderate selectivity against non-target influenza subtypes. Overall, this study highlights the critical role of electrode material selection in determining electrochemical immunosensor performance and supports the potential of SPCE-based platforms for the screening of an Influenza A (H1) antigen in point-of-care-oriented applications. Full article

Magnetic polymeric microspheres are pivotal solid-phase carriers in chemiluminescence enzyme immunoassays (CLEIA). However, their practical clinical application is frequently hindered by non-specific adsorption, irreversible aggregation, and the intrinsic susceptibility of exposed outermost Fe₃O₄ nanoparticles to oxidation. To overcome these critical bottlenecks, we rationally engineered highly original monodisperse P(St-MMA)@Fe₃O₄@P(St-MMA) sandwich-structured microspheres. The bespoke amphiphilic outer shell acts as an impenetrable shield against hydration and oxidation, while maintaining a topologically size-matched mesoporous network (average pore size of 13.11 nm) for optimal antibody anchoring. Strikingly, this architecture ensures exceptional long-term colloidal stability, completely preventing macroscopic agglomeration for over six months in buffer solutions. When evaluated in a carcinoembryonic antigen (CEA), CLEIA, our microspheres achieved an ultra-low limit of detection (LOD) of 0.055 ng·mL⁻¹ and high analytical recovery (93.37–108.25%). In a head-to-head comparison with industry-standard commercial magnetic beads, the engineered microspheres delivered stronger chemiluminescent signals and lower background noise, demonstrating excellent intra-assay (CV < 4.37%) and inter-assay (CV < 10%) precision. This work establishes a scalable, highly stable materials platform that effectively resolves the persistent oxidation limitations, holding immense practical importance for next-generation ultrasensitive clinical in vitro diagnostics. Full article

Klebsiella pneumoniae is a member of the six highly virulent and antibiotic-resistant bacterial pathogens group (ESKAPE) and poses a significant threat to public health due to its ability to cause both hospital and community-acquired infections. Recent health concerns have emerged about heat-tolerant bacterial contamination in hospital settings, particularly those associated with infant formula preparation. This study aims to evaluate the heat survival of 10 clinical K. pneumoniae strains in infant formula and to investigate the correlation between heat tolerance and the presence of heat shock resistance genes, particularly the clp family of ATPases. Ten strains of K. pneumoniae were exposed to heat at 55 °C for 30 min in infant formula. We assessed their survival rates and determined their D-values. Additionally, we screened for the presence of clpC family genes across representative strains. A wide variation in heat tolerance was observed among the strains. Strain 1701 (ST247, capsular antigen profile O3:K1) exhibited the highest heat tolerance, with a D-value of 12.9 min at 55 °C. The other strains exhibited moderate-to-low heat tolerance. Notably, strain 1701 was the only one that contained the clpC2 gene, suggesting a potential association between the clp gene family and heat resistance. Our results indicate that specific heat shock resistance genes, such as clpC2, may be associated with enhanced heat tolerance observed in K. pneumoniae strains. These findings highlight the potential role of heat shock proteins in bacterial persistence within neonatal healthcare environments. Full article

Glioblastoma (GBM) is characterized by a profoundly immunosuppressive tumor microenvironment that limits the efficacy of conventional and immune-based therapies. Oncolytic herpes simplex virus (oHSV) therapy has emerged as a strategy capable of both tumor-selective infection and immune microenvironment modulation. This systematic review aimed to synthesize current evidence on how oHSV therapy reshapes the immunosuppressive microenvironment of GBM. A systematic search of PubMed (MEDLINE) and Embase identified original studies published between 2016 and 2025 investigating immunological or microenvironmental effects of oHSV-based therapies in GBM (final search: 28 January 2026). Preclinical and clinical studies were included, whereas reviews, editorials, conference abstracts, and studies of non-herpes oncolytic viruses were excluded. Study selection and data extraction were performed independently by two reviewers. Due to heterogeneity in models, viral constructs, and outcome measures, a qualitative narrative synthesis was conducted. Of 214 records, 22 studies met the inclusion criteria. Most were preclinical studies using orthotopic immunocompetent murine models, with limited clinical evidence including an early-phase trial in recurrent high-grade glioma. Therapeutic efficacy frequently correlated with tumor microenvironment remodeling rather than viral replication alone. oHSV infection promoted inflammatory signaling, antigen presentation, macrophage polarization, and effector T-cell recruitment but also induced counter-regulatory mechanisms such as myeloid-mediated immunosuppression and vascular or stromal barriers. The clinical significance and durability of these effects remain to be established in larger prospective studies. Despite heterogeneous designs and limited clinical data, current evidence suggests that oHSV may function as a platform for immune microenvironment reprogramming in glioblastoma. Full article

The Republic of Korea has implemented routine vaccination against foot-and-mouth disease virus (FMDV) in livestock using a bivalent vaccine comprising serotypes O and A following the massive FMD outbreak in 2010, while antigens for the remaining serotypes are maintained in overseas antigen banks. The recent geographic expansion of FMDV Southern African Territories 1 (SAT1) beyond Africa underscores the need for enhanced preparedness in previously unaffected regions. In this study, we evaluated the SAT1 BOT-R strain as a candidate vaccine seed for potential domestic vaccine production by optimizing antigen production conditions, assessing scalability, determining virus inactivation parameters, and examining immunogenicity in pigs. Optimal antigen yield was achieved at 20 h−24 h post infection with a multiplicity of infection of 0.005−0.01, with production remaining stable under mildly alkaline conditions. Antigen productivity was consistently maintained during scale-up from shake flasks to a bioreactor, yielding up to 9.5 μg/mL. Complete virus inactivation was achieved using binary ethylenimine at 2 mM for 24 h at 26 °C. Vaccines formulated from both flask- and bioreactor-derived antigens elicited comparable neutralizing antibody responses in pigs, reaching a median titer of 1:500 following booster immunization. Collectively, these findings demonstrate that the SAT1 BOT-R strain is a viable and scalable candidate for SAT1 antigen banking and future domestic vaccine production, providing a practical framework for strengthening national preparedness against potential incursions of FMDV SAT1. Full article

Background/Objectives: Extraintestinal pathogenic Escherichia coli (ExPEC) is a major pathogen that causes septicemia, meningitis, and polyserositis in pigs. The increasing prevalence of antimicrobial resistance and the diverse serotypes of ExPEC highlight the urgent need for broadly protective vaccines. Methods and Results: In this study, an OmpC epitope vaccine based on the T4 phage display system was developed and evaluated. Two B-cell epitopes (OmpC-1 and OmpC-2) were identified by bioinformatic analysis and displayed on recombinant T4 phages. Immunization induced strong antigen-specific IgG responses, with the OmpC-1-T4 group showing significantly higher antibody titers than the OmpC protein group. In the O11 serotype PCN033 challenge model, survival rates reached 100% in the OmpC-1-T4 group, 60% in the OmpC-2-T4 group, and approximately 80% in the OmpC protein group. In the O18 serotype 2103 challenge model, both recombinant phage groups had survival rates of approximately 60%, whereas all the mice in the OmpC protein group died within three days. OmpC-1-T4 immunization also significantly reduced bacterial loads in lung and brain tissues after PCN033 infection and decreased TNF-α and IL-6 expression in lung tissues, accompanied by reduced inflammatory infiltration and tissue damage. Conclusions: Overall, the T4 phage-displayed OmpC epitope vaccine induced strong humoral immunity and provided protection against different ExPEC serotypes. Among the candidates, OmpC-1-T4 showed superior immune protection, bacterial clearance, and inflammation control, supporting its potential as a vaccine candidate against porcine ExPEC infection. Full article