Search Results (12)

Background/Objectives: Acute respiratory infections represent a global health burden due to their high incidence, morbidity and mortality. Despite advances in prevention and treatment, strategies providing broad protection against respiratory pathogens remain limited. This study evaluated the antiviral activity of nasal spray forms based on an antigen complex from opportunistic bacteria, with or without a mucoadhesive copolymer, and their influence on cellular and humoral components of the immune response. Methods: Antiviral activity was assessed in experimental models of SARS-CoV-2 infection in Syrian hamsters and influenza A pneumonia in mice under prophylactic and therapeutic–prophylactic regimens. Parameters of cellular and humoral immune response were assessed using delayed-type hypersensitivity, antibody-forming cell assays, and leukocyte phagocytic activity. Results: Both forms showed comparable influence on cellular and humoral components of the immune response. In a mouse model of lethal influenza pneumonia induced by A/California/04/2009 (H1N1)pdm09 virus, intranasal administration at 100 µg/kg increased mean survival time and survival rate, and reduced body weight loss. In SARS-CoV-2-infected hamsters, both forms reduced clinical signs, body weight loss, weight lung index, lung pathology, and decreased viral titers. The copolymer-containing form showed the most pronounced effect under the therapeutic–prophylactic regimen, with greater protection against body weight loss and stronger suppression of viral replication. Conclusions: The findings support further investigation of these nasal spray forms as candidates for prophylaxis and adjunctive therapy of respiratory infections, particularly when pathogen variability may limit the effectiveness of pathogen-directed treatments. Further mechanistic studies are needed to clarify the pathways underlying the observed antiviral effect. Full article

Verticillium dahliae is a devastating soil-borne fungal pathogen that causes severe yield losses in melon (Cucumis melo L.) and other crops. Identifying novel resistance genes is crucial for sustainable disease management. In this study, we characterized the function of CmVQ23, a candidate gene previously identified through QTL mapping, in mediating defense against V. dahliae using heterologous expression in Arabidopsis thaliana. Subcellular localization assays revealed that the CmVQ23-eGFP fusion protein predominantly localized to the nucleus, consistent with its predicted role as a co-factor of transcription factor. Upon V. dahliae inoculation, CmVQ23-overexpressing Arabidopsis lines exhibited significantly reduced disease indices and restricted fungal proliferation compared with wild-type and mutant plants, although these lines displayed altered vegetative growth, including delayed bolting and reduced plant height. Mechanistically, CmVQ23 overexpression promoted reactive oxygen species (ROS) accumulation and hypersensitive response (HR)-mediated cell death at infection sites, as evidenced by intensified DAB and trypan blue staining. Furthermore, transgenic lines maintained higher photosynthetic efficiency, enhanced antioxidant enzyme activities, and increased lignin deposition via upregulation of phenylalanine ammonia-lyase (PAL) and polyphenol oxidase (PPO). Notably, CmVQ23 overexpression markedly upregulated both salicylic acid (SA)- and jasmonic acid/ethylene (JA/ET)-responsive marker genes, including AtPR1, AtPR2, AtPR5, AtPAD4, AtPDF1.2, and AtVSP2 upon infection. Collectively, these findings demonstrate that CmVQ23 functions as a positive regulator of resistance to Verticillium dahliae by orchestrating ROS/HR-mediated cell death, antioxidant defense, phenylpropanoid pathway activation, and phytohormone signaling crosstalk, offering a promising genetic resource for improving Verticillium wilt resistance in crops. Full article

Latent tuberculosis infection (LTBI) represents a major global health concern as it constitutes the principal reservoir for future tuberculosis (TB) disease. Its identification is particularly important in Bacille Calmette–Guérin (BCG)-vaccinated populations, where cross-reactivity of purified protein derivative limits the specificity of the tuberculin skin test and hampers targeted preventive therapy. Early Mycobacterium tuberculosis antigens encoded within the RD1 region, especially ESAT-6, CFP-10 and TB7.7, have enabled the development of antigen-specific interferon-gamma release assays (IGRAs) and recombinant skin tests with improved BCG-independent specificity. This narrative review integrates and critically appraises current evidence on the immunobiological properties of early and latency-associated antigens, the cellular mechanisms underlying T-cell-dependent immune reactivity, and the diagnostic performance of IGRAs and ESAT-6/CFP-10-based skin tests, rather than merely summarizing individual studies. Although these platforms rely on different assay principles (in vitro cytokine release versus in vivo delayed-type hypersensitivity), both measure antigen-specific T-cell memory and do not define the biological stage of infection or reliably distinguish latent from incipient or active TB. Across most adult populations, IGRAs demonstrate high specificity and acceptable sensitivity, whereas reduced sensitivity and higher rates of indeterminate results are observed in young children and immunocompromised individuals. ESAT-6/CFP-10-based skin tests show diagnostic accuracy comparable to IGRAs and may offer operational advantages in resource-limited settings. Latency-associated antigens and host biomarkers such as IP-10, together with multi-analyte immune signatures, represent promising avenues for improving diagnostic sensitivity and prognostic stratification but currently lack sufficient validation for routine clinical use. Overall, RD1-encoded antigens remain central to LTBI immunodiagnosis, while future research should focus on developing stage-resolving and prognostic biomarkers, optimized antigen panels, and standardized interpretive frameworks. Full article

Penicillium expansum, a major postharvest pathogen, causes blue mold decay in apples, resulting in substantial economic losses and mycotoxin contamination. Despite the importance of effector proteins in fungal pathogenicity, the role of metalloproteases in P. expansum remains unclear. Here, we characterize an effector candidate, PeMep, through whole genome sequencing and functional analyses. Functional validation confirmed the secretory capacity of its signal peptide via yeast assays and subcellular localization. Deletion of PeMep significantly impaired fungal growth (23% reduction), conidiation (23.3% decrease), and germination efficiency. The ΔPeMep mutant exhibited hypersensitivity to osmotic, oxidative, and thermal stresses, highlighting its vital role in environmental adaptability. Importantly, pathogenicity assays revealed attenuated virulence in the ΔPeMep mutant, with 15–30% smaller lesion sizes on apples and delayed hyphal penetration compared to the wild-type, demonstrating that PeMep is essential for the pathogenic process of P. expansum 3.3703. These findings identify PeMep as a potential multifunctional effector protein crucial for fungal development, environmental adaptation, and pathogenicity in P. expansum 3.3703, providing a novel target for postharvest disease management. Full article

The dysregulation of immune responses are responsible for the development of several diseases, such as allergic and autoimmune diseases. The medications used to treat these conditions have numerous side effects, creating the need for new drugs. Physalins are natural compounds with various pharmacological activities already described. Here, we aimed to investigate the immunomodulatory effects of physalin F in mouse splenocytes and in a delayed-type hypersensitivity (DTH) model. In a cytotoxicity assay, physalin F had low cytotoxicity to mouse splenocytes in concentrations equal to or below 2 µM. It significantly inhibited lymphocyte proliferation in a concentration-dependent manner and reduced the production of cytokines, including IL-2, IL-4, IL-10, and IFN-γ, in activated splenocytes. The combined therapy of physalin F with dexamethasone was investigated in vitro, showing a synergistic action of the two compounds. Mechanistically, physalin F reduced calcineurin activity in concanavalin A-stimulated splenocyte cultures. Finally, in vivo, the intraperitoneal administration of physalin F in a DTH model reduced paw edema induced by bovine serum albumin immunization. Our results demonstrate the potential of physalin F as an immunosuppressive agent, to be used alone or in combination with glucocorticoids. Full article

Delayed-type hypersensitivity (DTH) is caused by a broad number of drugs used in clinic, and antineoplastic drugs show an elevated proportion of DTH, which potentially affects the quality of life of patients. Despite the serious problem and the negative economic impact deriving from market withdrawal of such drugs and high hospitalization costs, nowadays, there are no standard validated methods in vitro or in vivo to evaluate the sensitizing potential of drugs in the preclinical phase. Enhanced predictions in preclinical safety evaluations are really important, and for that reason, the aim of our work is to adapt in vitro DPRA, ARE-Nrf2 luciferase KeratinoSens^TM, and hCLAT assays for the study of the sensitizing potential of antineoplastic agents grouped by mechanism of action. Our results reveal that the above tests are in vitro techniques able to predict the sensitizing potential of the tested antineoplastics. Moreover, this is the first time that the inhibition of the VEGFR1 pathway has been identified as a potential trigger of DTH. Full article

Previously, the delayed-type hypersensitivity (DTH) cutaneous test with the spike protein of SARS-CoV-2 has been shown to be a simple in vivo method to measure T-cell functionality after natural infection and in vaccinated individuals. Methods: Twenty-five kidney-transplanted recipients were immunized with two doses of the mRNA-based Pfizer–BioNTech COVID19 vaccine three weeks apart. Cell-immune response (CIR) was evaluated ten weeks later using an in vivo DTH skin test and in vitro with an interferon gamma release assay (IGRA). Humoral Immune Response (HIR) was determined by the measurement of specific IgG anti-S1 SARS-CoV-2. Results: Ten weeks after the second dose of the vaccine, 23 out of 25 transplanted patients had a positive DTH skin test, while in vitro CIR was considered positive in 20 patients. Unspecific stimulation was positive in all 25 patients, showing no T-cell defect. Seven out of twenty-five patients had a negative specific anti-spike IgG. CIR was positive in all immune-competent control patients. Conclusions: DTH is a useful, simple, and cheaper tool that can be used to assess cellular immune response, with an excellent correlation with the in vitro CIR. CIR assessment after vaccination in these immunocompromised patients is an excellent complement to HIR-based methods. This skin test could be used if classical in vitro methods cannot be applied. Full article

Deeper understanding of mold-induced cytokine signatures could promote advances in the diagnosis and treatment of invasive mycoses and mold-associated hypersensitivity syndromes. Currently, most T-cellular immunoassays in medical mycology require the isolation of mononuclear cells and have limited robustness and practicability, hampering their broader applicability in clinical practice. Therefore, we developed a simple, cost-efficient whole blood (WB) assay with dual α-CD28 and α-CD49d co-stimulation to quantify cytokine secretion in response to Aspergillus fumigatus antigens. Dual co-stimulation strongly enhanced A. fumigatus-induced release of T-cellular signature cytokines detectable by enzyme-linked immunosorbent assay (ELISA) or a multiplex cytokine assay. Furthermore, T-cell-dependent activation and cytokine response of innate immune cells was captured by the assay. The protocol consistently showed little technical variation and high robustness to pre-analytic delays of up to 8 h. Stimulation with an A. fumigatus lysate elicited at least 7-fold greater median concentrations of key T-helper cell signature cytokines, including IL-17 and the type 2 T-helper cell cytokines IL-4 and IL-5 in WB samples from patients with Aspergillus-associated lung pathologies versus patients with non-mold-related lung diseases, suggesting high discriminatory power of the assay. These results position WB-ELISA with dual co-stimulation as a simple, accurate, and robust immunoassay for translational applications, encouraging further evaluation as a platform to monitor host immunity to opportunistic pathogens. Full article

Antigen-specific regulatory T cells play key immune suppressive roles in autoimmune disease models and regulate the peripheral tolerance achieved via anterior chamber-associated immune deviation (ACAID). Articular cartilage has type II collagen (CII), which is a potent autoantigen protein in arthritis. There has not been much research on the clinical importance of CII-associated diseases. Moreover, the capability of CII to induce immune tolerance has not been previously assessed. We reported that delivery of CII either directly into the eye or via intravenous injection of CII-specific ACAID antigen presenting cells (APCs) can induce ACAID. Here, we hypothesized that peripheral tolerance can be induced following adoptive transfer of in vitro generated CII-specific ACAID B cells to naive mice. Delayed hypersensitivity (DTH) assays were used to assess the suppressive ability of adoptively transferred B cells. Immune responses of ACAID B cell-injected mice were significantly suppressed following challenges with CII as compared to positive controls. This effect was replicated in three different strains of mice (C57BL/6, BALB/c, and DBA/1). Thus, CII-specific ACAID B cells were able to induce immune tolerance in Th1-skewed, Th2-skewed, and arthritis-prone mice. ACAID B cell-mediated tolerance induced by CII could have therapeutic implications for the treatment of CII-mediated autoimmune diseases. Full article

Since antigens are negatively charged, they combine well with positively charged adjuvants. Here, ovalbumin (OVA) (0.1 mg·mL⁻¹) and poly (diallyldimethylammonium chloride) (PDDA) (0.01 mg·mL⁻¹) yielded PDDA/OVA assemblies characterized by dynamic light scattering (DLS) and scanning electron microscopy (SEM) as spherical nanoparticles (NPs) of 170 ± 4 nm hydrodynamic diameter, 30 ± 2 mV of zeta-potential and 0.11 ± 0.01 of polydispersity. Mice immunization with the NPs elicited high OVA-specific IgG1 and low OVA-specific IgG2a production, indicating a Th-2 response. Delayed-type hypersensitivity reaction (DTH) was low and comparable to the one elicited by Al(OH)₃/OVA, suggesting again a Th-2 response. PDDA advantages as an adjuvant were simplicity (a single-component adjuvant), low concentration needed (0.01 mg·mL⁻¹ PDDA) combined with antigen yielding neglectable cytotoxicity, and high stability of PDDA/OVA dispersions. The NPs elicited much higher OVA-specific antibodies production than Al(OH)₃/OVA. In vivo, the nano-metric size possibly assured antigen presentation by antigen-presenting cells (APC) at the lymph nodes, in contrast to the location of Al(OH)₃/OVA microparticles at the site of injection for longer periods with stimulation of local dendritic cells. In the future, it will be interesting to evaluate combinations of the antigen with NPs carrying both PDDA and elicitors of the Th-1 response. Full article

Food intolerance is delayed adverse food reactions which follow consumption of specific foods. The underlying mechanisms are not well understood, but food intolerance is often considered as a type 2 hypersensitivity reaction mediated by immunoglobulin G (IgG) antibody. To understand the causes of food intolerance, it is important to investigate sensitization patterns of food-specific IgGs (sIgG) in relation to dietary patterns and physical conditions. Conventional approaches to measure serological IgGs often require large volumes of serum, thus are not suitable for highly multiplexed assays. To overcome this impracticality, we developed a highly sensitive method to screen the sIgGs and other antibody isotypes against 66 antigens with minimal amount of serums. We prepared a microarray by immobilizing food antigens on activated glass slides. Human sera and their dietary information were obtained from 30 subjects. Aliquots (200 nl) of sera were analyzed against 66 food antigens in parallel. sIgG levels were determined and analyzed in relation to subjects’ dietary patterns. The levels of antibody isotypes were also examined to understand the relationship between allergy and food intolerance. The developed microarray showed exceptional performances in antibody screening and demonstrated the potential to be used as an automated assay system. Full article

Response surface methodology was employed to optimize the conditions for alkaline extraction of polysaccharides from Ganoderma lucidum. The results indicated that the optimum conditions were an extraction temperature of 60.1 ºC, an extraction time of 77.3 min, a sodium hydroxide (NaOH) concentration of 5.1% and a substrate/liquid ratio of 1:21.4. Immunological assays results have shown that the alkaline soluble polysaccharides have no noticeable effects on monocyte phagocytosis and immune organ (spleen, thymus) weight of of immunocompromised mice at the tested dosages. However, they could restore delayed type hypersensitivity reaction to dinitrofluorobenzene (DFNB), hemolysis antibody levels at the three doses applied, and improve the natural killer cell activity at the high-dose and medium dose. Full article