Search Results (2,365)

Feline calicivirus (FCV) is a major pathogen that threatens feline health worldwide. Its global prevalence, extensive genetic variability, and limited cross-protection among strains present significant challenges for vaccine development. In this study, an infectious clone of the FCV-GDJM202201 strain was constructed using the eukaryotic expression plasmid pcDNA3.1 under the control of the cytomegalovirus (CMV) promoter. The rescued virus, rGDJM-A4822T, exhibited growth kinetics comparable to those of the parental strain in vitro. Subsequently, two recombinant viruses, rGDJM-VP1_JL and rGDJM-VP1_SH, were generated by replacing the VP1 gene in the GDJM202201 backbone with those from heterologous FCV strains. Notably, these recombinant viruses exhibited reduced viral titers compared to rGDJM-A4822T. Finally, neutralization assays revealed differential neutralizing antibody titers among the recombinant FCVs, with rGDJM-A4822T inducing higher neutralizing antibody titers and cross-neutralizing activity. Collectively, this study establishes an FCV infectious clone that can be used to rescue recombinant viruses carrying heterologous VP1 proteins and to evaluate neutralizing antibody responses. Full article

Background: Respiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract infections in infants, seniors, and immunocompromised individuals, contributing substantially to the global disease burden. Given the limited preventive options available, developing an effective and safe vaccine remains a public health priority. Methods: An mRNA vaccine encoding the RSV PreF protein was designed and prepared. Antigen properties were evaluated in silico, and the coding sequence was optimized using NLP algorithms. The stability and translational efficiency of the mRNA constructs were verified through in vitro and in vivo assays, followed by immunogenicity evaluation of the formulated mRNA vaccines in a BALB/c mouse model. Results: The optimized mRNA showed predicted improvements in structural stability and a lower free energy state, which were associated with increased translational efficacy in vitro. Correct antigen conformation and retention of key epitopes were confirmed by intracellular staining followed by flow cytometry. A balanced Th1-biased immune response was induced in mice, characterized by high levels of neutralizing antibodies and antigen-specific T-cell immunity, along with enhanced memory T-cell proliferation and differentiation, indicating long-term immunological memory. Conclusions: A novel RSV PreF mRNA vaccine was successfully developed via optimization of protein structure and mRNA sequence. Superior immunogenicity was demonstrated in the BALB/c mouse model, together with promising potential in terms of vaccine safety and immunological persistence. These findings represent a promising step forward in the pursuit of an effective RSV vaccine and suggest the potential of the developed mRNA vaccine to induce substantial immune responses that may correlate with protection in future challenge studies. Full article

Porcine circovirus type 2 (PCV2) is a pathogen of major importance in swine that is characterized by ongoing genetic evolution. To provide an updated epidemiological assessment for China, our study analyzed 1051 clinical samples collected from 27 provincial-level regions between 2023 and 2024. The overall PCV2 positivity rate was 65.18%, with detection rates showing significant seasonal variation, with higher rates in spring and summer. Genotypic analysis of 379 open reading frame 2 (ORF2) sequences identified PCV2d as the dominant genotype (78.89%), and no significant geographic clustering was observed. Coinfection with porcine reproductive and respiratory syndrome virus (PRRSV) is common, yet statistical tests have revealed an epidemiologically independent relationship between the two viruses. Notably, analysis of the capsid (Cap) protein revealed that high-frequency amino acid mutations were concentrated in immunodominant loop regions. These mutations resulted in genotype-specific substitutions within key neutralizing epitopes. This study provides the latest large-scale national baseline data on PCV2 in China for 2023–2024. It systematically analyzes the epidemiological characteristics of the dominant PCV2d genotype in the post-African Swine Fever era, the patterns of antigenic epitope mutations in the Cap protein, and their potential impact on vaccine efficacy. The study fills a gap in recent national epidemiological data on PCV2 in China and provides a basis for the targeted prevention and control of PCV2 and the updating of vaccine strains. Full article

While current influenza vaccines often lack broad protection against antigenically drifted strains, some modified hemagglutinin (HA) protein antigens have shown promise in eliciting broadly neutralizing antibodies against conserved epitopes. During infection, the mildly acidic environment of the late endosome triggers irreversible HA conformational changes resulting in a post-fusion structure with altered antigenicity. While enhancing the stability of other structural class I viral fusion protein antigens has been instrumental in improving the effectiveness of COVID-19 and RSV vaccines, the role of HA stability in influenza vaccine immunogenicity is relatively unclear. Here, we used the nucleoside-modified mRNA-LNP platform to test engineered HA antigens with specific acid-stabilizing mutations (E47K, K58I, R106K, and K153E) in the HA stalk. All mutations increased HA acid stability, but E47K and R106K did not increase immunogenicity. K153E and K58I, but not E47K and R106K, enhanced the cell-surface expression of the HA protein in vitro. In mice, K153E- and K58I-containing mRNA-LNP vaccines elicited increased neutralizing antibody titers against homologous virus. K153E conferred greater protection than wild-type vaccine against lethal heterologous A/PR/8/34 challenge at low doses (0.5–1.0 µg), despite the absence of neutralizing antibodies against the challenge strain. K153E also elicited greater expansion of antigen-specific antibody-secreting cells (ASCs) in the bone marrow, as well as cross-reactive T follicular helper (Tfh) cells in the spleen. For the vaccines studied, increased HA expression was a stronger correlate of mRNA-LNP enhancement than increased HA stability. Full article

Background/Objectives: Our previous study demonstrated that while the third SARS-CoV-2 booster effectively enhanced immunity against the Delta subvariant, its protection declined over time. This study aimed to evaluate and compare the humoral and cellular immune responses, as well as reactogenicity, of the mRNA-1273 vaccine administered as a fourth booster in healthy Thai adults previously vaccinated with two doses of CoronaVac (CV) followed by a third dose of either AZD1222 (AZ) or BNT162b2 (BNT). Methods: Participants received a single 100 µg (0.5 mL) intramuscular dose of mRNA-1273. Blood samples were collected at baseline (D0), D14, D90, and D180 to assess anti-RBD IgG, conduct a surrogate virus neutralization test (sVNT) against the Delta and Omicron variants, and assess IFN-γ levels and reactogenicity. Results: Both 2CV/AZ- and 2CV/BNT-primed groups exhibited comparable local and systemic reactogenicity. The fourth mRNA-1273 dose markedly increased Delta variant inhibition within 14 days in both groups and remained at high levels at Days 90 and 180. sVNT inhibition against Omicron rose similarly in both groups at Day 14; it declined sharply by Days 90 and 180, with the 2CV/AZ-primed group showing significantly lower levels than the 2CV/BNT-primed group. Baseline anti-RBD IgG levels were lower in the 2CV/AZ group (p = 0.003) but surpassed those of the 2CV/BNT group by Day 14, with no significant differences at later time points. IFN-γ responses followed a similar pattern to anti-RBD IgG Conclusions: A heterologous fourth mRNA-1273 booster in both 2CV/AZ- and 2CV/BNT-primed groups effectively enhances B-cell and T-cell responses against SARS-CoV-2. However, emerging variants such as Omicron may still pose challenges. The trial was registered with the Thai Clinical Trials Registry: the name of the registry: “The comparison of immune response to the 4th dose booster with mRNA-1273 COVID-19 vaccine in individuals who had received 2 doses of CoronaVac and booster with ChAdOx-1 or BNT162b2 COVID-19 vaccine”, TCTR20220205002 on 5 February 2022. Full article

Antiretroviral therapy (ART) effectively controls Human Immunodeficiency Virus Type-1 (HIV-1) infection in people with HIV-1 (PWH), preventing the progression of their infections to AIDS. However, as PWH age, they experience lifestyle- and age-related diseases, notably various types of cancer beyond those traditionally associated with AIDS, with greater incidence and mortality than their non-HIV-1-positive counterparts, despite effective arrest of HIV-1 infection by ART. Dysregulation of redox enzymes presents an underexplored linkage between HIV-1 infection and cancer comorbidity, impacting reactive oxygen/nitrogen species (RONS) management, inflammation, immune function, and mitochondrial function. Chronic HIV-1 infection increases both RONS production and RONS neutralization responses, accelerating development of a sustained RONS-rich environment that still possesses sufficient dampening to prevent outright cytotoxic effects. Such an environment promotes both tumor proliferation and resistance adaptations to chemo- and radiotherapies. This review considers the effects of chronic HIV-1 infection on redox enzyme function and links these effects to tumorigenic mechanisms as potentially shared pathways. We then examine current methods of modulating redox function, consider how these could potentially impact both HIV-1 infection and cancer progression, and lastly propose future methods of co-treatment that could be explored. Full article

Background/Objectives: Bovine viral diarrhea (BVD) is a major infectious disease of cattle caused by bovine viral diarrhea virus genotypes 1 and 2 (BVDV-1 and BVDV-2). Current inactivated and live attenuated vaccines provide incomplete cross-genotype protection and may exhibit limitations related to durability of immunity or safety. This study evaluated whether co-expression of the BVDV envelope glycoproteins E1 and E2 in a Modified Vaccinia Ankara (MVA) vector could support antigen expression and induce immune responses in a proof-of-concept model. Methods: Recombinant Modified Vaccinia Ankara (MVA) viruses expressing BVDV-1 E1E2 or BVDV-2 E1E2 were generated by homologous recombination. Recombinant viruses were purified and characterized for antigen expression, genetic stability, and growth properties in vitro. Immunogenicity was evaluated in a BALB/c mouse model by measuring E2-specific antibody responses, virus-neutralizing antibodies, and antigen-responsive cellular immune responses. Results: Both recombinant MVA constructs showed detectable E2 expression when E1 and E2 were co-expressed, and exhibited growth characteristics comparable to parental MVA with stable maintenance after serial passage. In contrast, recombinant MVA expressing E2 alone did not yield detectable E2 protein under the same experimental conditions. Immunization induced detectable humoral and cellular immune responses, including E2-specific IgG antibodies, virus-neutralizing antibodies, and increased frequencies of antigen-responsive CD8⁺ T cells with a tendency toward a Th1-biased profile. Conclusions: These findings indicate that co-expression of BVDV E1 and E2 in an MVA vector can support detectable antigen expression and induce measurable immune responses in a mouse proof-of-concept model. Further studies in cattle, including challenge experiments, will be required to determine the protective efficacy and practical applicability of this platform for BVDV vaccine development. Full article

Background/Objectives: Seasonal waves of respiratory viruses—including SARS-CoV-2, influenza A virus (IAV), and respiratory syncytial virus (RSV)—continue to pose a global health burden and highlight the need for antiviral agents that are effective, safe, broadly active, affordable, and widely accessible. Current interventions are limited by the need for their early administration, the risk of resistance, their costs, and the restricted availability in large parts of the world. For certain natural products, such as European black elderberry (Sambucus nigra L.) fruit extract (ElderCraft^®; EC) and the seaweed-derived sulfated polymer iota-carrageenan (IC), antiviral activities against respiratory viruses, particularly IAV and SARS-CoV-2, have previously been shown. Here, we assessed the antiviral activity of IC and an anthocyanin-standardized EC extract against SARS-CoV-2, IAV, and RSV, either as monotherapy or in multiple-dose combinations. Methods: MDCKII cells were infected with IAV_PR8, human Calu-3 lung epithelial cells with the SARS-CoV-2 Omicron variant, and HEp-2 cells with RSV (A2 strain). Inhibitors were administered either by pre-incubation of cell-free virions prior to infection or, in separate time-of-addition experiments, during or post-infection. Viral replication was quantified by qRT-PCR or intracellular immunostaining. Cytotoxicity was evaluated using a neutral red uptake assay. Results: Most intriguingly, both EC and IC are able to neutralize virions derived from SARS-CoV-2, IAV, or RSV extracellularly in a dose-dependent manner. Notably, EC and IC alone exhibited strong anti-RSV activity, which was not reported previously. Most importantly, combined treatment with IC and EC caused a pronounced synergistic antiviral effect against the tested viruses, as confirmed by the Bliss independence model, without any detectable impact on cell viability. Finally, solutions prepared from matrix-standardized mono- or combi-lozenges, containing IC and/or EC in high or low doses, reproduced the antiviral and synergistic combination effects observed with the pure compounds. Conclusions: In summary, these findings support further development of EC and IC as a topically accessible, virion-neutralizing combination (e.g., lozenges) to provide additional protection against major respiratory viruses and potentially strengthen pandemic preparedness. Full article

Background/Objectives: Prevention of virus-related cancers is a multifaceted process shaped by vaccination and public awareness. This study assessed awareness of virus–cancer relationships and willingness to vaccinate against a cancer-associated virus among medical and non-medical students. We also evaluated whether human papillomavirus (HPV)-vaccinated students demonstrate greater awareness of the HPV-cancer link compared to unvaccinated students, and examined willingness to vaccinate against a certain cancer-associated virus according to HPV vaccination status. Methods: This cross-sectional survey was conducted in Poland (October 2023–June 2024) and included 1013 first- and second-year university students recruited via convenience sampling. Participation was voluntary and anonymous. Results: Awareness of virus–cancer relationships was low, ranging from 19% for Epstein–Barr virus-related cancers to 43.8% for HPV-related cervical cancer. Women were more likely than men to recognize the HPV–cervical cancer link (OR = 2.08, p < 0.001), supporting gender differences and the need for gender-neutral HPV education with targeted strategies for men. Medical students demonstrated higher awareness than non-medical students. HPV vaccination coverage was low (14.5%), with higher uptake among medical students (21.2% vs. 8.2%). Notably, 41.3% of non-medical students and 7.5% of medical students had never heard of HPV vaccination. Willingness to vaccinate against a cancer-associated virus varied according to perceived infection risk. Conclusions: These findings highlight the need for targeted educational interventions to improve awareness of HPV–cancer links and risk perception, as well as to ensure ongoing education of both HPV-vaccinated and unvaccinated individuals to support informed health decisions and vaccine acceptance. Full article

Avian influenza infections cause substantial economic losses in the poultry industry and raise public health concerns due to viral adaptation and cross-species transmission. The frequent antigenic drift of influenza viruses further complicates the prevention and treatment of avian respiratory infections. In this study, we generated high-affinity heavy-chain variable domain (VHH) nanobodies from naïve alpaca/camelid VHH libraries using phage display combined with H9N2 influenza A virus (IAV)-infected Madin-Darby Canine Kidney (MDCK) cells. Based on binding affinity and neutralization potential, we identified seven hemagglutinin (HA)-specific and two neuraminidase (NA)-specific VHHs. Molecular docking predicted the interaction sites of HA-specific VHHs (L1-2, L1-4, A5) and NA-specific VHHs (L1-3, L2-2), providing mechanistic insights. Notably, the three HA-specific VHHs (L1-2, L1-4, A5) showed cross-reactivity to representative HA subtypes (H1, H3, and influenza B), indicating recognition of conserved epitopes across divergent influenza strains. For the first time, these camelid nanobodies were fused to the chicken IgY Fc domain, and the expression cassette was integrated into the Saccharomyces cerevisiae genome, achieving a secretion yield of 15–20 mg/L of VHH-Fc antibodies. Experimental validation confirmed that the three HA-specific VHHs-Fc constructs effectively blocked viral infection, while the two NA-specific VHH-Fc constructs (L1-3, L2-2) inhibited NA activity, demonstrating the functional efficacy of the yeast-secreted VHH–IgY Fc platform. This novel IgY Fc fusion approach offers a scalable platform with enhanced stability, extended circulation potential, and applicability in poultry. Full article

Vaccination remains the core strategy for the prevention and control of Newcastle disease (ND). The inherent thermosensitivity of traditional Newcastle disease virus (NDV) vaccines imposes major limitations on their transportation, storage, and field application. To address these challenges, a novel liquid, thermostable, live ND vaccine was developed in the present study. Firstly, Tris/HCl buffer at near-neutral pH was identified as the optimal basic buffer system. On this basis, further screening and formulation optimization of vaccine stabilizers were conducted, and NDV strains with excellent thermal stability were used to verify the stability-conferring properties of the developed stabilizer. The results showed that the formulation composed of 0.5% gelatin, 4% trehalose, 0.1% L-glutamic acid, and 0.5% thiourea was confirmed as the optimal stabilizer for ND liquid vaccines. This formulation maintained the stable storage of the tested NDV for 12 months at 4 °C and exhibited promising stability for 30 days at 25 °C, marking a significant advancement toward development thermostable NDV vaccines that are independent of a continuous cold chain. More importantly, the liquid vaccine stored at 4 °C for 12 months still induced high levels of NDV-specific antibodies in specific pathogen-free chicks and provided 100% protective efficacy against challenge with virulent NDV. In conclusion, the liquid vaccine stabilizer developed in this study not only significantly enhanced the thermostability of the vaccine but also effectively maintained its immunogenicity, thereby providing an important theoretical basis for the research and development of liquid ND vaccines. Full article

Background: Rabies is a highly fatal zoonotic disease that causes approximately 59,000 human deaths worldwide each year. Current inactivated rabies vaccines require multiple doses and are associated with high costs. The full-length rabies virus glycoprotein (RVG), a membrane protein, exhibits substantial instability in its trimeric structure during recombinant expression. This instability makes it difficult to obtain high-purity, correctly folded antigens. Objectives: This study focuses on the preparation of a full-length recombinant RVG subunit vaccine candidate expressed in a glycoengineered Pichia pastoris system with mammalian-like glycosylation. Methods: The full-length RVG gene (including the transmembrane domain and cytoplasmic tail) from the Challenge Virus Standard-11 (CVS-11) strain was codon-optimized and inserted into the pPICZαA vector to construct the recombinant expression plasmid pPICZαA-RVG. The plasmid was transformed into glycoengineered Pichia pastoris X33-7 (low-mannose type) by electroporation for inducible expression. The target protein was purified by nickel affinity chromatography, anion-exchange chromatography, and Superdex-200 size-exclusion chromatography. The structural characteristics of the purified protein were analyzed by dynamic light scattering (DLS) and transmission electron microscopy (TEM). The purified antigen was formulated with the adjuvants AS03 or MF59. BALB/c mice (n = 5 per group) were immunized intramuscularly following a four-dose schedule (days 0, 7, 14, and 28). Antigen-specific IgG antibody titers were measured by ELISA, and neutralizing antibody titers were determined using the rapid fluorescent focus inhibition test (RFFIT). Results: Glycoengineered Pichia pastoris yeast strains expressing wild-type RVG (RVG-WT) or a mutant variant (RVG-M6: R84S, R199S, H270P, R279S, K300S, and R463S) were successfully constructed. The purified RVG antigen formed nanoparticles with an average particle size of approximately 75 nm. Immunized mice generated robust RVG-specific IgG responses, with titers reaching approximately 6.31 × 10⁵ for RVG-WT after the fourth immunization, compared to 3.16 × 10³ for RVG-M6 and 5.62 × 10³ for the RVG-WT-PEG control. Two weeks after the fourth immunization, RVG-WT formulated with AS03 or MF59 induced significant neutralizing antibody responses compared with the control group (p < 0.0001 and p < 0.01, respectively). The neutralizing antibody titers reached 1:79.43 in the AS03 group and 1:33.11 in the MF59 group, whereas the WT-PEG + AS03 control group showed a low titer of 1:3.72. In contrast, RVG-M6 formulated with MF59 failed to induce detectable neutralizing antibodies (1:3.02). Furthermore, RVG-WT + AS03 induced significantly higher neutralizing antibody responses than the WT-PEG + AS03 control group (p < 0.0001), and a significant difference was also observed between the RVG-WT + MF59 and RVG-M6 + MF59 groups (p < 0.01). Conclusions: The glycoengineered Pichia pastoris expression system successfully produced uniform full-length rabies virus glycoprotein nanoparticles with high purity. When formulated with the AS03 adjuvant, RVG-WT induced high-titer neutralizing antibodies in mice, suggesting a promising strategy for the development of recombinant subunit vaccines against rabies. However, this study is limited by the absence of challenge studies and validation in target animal species, which will be further investigated in future work. Full article

Gene therapy using recombinant adeno-associated virus (rAAV) vectors has emerged as a transformative therapeutic modality for genetic disorders, demonstrating high transduction efficiency and a generally favorable safety profile during pre-clinical development. However, serious adverse events, including thrombotic microangiopathy, acute respiratory distress syndrome, hepatotoxicity, myocarditis, cytokine storm, and hemophagocytic lymphohistiocytosis, have been observed across multiple gene therapy clinical trials. Significant efforts have been made to understand the toxicities that cause these adverse events and clinical care for patients receiving gene therapies has evolved to mitigate their effects. These toxicities arise from a complex interplay between the innate and adaptive immune responses directed against the viral capsid and transgene products and are often compounded by pre-existing anti-AAV immunity. Immunomodulatory strategies have been developed to combat these responses to improve the long-term success of gene therapies, and this review provides clinicians managing gene therapy patients with an overview of mechanisms underlying AAV-associated immunotoxicities and a discussion of syndromes and mitigation strategies that have been reported in the clinical care of patients. Full article

Background/Objectives: Pathogenic mammarenaviruses cause severe hemorrhagic and neurologic disease in humans. Machupo virus (MACV), a New World (NW) mammarenavirus, causes Bolivian hemorrhagic fever in humans, and there are no approved vaccines. Methods: Here, we describe and compare the immunogenicity of three vaccines expressing the MACV glycoprotein complex (GPC) in C57BL/6 mice: a recombinant vesicular stomatitis virus (rVSV) and two different lipid nanoparticle (LNP)-encapsulated nucleoside-modified mRNA (mRNA-LNP) vaccines. The first mRNA-LNP vaccine, designated MACV mRNA, expresses the full-length MACV GPC. The second mRNA-LNP vaccine, called MACV VLP mRNA, encodes MACV GPC with appended sequences that induce the budding of virus-like particles (VLPs) with MACV GPC on the surface. This is the first description of any mRNA-LNP vaccine for MACV and the first comparison of mRNA and rVSVs as vaccine candidates for MACV. Results: We find that two doses of either MACV mRNA or MACV VLP mRNA are required for the induction of robust humoral and cellular immune responses including total MACV GPC IgG, neutralizing antibodies, cross-reactive antibodies that bind the related Junín virus GPC, and MACV-specific T-cell responses. To further investigate vaccination strategies for MACV, we also evaluated a heterologous prime-boost regimen involving the MACV mRNA vaccine coupled with the rVSV-based MACV vaccine. We find that the highest levels of MACV GPC-specific IgG and neutralizing titers were achieved when heterologous mRNA and rVSV prime-boost regimens were employed. Conclusions: These results elucidate differences in the immune response to different vaccine platforms for MACV and can inform future vaccine development for NW arenaviruses. Full article

Background: Highly pathogenic avian influenza H5N1 viruses of clade 2.3.4.4b have spread globally since 2021, causing extensive outbreaks in avian populations and repeated spillovers into diverse mammalian hosts, including humans. These cross-species transmission events highlight ongoing pandemic risks and underscore the need for vaccine strategies that reflect viral evolution at the human–animal interface. Despite the availability of licensed H5 vaccines and newly recommended World Health Organization (WHO) candidate vaccine viruses (CVVs), the extent to which these vaccines elicit cross-reactive antibody responses against contemporary clade 2.3.4.4b viruses, including mammalian spillover isolates of avian origin, remains incompletely characterized. Method: In this study, ferret antisera were generated using four WHO-recommended H5 CVVs, including a clade 1 strain (A/Vietnam/1194/2004) and three clade 2.3.4.4b strains (A/Astrakhan/3212/2020, A/American wigeon/South Carolina/22-000345-001/2021, and A/Ezo red fox/Hokkaido/1/2022), formulated with alum adjuvant to reflect licensed vaccine formulation used in national preparedness programs. Antibody responses and cross-reactive activity were evaluated using hemagglutination inhibition (HI) and microneutralization (MN) assays against homologous vaccine strains and a feline-origin clade 2.3.4.4b H5N1 field isolate from Korea, A/Feline/Korea/SNU-01/2023. Results: Antisera induced by clade 2.3.4.4b CVVs showed cross-reactive antibody responses against homologous and heterologous clade 2.3.4.4b viruses and demonstrated measurable HI and MN responses against the feline-origin field isolate. In contrast, antisera raised against the clade 1 Vietnam CVV exhibited limited cross-reactivity against clade 2.3.4.4b viruses. Overall, clade 2.3.4.4b CVVs generally showed higher antibody responses than the clade 1 vaccine strain across multiple panels. Conclusions: These findings provide descriptive insights into antigenic differences between clade 1 and clade 2.3.4.4b viruses and support the antigenic relevance of clade 2.3.4.4b CVVs for contemporary H5N1 strains. This study highlights the importance of ongoing antigenic evaluation to inform vaccine strain selection within a One Health framework. Full article