Search Results (2,651)

A cell-based screening system for viral protease inhibitors was developed using firefly luciferase fragment complementation and validated on the SARS-CoV-2 3CLpro model. The optimal luciferase variant incorporating the VLQSGF proteolytic site (Luc III) retained 88% of its native activity. A critical requirement for system performance was the use of an extended nsp4–nsp6 fragment of the viral polyprotein rather than the mature protease, underscoring the importance of the native context for 3CLpro activity. The bicistronic construct pCAG-Luc-III-IRES-nsp4-6 enables coordinated expression of the reporter and protease, thereby increasing assay reproducibility. IC₅₀ values obtained in this system for nirmatrelvir and GC376 correlated with live-virus assay data but differed significantly from those of a cell-free FRET assay, reflecting the impact of cellular barriers. This approach combines simplicity, a standard substrate, and high reproducibility, making it promising for high-throughput screening in basic laboratory settings and adaptable to other viral proteases. Full article

Background/Objectives: The emergence of SARS-CoV-2 variants and breakthrough infections underscores the need for next-generation vaccines capable of protecting from natural infection and/or preventing virus transmission. Intranasal vaccination offers a promising approach by eliciting local immune responses in the nasal mucosa, the primary site of infection and reservoir for transmissible virus. We evaluated two live-attenuated, respiratory syncytial virus-vectored vaccines in which the RSV F and G surface glycoproteins were replaced with a chimeric SARS-CoV-2 Spike protein from the ancestral USA/WA-1/2020 strain (MV-014-212) or the Delta variant (MV-014-212-delta). Methods: K18-hACE2 mice and LVG Syrian hamsters were vaccinated with a single intranasal dose of MV-014-212 or MV-014-212-delta. Systemic and mucosal immunity were assessed following vaccination, and protection was evaluated following Delta SARS-CoV-2 challenge. In vaccinated hamsters, morbidity, viral shedding, and lung inflammation and injury were also assessed following natural exposure to infected cagemates. Results: A single intranasal dose of either vaccine elicited systemic and mucosal immunity in K18-hACE2 mice, including serum neutralizing antibodies, Spike-specific memory B cells and plasmablasts, and Spike-specific CD8⁺ lung-resident memory T cells. Although MV-014-212-delta vaccination provided the best protection against the Delta variant virus challenge, both vaccines decreased viral loads in nasal discharge, lung, and brain, and reduced weight loss and mortality. In naturally acquired infection studies, vaccinated hamsters exposed to infected cagemates exhibited minimal weight loss, limited viral replication within the nasal mucosa, and attenuated lung pathology. Conclusions: Intranasal RSV-vectored vaccines can elicit broad protective respiratory immunity, suggesting that this platform could be leveraged for other respiratory pathogens. Full article

Porcine epidemic diarrhea virus (PEDV) is the pathogen responsible for porcine epidemic diarrhea, causing significant economic losses to the swine industry. During the replication of PEDV, the genome mutates rapidly, making the effectiveness of commercial vaccines uncertain when facing newly emerging prevalent variants. More importantly, there are currently no safe and effective specific antiviral drugs available. YT1418, a pyrido[1,2-c]pyrimidin-1-one (PPO) compound, exhibited anti-PEDV activity in a previous study. To expand the chemical space of the PPO scaffold and clarify the influence of substituents at different positions on the antiviral activity of the compounds, 36 new compounds were designed and synthesized, and then their abilities to inhibit viral replication in a PEDV-infected cell model were evaluated. Furthermore, the hepatic microsomal metabolic stabilities of compounds with potent antiviral activity were assessed. The results showed that compounds N1 and N2 exhibited antiviral activity (EC₅₀ = 0.32, 0.37 μM, respectively) superior to that of YT1418, with selective index values of 43.78 and 42.89, respectively. Meanwhile, compound J4 demonstrated good hepatic microsomal stability and low cytotoxicity, which requires further investigation. This study identified lead compounds featuring a novel PPO core and established their structure–activity relationships, providing important insights for the development of anti-PEDV drugs. Full article

Dengue virus (DENV) and SARS-CoV-2 are emerging viral pathogens that share overlapping clinical features, including fever, fatigue, and respiratory symptoms, complicating differential diagnosis in endemic regions. Their co-circulation has increased the risk of co-infections, which may result in unpredictable disease progression, increased morbidity, and mortality. This overlap presents a significant challenge in managing outbreaks, as both viruses pose a major public health threat. Vaccines and direct-acting antivirals may be rendered ineffective by viral mutations, making it difficult to address evolving strains. Host-directed antivirals offer a promising alternative, potentially maintaining efficacy against a multitude of variants. Both DENV and SARS-CoV-2 rely on host proteases for viral maturation and entry, with furin playing a crucial role in viral glycoprotein cleavage. In DENV, furin cleaves the prM protein, facilitating virion maturation, while in SARS-CoV-2, the polybasic furin cleavage site in the spike protein enhances viral entry. This makes furin a compelling pan-viral target, where inhibiting furin could reduce viral fitness without relying on viral mutations. This review highlights the therapeutic rationale for targeting furin and discusses luteolin, a furin inhibitor showing antiviral activity against both viruses. Furin-targeted therapies may offer a durable antiviral strategy effective across DENV serotypes, SARS-CoV-2 variants, and co-infection settings. Full article

The COVID-19 pandemic highlighted the critical role of serological assays in understanding antiviral immune responses, monitoring vaccine efficacy, and informing public health strategies. This review provides a comprehensive overview of commonly used SARS-CoV-2 antibody detection methods, focusing on binding and neutralization assays. Antibody binding assays, including enzyme-linked immunosorbent assays (ELISAs), chemiluminescence immunoassays (CLIAs), lateral flow immunoassays (LFAs), and multiplex platforms, enable the rapid and high-throughput detection of immunoglobulin isotypes against various viral antigens. Neutralization assays, including live-virus, pseudovirus (PsV), and surrogate assays, offer functional insights into the ability of antibodies to prevent viral entry, though they often require higher biosafety levels and optimization. Serological assays, primarily antibody binding assays and several surrogate neutralization assays, received Emergency Use Authorization (EUA) during the pandemic, supporting seroprevalence efforts. Antibody binding assays and neutralization assays were also widely used in vaccine immunogenicity studies. Despite many standardization initiatives, assay standardization and data harmonization remain challenging and require further efforts. The choice of assay should be guided by study goals: antibody binding assays are preferred for high-throughput monitoring and epidemiological studies, while neutralization assays are essential for assessing functional immunity and variant-specific neutralization and protection. Full article

Background: Influenza A(H3N2) viruses remain a major public health concern due to their rapid antigenic evolution and association with severe disease, particularly among high-risk populations. During the 2025–2026 influenza season, a marked epidemiological shift was observed in Europe, with the emergence and predominance of the A(H3N2) subclade K (J.2.4.1). Objectives: This narrative review aims to provide an integrated overview of the epidemiology, evolutionary dynamics, and public health implications of subclade K, with a particular focus on its impact on vaccine effectiveness, in comparison with the 2024–2025 influenza season. Methods: A non-systematic literature review was conducted using major scientific databases and official public health sources, including WHO and ECDC reports. Recent surveillance data, genomic analyses, and epidemiological updates were included. Given the rapidly evolving evidence base, selected preprint studies were also considered and interpreted with caution. Results: The 2025–2026 influenza season in Europe was characterized by a relative genetic convergence, with subclade K accounting for the majority of A(H3N2) sequences. This variant demonstrated a clear selective advantage and was associated with an earlier and more intense epidemic peak. Molecular analyses indicate the accumulation of multiple mutations in the hemagglutinin protein, particularly within key antigenic sites, contributing to immune escape. These evolutionary changes have important implications for vaccine effectiveness, with current estimates suggesting moderate protection against infection but preserved effectiveness against severe outcomes. Antigenic mismatch, manufacturing constraints, and host-related factors further contribute to reduced vaccine performance. Conclusions: The emergence and rapid spread of subclade K highlight the dynamic nature of influenza virus evolution and its impact on public health. Continuous genomic surveillance and timely vaccine updates remain essential. Despite suboptimal effectiveness against infection, influenza vaccination continues to provide significant protection against severe disease and should remain a cornerstone of prevention strategies. Full article

Infectious bronchitis virus (IBV) is an avian coronavirus associated with respiratory, renal, and reproductive disease in chickens, with important economic consequences in intensive poultry production. This study aimed to estimate the seroprevalence of anti-IBV antibodies in vaccinated broiler flocks reared in south-western Romania and to characterize its distribution across counties and study years. Between 2018 and 2021, a total of 2466 blood samples were collected from Ross 308 broilers aged 35–45 days originating from five commercial farms (one per county) located in Caraș-Severin, Dolj, Gorj, Hunedoara, and Vâlcea. Samples were obtained from 137 production halls/series. Sera were tested using a commercial indirect ELISA kit and classified according to the manufacturer’s criteria based on the sample-to-positive (S/P) ratio and the corresponding antibody titer threshold. Overall, 2115/2466 sera were positive (85.77%) and 351/2466 were negative (14.23%). Anti-IBV antibodies were detected in all halls (137/137, 100%), although negative sera were recorded in 87/137 halls (63%). Seroprevalence by county ranged from 78.67% (Vâlcea) to 89.24% (Hunedoara). Significant differences in the proportions of positive and negative sera were identified between several county pairs by Fisher’s exact test. These findings indicate widespread serological evidence of anti-IBV antibodies in vaccinated broiler flocks from south-western Romania, which may reflect vaccine-induced immunity and/or field exposure. The results support the need for continued serological monitoring, alongside targeted molecular investigations to differentiate vaccine strains from circulating field variants. Full article

The single von Willebrand factor C (SVWC) domain-containing protein family represents a crucial class of immune molecules recently identified in insects and crustaceans. Initially regarded as functional analogs of vertebrate interferons (IFNs) due to their virus-induced expression and activation of the Janus kinase-signal transducer and activator of the transcription (JAK-STAT) pathway, recent studies have revealed that SVWC proteins possess far more complex functions. Many SVWC members are themselves a novel class of pattern recognition receptors (PRRs) that can directly bind to viruses and bacteria. Importantly, SVWCs are not a single entity but a highly diverse family—multiple subtypes exist in Drosophila, Bombyx mori, and shrimp—a gene expansion that implies functional differentiation. This review systematically examines the multifunctionality of SVWC proteins in insects and crustaceans, with a particular focus on the functional specialization driven by subtype diversity. We delve into the complex regulatory networks governing SVWC expression, including the differential activation by nuclear factor kappa B (NF-κB) pathways (Dorsal, Rel-2, Relish) and interferon regulatory factor (IRF) pathways. We detail the unique signaling mechanism by which SVWCs activate the JAK-STAT pathway via integrins, rather than the canonical Domeless receptor. Furthermore, we extend the discussion to the emerging roles of SVWCs as PRRs in humoral immunity (activating Toll/IMD pathways to induce antimicrobial peptides) and cellular immunity (mediating hemocyte phagocytosis). Based on current evidence, We propose that diverse SVWC subtypes may recognize distinct pathogens, bind to different integrin receptors, and activate specific STAT variants via disparate upstream induction pathways, thereby establishing a systematic and hierarchical immunoregulatory network. This understanding positions the SVWC protein family as a central hub in the insect immune network and offers a novel perspective on the complexity and evolution of invertebrate immunity. Full article

For over a decade, non-integrating Sendai virus vectors have been the gold standard for induced pluripotent stem cell (iPSC) reprogramming. However, as the field shifts toward regenerative and precision medicine and large-scale biorepositories, Sendai virus workflow necessitates dedicated viral-clearance testing, specialized manufacturing controls, and heightened regulatory oversight, leading to increased cost. While mRNA-based reprogramming offers a non-viral alternative, traditional mRNA delivery methods like electroporation are often physiologically disruptive. This study evaluates an mRNA-reprogramming platform that delivers lipid nanoparticles (mRNA-LNPs) via receptor-mediated endocytosis. By utilizing both Sendai virus and mRNA-LNP approaches to reprogram PBMCs from the same donor, we established a genetically identical starting point. Results demonstrate that mRNA-LNP-reprogrammed iPSCs maintain genomic integrity, retain the donor KCNH2 c.2398+5G>T variant, and exhibit characteristic colony morphology, pluripotency markers, and trilineage differentiation capacity consistent with the Sendai-reprogrammed counterparts. The mRNA-LNP-reprogrammed iPSCs differentiate into iPSC-derived cardiomyocytes presenting sarcomeric structures and electrophysiological activity, recapitulating a disease-specific phenotype. Notably, the mRNA-LNP workflow reached these milestones in significantly fewer passages than the Sendai virus workflow, markedly shortening timelines and reducing costs. These findings highlight mRNA-LNP reprogramming as a potentially attractive and effective, virus-independent platform to support future regenerative and precision medicine initiatives and scalable biobanking. Full article

The 5′-proximal region of the citrus tristeza virus (CTV) RNA genome is a hub where several elements involved in different facets of the virus cycle reside, including the sequences driving the production of the viral long non-coding RNA (lncRNA) LMT1. The sequence of this region is one of the most divergent genome areas, allowing for strain differentiation. Beyond its use in assessing viral population diversity, the region provides a valuable model for studying the conservation of RNA structure and function despite sequence variation. Here, we integrated comparative in silico analysis of the LMT1 region from variants of eight CTV strains with selective 2′-hydroxyl acylation, analyzed by primer extension and mutational profiling (SHAPE-MaP) probing of in vitro–generated LMT1 RNAs from two divergent strains, T36 and T68. The predicted consensus structures revealed 19 putative, conserved stem-loops. The SHAPE-MaP reactivity data supported and substantiated the thermodynamics-based predictions for the 15 previously uncharacterized stem-loops and two functional elements identified earlier. The strong structural conservation across strains highlights that the LMT1 RNA structure contributes to its function during CTV infection. These results provide the first experimentally supported structure of this viral lncRNA and lay the foundation for defining how individual RNA motifs influence CTV biology. Full article

Background: Despite the effectiveness of current SARS-CoV-2 vaccines, the genetic variability of the viral target has led to the emergence of variants capable of evading vaccine-induced protection. To ensure broader and more durable protection, we investigated the efficacy of a novel vaccine strategy. Methods: This vaccine utilizes the highly conserved nucleocapsid (N) protein as its primary antigen, rather than the spike (S) protein. It incorporates the Papaya Mosaic Virus (PapMV) nanoparticle, a Toll-like receptor (TLR) 7/8 agonist with intrinsic adjuvant properties, as a vaccine platform. Results: The vaccine formulations, comprising PapMV nanoparticles and the N antigen covalently or non-covalently attached to the PpaMV nano, generated robust humoral (antibody) and cellular (T-cell) immune responses. Protective efficacy was evaluated in K18-hACE2 transgenic mice challenged with either the ancestral SARS-CoV-2 strain or the Omicron XBB.1.5 variant. In both cases, the vaccine significantly reduced inflammation and viral titers in the lungs of vaccinated animals. Conclusions: These results highlight the potential of this PapMV-N vaccine to induce broad protection against diverse SARS-CoV-2 variants. 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

SARS-CoV-2 is a zoonotic virus with a proven ability to infect various animal species, including domestic cats. In the post-pandemic period of COVID 19, limited data still exist on the clinical course, shedding of infectious virus and diagnostic features in cats. The aim of this study was to investigate the spread of SARS-CoV-2 in cats in 2023, the clinical manifestations of the infection, the diagnostic algorithm, including molecular detection of viral components, the differential diagnosis of co-infection with FHV, FCV, Mycoplasma spp. and Chlamydia felis, serology, and the isolation of infectious SARS-CoV-2. The immunomodulatory therapy in animals with a standalone SARS-CoV-2 infection was applied. The study included oropharyngeal, conjunctival and nasal swab samples from 102 domestic cats with clinical signs. Of them, 20.6% (21/102) were positive for SARS-CoV-2, with 16.67% (17/102) of the cats showing various variants of co-infection with FHV, FCV, Mycoplasma spp. and Chlamydia felis. Four of the cats had a standalone SARS-CoV-2 with mild clinical manifestations that included serous discharges from the eyes, without change in the general condition. The virus was isolated from these samples. These four cats and their owners were positive for antibodies to the virus, and the owners were PCR-negative. The treatment of SARS-CoV-2 infection included the preparations Viusid, RX immunosuport, Vetomun and Lisymun. This is one of the first post-pandemic studies covering FHV, FCV, Mycoplasma spp. and Chlamydia felis in domestic cats with SARS-CoV-2 infection and further expands on the essential main idea including the specified pathogens of interest. Full article

Fibropapillomatosis, a disease associated with Scutavirus chelonidalpha5, commonly known as Chelonid alphaherpesvirus 5 (ChHV5), manifests as benign tumors that impair the motor, visual, and physiological functions of affected sea turtles. In this study, blood and tissue samples were collected from turtles exhibiting fibropapilloma-like lesions as well as from clinically healthy individuals. A nested PCR approach was employed to amplify the viral UL30 and UL28 genes for the detection and characterization of the virus variants. The mitochondrial control region was used to assess the relationship between the turtle population and the viral variant. Among the 19 turtles analyzed, six tested positive for ChHV5, including both symptomatic and asymptomatic turtles. Phylogenetic analysis revealed that three positive samples belonged to the Western Atlantic/Caribbean clade, whereas the other three grouped within the Atlantic clade. New oligonucleotides and probes were designed for ChHV5 qPCR detection, accounting for the globally accumulated genetic variability. The qPCR test parameters demonstrated an optimized assay with an efficiency of 101.4% and a detection limit of 2.4 genome copy equivalents (GCE)/μL. This study confirms the presence of two ChHV5 viral variants in rescued turtles from the Caribbean region of Colombia, including both clinically affected and asymptomatic individuals. Therefore, these results support the association between ChHV5 and fibropapillomatosis. Furthermore, analysis of the mitochondrial control region supports the hypothesis of horizontal transmission of the virus. A novel qPCR protocol with a synthetic control is proposed to improve early diagnosis and strengthen conservation and prevention strategies. Full article