Search Results (2,025)

Background: Respiratory syncytial virus (RSV) RNA-dependent RNA polymerase (RdRp) complex is an essential molecular machine for viral genome replication. The L protein, the catalytic subunit of this complex (L-RdRp), is well-characterized structurally and represents a highly promising target for the development of novel small-molecule drugs against RSV. Methods: To address the limitations of current QSAR-based virtual screening strategies for RSV L-RdRp inhibitor development, we established a multi-dimensional computer-aided drug screening framework integrating activity, toxicity, drug-likeness, and stability. Results: Two OECD-compliant 2D-QSAR models were developed and rigorously validated to predict inhibitory activity and cytotoxicity, respectively. The optimal inhibitory activity model exhibited strong statistical performance, with R² = 0.8281, $Q_{\mathrm{L}\mathrm{O}\mathrm{O}}^2=$ 0.7653, $R_{\mathrm{t}\mathrm{e}\mathrm{s}\mathrm{t}}^2=$ 0.8713, $Q_{\mathrm{F}\mathrm{n}}^2=$ 0.8594 ∼ 0.8837, CCC_test = 0.9301, MAE_test = 0.1966. Similarly, the best cytotoxicity model achieved R²= 0.8263, $Q_{\mathrm{L}\mathrm{O}\mathrm{O}}^2$ = 0.7422, $R_{\mathrm{t}\mathrm{e}\mathrm{s}\mathrm{t}}^2$ = 0.8951, $Q_{\mathrm{F}\mathrm{n}}^2$ = 0.8108~0.8530, CCC_test = 0.9081, MAE_test = 0.1685. Based on these models, a four-step screening workflow—QSAR-based filtering and molecular docking (15,758 → 2446 → 162 → 19 compounds), ADMET evaluation (19 → 5), and molecular dynamics simulations (MDSs)—was implemented to identify promising L-RdRp inhibitors. Conclusions: Ultimately, five candidate compounds were selected, all of which demonstrated predicted higher inhibitory activity, lower predicted cytotoxicity, a stable predicted binding mode, and favorable oral bioavailability compared with the reference drug remdesivir. These findings provide valuable in silico-derived lead candidates and a reliable computational workflow for identifying experimental L-RdRp inhibitors targeting RSV. Full article

Goose astrovirus (GoAstV) is a newly identified virus affecting geese, leading to goose gout, which is marked by urate deposits in organs and joints. This research investigates the role of autophagy in renal damage caused by GoAstV in goslings. The findings indicated that GoAstV infection in goslings resulted in characteristic clinical manifestations, with renal tissues displaying tubular swelling, inflammatory infiltration, and autophagosome formation. In vivo experiments demonstrated a significant upregulation of mRNA levels for autophagy-related factors, including AMPK, LC3A, ATG5, ATG7, P62, Beclin1, AMBRA1 and GABARAPL1, while mTOR and LC3B levels were notably decreased. At 3 dpi, the protein expression levels of ATG5, Beclin1, and LC3B II/I increased, while P62 levels decreased, suggesting autophagy activation. In vitro analyses revealed that GoAstV infection led to enhanced autophagy; however, the concurrent upregulation of LC3B II/I and P62 proteins suggested an obstruction in the autophagic flux. Upon the inhibition of autophagy with 3-methyladenine (3-MA, autophagy inhibitor), there was a significant reduction in the expression of autophagy-related factors, accompanied by a marked decrease in viral replication rates. In conclusion, GoAstV infection in gosling kidney cells initially triggers enhanced autophagy and subsequently causes a blockage in the autophagic flux. The virus exploits autophagosomes for replication, ultimately resulting in kidney damage. The application of 3-MA effectively inhibits this autophagic process and diminishes viral replication. Full article

Purpose: Nucleotide metabolism plays a critical role in cancer development, but the prognostic significance of genetic variants in nucleotide metabolism genes for hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) patients remains unclear. Methods: We performed Cox proportional hazards regression analyses to evaluate the association between genetic variants in 94 nucleotide metabolism-related genes and overall survival (OS) in 866 HBV-HCC patients. To assess the potential biological relevance of the identified variants, the Bayesian false discovery probability and false-positive report probability were applied for multiple testing correction. Results: Two independent SNPs, DCTD rs17074255 G>A (HR = 1.22, 95% CI: 1.06–1.40, p = 0.005) and ENTPD2 rs3763662 G>A (HR = 1.18, 95% CI: 1.03–1.34, p = 0.015), were significantly associated with OS. A significant dose-dependent association between the number of risk genotypes and poorer OS was observed (P_trend < 0.001). Luciferase reporter assays demonstrated allele-specific regulatory effects of rs3763662 on ENTPD2 expression (p < 0.001). DCTD and ENTPD2 mRNA expression levels were significantly elevated in HCC tumors in the UALCAN database and in our 103 paired samples. Higher expression levels of both genes were associated with poorer survival in the TCGA cohort (p = 0.003 and p < 0.001). Conclusions: Our findings suggest that ENTPD2 rs3763662 (supported by direct functional evidence) and DCTD rs17074255 (supported by eQTL and expression associations) may serve as potential prognostic indicators for HBV-HCC through the regulation of mRNA expression. These findings provide new insights into the role of nucleotide metabolism-related genetic variation in HBV-HCC progression and may facilitate prognostic assessment, pending replication in independent cohorts. Full article

Respiratory syncytial virus (RSV) and human metapneumovirus (hMPV), members of the family Pneumoviridae, represent a foremost global cause of acute lower respiratory tract infection in infants, young children, the elderly, and immunocompromised individuals. Despite the recent approval of preventive vaccines and monoclonal antibody prophylactics targeting the viral fusion protein, no widely adopted, RSV-specific direct-acting antiviral is currently approved for routine post-infection treatment. The large (L) protein of the viral RNA polymerase complex, which catalyzes genome replication and mRNA transcription in concert with its obligate cofactor, the phosphoprotein (P), constitutes an ideal drug target owing to its essential and multifunctional enzymatic activities and its absence from host cells. Over the past decade, Cryo-electron microscopy (Cryo-EM) has yielded a series of landmark structures of Pneumoviridae L–P complexes, including apo forms of RSV (at 3.2–3.67 Å) and hMPV (at 3.7 Å) polymerases, among the first promoter-bound non-segmented negative-sense (nsNSV) RNA virus polymerase structures (at 3.40–3.41 Å), and inhibitor-bound complexes that illuminate the molecular basis of non-nucleoside inhibitor (NNI) action at sub-nanomolar potency. This review synthesizes the structural biology of Pneumoviridae RNA polymerases from a chronological and mechanistic perspective, compares RSV and hMPV L protein active sites at near-atomic resolution, and critically evaluates how structural insights are being translated into next-generation antiviral drug candidates, including nucleoside analog inhibitors, allosteric non-nucleoside inhibitors, and emerging candidates at various stages of preclinical and clinical investigation. Full article

Bats are important reservoirs of zoonotic pathogens and valuable models for studying antiviral tolerance and neuroinflammation within a One Health framework. However, chiropteran neural 2D-in vitro models remain limited. Here, we established and characterized the first chiropteran primary (CpBCs) and immortalized (CiBCs) cell lines from Hypsugo savii species. To overcome the limited lifespan of CpBCs, immortalization strategies based on human telomerase reverse transcriptase (hTERT) and Simian virus 40 large T antigen (SV40) were evaluated. Electroporation-mediated transfection with SV40 successfully generated CiBCs, whereas liposome-mediated and hTERT-based approaches were unsuccessful. RNA sequencing revealed marked transcriptional changes comparing CiBCs with CpCBs, such as the upregulation of pathways related to cell cycle progression, DNA replication, and proliferation in CiBCs, together with the downregulation of apoptosis, inflammatory signaling, and immune-related pathways. Immortalized cells also exhibited enrichment of neural stem cell-like and cancer-associated signatures, suggesting partial dedifferentiation induced by SV40-mediated immortalization. Overall, this study provides a novel chiropteran brain-derived 2D-in vitro platform for investigating bat neurobiology, host–pathogen interactions, viral tolerance, and neurotropic infectious diseases relevant to emerging zoonoses. Full article

Background: Oncolytic viruses (OVs), a promising anti-cancer therapeutic, replicate more efficiently in cancer cells rather than in healthy cells due to the alterations in antiviral response mechanisms and dysregulation of signaling pathways. Vesicular stomatitis virus (VSV) is known for low pathogenicity, tropism to various cancer cells, and the ability to lyse cells in the hypoxic tumor microenvironment (TME). Targeted delivery of immune checkpoint and co-stimulatory molecules can enhance the anti-tumor immune response and remodel the immunosuppressive TME. The aim of this study was to compare the activity of rVSV-GFP with rVSV, encoding the programmed cell death protein 1 (PD-1) and tumor necrosis factor ligand superfamily member 9 (4-1BBL). Methods: The oncolytic efficacy of these rVSV variants used at 10⁵, 10⁶, and 10⁷ TCID50 was evaluated at 24 and 48 h post-infection by flow cytometry in a panel of mouse and human cancer cell lines. Quantitative real-time polymerase chain reaction (qPCR) was used to evaluate mRNA expression levels of certain genes at 12 and 48 h post-infection. Results: Murine hepatocellular carcinoma (H22) and human melanoma (A375) or human lung carcinoma (A549) were the most sensitive to rVSV therapy cell lines. The higher relative expression of the antiviral response genes RIG-I and IFIT1 within each biological species (mouse or human) correlated with lower sensitivity to rVSV. No such effect was observed for the type I interferons (IFNs), despite their proposed key role in resistance to OV therapy. Conclusions: H22, A375, and A549 are more susceptible to the oncolytic activity of the novel rVSV-PD-1-4-1BBL. Full article

Host microRNAs (miRNAs) are widely proposed as innate antiviral effectors against SARS-CoV-2, yet whether they actually restrict infection in lung epithelial cells remains unresolved. Two of the most-cited candidates, miR-29a-3p and miR-15b-5p, are predicted to bind both the viral genome and key entry/trafficking factors such as Furin and ATG9A, but functional evidence is fragmented and often contradictory. Here, we put both miRNAs to the test in human Calu-3 cells infected with the SARS-CoV-2 Beta and Omicron BA.1 variants, using parallel gain- and loss-of-function strategies coupled to RT-qPCR of viral and cellular transcripts and back-titration of infectious progeny on VeroE6/TMPRSS2 cells. Both miRNAs transiently suppressed viral gene expression at 6 hpi, but this early dampening was followed by a marked transcript rebound at 24 hpi, especially for Omicron, with virtually no impact on total extracellular viral RNA. More strikingly, miR-15b modulation enhanced infectious virus output during Beta infection, and miR-29a overexpression boosted Omicron BA.1 infectivity, while Furin, ATG9A, AKT3, and TFEB showed only modest, condition-dependent shifts. Rather than acting as clean antiviral effectors, miR-29a and miR-15b emerge as context-dependent modulators that can paradoxically favor SARS-CoV-2 replication—a cautionary signal for miRNA-based antiviral strategies. Full article

The recurrent emergence of viral outbreaks, including SARS-CoV-2, influenza, Ebola, and respiratory syncytial virus (RSV), continues to expose critical limitations in conventional antiviral therapies, particularly in terms of targeting specificity, bioavailability, and resistance development. Nanotechnology has emerged as a transformative approach to overcome these challenges. This review provides a comprehensive and critical analysis of nanoparticle-based antiviral systems, including lipid-based, polymeric, inorganic, and hybrid nanocarriers, with a focus on their roles in enhancing drug delivery, targeting precision, and therapeutic efficacy. These platforms exert antiviral effects through multiple coordinated mechanisms, including inhibition of viral entry, suppression of replication, gene silencing, and modulation of host immune responses. The clinical success of lipid nanoparticle-based mRNA vaccines highlights the translational potential of nanotechnology, while emerging nanotherapeutic strategies demonstrate increasing versatility across diverse viral pathogens. However, key challenges—including safety, scalability, formulation stability, and regulatory constraints—continue to limit widespread clinical implementation. Overall, nanoparticle-mediated antiviral systems represent a multifunctional and adaptable platform capable of addressing the limitations of conventional therapies and enabling more effective, resilient, and precision-driven strategies for future pandemic preparedness. Full article

The molecular mechanisms governing the efficient packaging of the large SARS-CoV-2 RNA genome into progeny virions remain incompletely understood, with the role of untranslated regions (UTRs) being particularly enigmatic. Leveraging proximity ligation sequencing data, we identified direct, high-frequency interactions between the viral packaging signal PS9 and both the 5′ and 3′ UTRs during intracellular replication stages. Functional validation using an infectious virus-like particle (iVLP) system demonstrated that genomes incorporating SARS-CoV-2 UTRs exhibited significantly enhanced packaging efficiency, yielding an increase in both packaged RNA copies and reporter gene expression post-infection. Competitive packaging assays confirmed the UTRs confer a selective advantage during particle assembly. Mechanistically, Western blot and digital Western analysis revealed that UTR-containing iVLPs incorporated approximately 2-fold more nucleocapsid (N) proteins, suggesting enhanced N recruitment or retention. The deletion of specific core sequences within the UTRs predicted to form a base pair with PS9 abrogated this enhancement, suggesting the functional significance of the UTR-PS9 interaction interface. Collectively, these results establish that the 5′ and 3′ UTRs act synergistically through direct RNA-RNA interactions with PS9 to promote N protein recruitment and enhance packaging efficiency in a PS9-dependent iVLPs system. This UTR-PS9 regulatory axis presents a novel target for therapeutic intervention against SARS-CoV-2 and related coronaviruses. Full article

Andes virus (ANDV) is the only hantavirus with well-documented evidence of person-to-person transmission. However, key parameters related to transmission timing, viral shedding, exposure contexts, and public health management remain incompletely defined. We conducted a systematic review in accordance with PRISMA 2020. MEDLINE/PubMed, Scopus, and Web of Science were searched from database inception up to 14 May 2026. Eligible studies reported epidemiological, virological, clinical, or public health data relevant to ANDV infection, person-to-person transmission, viral shedding, and/or outbreak control. Thirty-three studies, including 17,204 individuals, 2221 laboratory-confirmed ANDV cases, and 135 documented secondary cases, were included. Person-to-person transmission was identified as a primary or co-occurring route in 20 papers. The median incubation period among ANDV cases was 20.8 days, and the median serial interval was 21.8 days (upper bounds near 40 days). Secondary attack rates were higher among sexual and other close contacts. ANDV RNA was consistently detected in blood and occasionally in saliva, respiratory secretions, urine, breast milk, and semen, although RNA detection alone does not necessarily imply infectious virus. Rare reports of culture-confirmed isolation of replication-competent virus support the biological plausibility of transmission via close mucosal or respiratory exposure. Unlike other hantaviruses, Andes virus can spread person to person through close contact, supporting prolonged monitoring and risk-stratified follow-up of high-risk contacts based on ANDV-specific epidemiological evidence. Possible recommendations, including post-discharge counselling regarding possible sexual transmission, remain provisional and require further evidence. Preparedness activities against outbreaks should also be implemented in non-endemic regions, while future research should prioritize prospective contact studies, standardized virological sampling, and genomic confirmation. Full article

The COVID-19 pandemic highlighted the urgent need to develop effective and broad-spectrum antiviral therapies against coronaviruses. One strategy to address this concern is a combination therapy using repurposed drugs against zoonotic viruses with pandemic potential. We previously demonstrated that the combination of Remdesivir and Ivermectin is highly potent and synergistic in inhibiting the replication of murine hepatitis virus (MHV) in RAW264.7 macrophages. This study investigated the interactions between the drug combination, coronavirus and host by proteomics and RNA sequencing of MHV-infected H2.35 murine liver epithelial cells. Time-of-addition and time-of-removal assays suggested that the drug combination likely affected the synthesis of viral RNA and viral protein. This combination drastically diminished the live virus titer greater than the respective monotherapies in MHV-infected H2.35 cells (by ~4 log₁₀), as well as in SARS-CoV-2-infected VeroE6 cells and human nasal epithelial cells. Proteomic and transcriptomic analyses revealed that viral protein and RNA levels were significantly depressed upon combination treatment. The drug combination exhibited considerable negative effects upon host RNA processes and resulted in the upregulation of host protein processes (e.g., response to unfolded protein; protein insertion into ER membrane). Molecular pathways affected by the combination treatment were markedly distinct from the monotherapies and indicated that Ivermectin enhances Remdesivir by modulating critical host processes to synergistically exert its inhibitory effect on the coronavirus replication cycle. Full article

Dengue virus (DENV) and chikungunya virus (CHIKV) co-circulate in many regions and present with overlapping clinical features, which complicate accurate diagnosis and disease management. This study develops an integrative transcriptomic framework to identify robust host gene signatures that distinguish between dengue, chikungunya, and healthy states. Publicly available RNA sequencing (RNA-seq) datasets derived from human blood samples were analyzed using a cross-validation design to ensure robustness and prevent information leakage. Differential expression analysis was performed independently within each dataset using the Generalized Linear Models with Quasi-Likelihood F-tests and Magnitude–Altitude Scoring (GLMQL-MAS) framework, followed by Cross-Magnitude–Altitude Scoring (Cross-MAS) integration to identify shared and virus-specific gene signatures. A strict consensus approach across folds was applied to derive reproducible gene sets. These signatures were used for dimensionality reduction and multinomial logistic regression to evaluate classification performance. A small subset of selected genes showed strong discriminative performance within the cross-validation framework, with test balanced accuracy reaching 0.97, which improved upon models using all genes. Biologically, both infections exhibited a shared antiviral response characterized by interferon signaling and innate immune activation. However, distinct virus-specific patterns were identified. Dengue infection was associated with cell-cycle and DNA replication pathways, while chikungunya infection showed stronger enrichment of inflammatory and immune signaling pathways, including NF-kappaB and Toll-like receptor signaling. Overall, this study provides a cross-validation-based framework for integrative transcriptomic analysis and identifies compact, reproducible host-response signatures with strong discriminative signals in the analyzed cohorts. These signatures require validation in larger independent cohorts before any clinical or diagnostic application. Full article

Background/Objectives: Self-amplifying RNA (saRNA) vectors enable high-level transgene expression from minimal initial doses. While alphavirus-based saRNA systems are widely used, they suffer from limitations, including large genome size, complex replicase machinery, and cellular toxicity. Nodamura virus (NoV) offers a promising alternative due to its compact genome (3.2 kb) and low cytotoxicity. This study aimed to elucidate NoV RNA1 replication mechanisms and develop a novel NoV-based saRNA vector platform. Methods: We established a Schizosaccharomyces pombe system to investigate NoV RNA1 replication and protein A localization. N-terminal deletion mutants and ER-targeting chimeras were constructed to characterize membrane targeting determinants. Based on mechanistic insights, we developed NovaVec by inserting transgenes at the RNA3⁴²² site within the subgenomic RNA3 region. In vivo performance was evaluated using lipid nanoparticle-encapsulated NovaVec expressing nanoluciferase or monkeypox A33R antigen in BALB/c mice. Results: We identified redundant mitochondrial targeting domains (amino acids 2-15 and 16-33) in NoV protein A, where either domain was sufficient for proper localization and replication. The replication machinery could be functionally redirected to the endoplasmic reticulum while maintaining replication competence. Lipid nanoparticle-encapsulated NovaVec achieved sustained transgene expression for 54 days in mice, significantly outperforming conventional mRNA vectors that lost signal within 14 days. The NovaVec-based monkeypox A33R vaccine elicited robust antigen-specific humoral immunity with titers reaching approximately 1:12,800 following booster immunization. Conclusions: With its compact genome encoding only a single replicase protein, minimal cytopathic effects, and demonstrated capacity for long-term protein expression, NovaVec represents a highly promising next-generation saRNA platform for vaccines. Full article

The Chinese bahaba (Bahaba taipingensis), an endangered marine fish, is highly vulnerable to infectious spleen and kidney necrosis virus (ISKNV). In this work, we developed a gill filament-derived cell line, designated BTG, to investigate how these cells respond to ISKNV over time, specifically from 3 to 24 h post-infection (hpi). BTG cells grew steadily, displayed a diploid chromosome number of 2n = 48, demonstrated high transfection efficiency, and were highly susceptible to viral infection. Characteristic cytopathic effects (CPEs) became noticeable as early as 6 hpi at 27 °C. RNA-seq profiling showed that the number of differentially expressed genes (DEGs) steadily increased with time. Standard enrichment analysis at individual time points (3, 6, 12, and 24 hpi) highlighted pathways mainly involved in DNA replication, cell cycle control, ribosome assembly, transcription and translation, mismatch repair, and cell adhesion. Temporal clustering analysis, however, revealed hidden patterns in immune gene expression. Genes that were consistently downregulated were enriched in immune-related pathways, including ECM–receptor interaction, cytokine–receptor signaling, PI3K–AKT, and Wnt signaling, indicating prolonged suppression of host defense mechanisms. In contrast, clusters of genes transiently upregulated during the first 6 h post-infection were associated with antiviral and innate immune pathways, such as NF-κB, JNK, IRF3, IRF7, caspases, JAK, MHC I, and lysosome-related functions, suggesting a rapid but short-lived antiviral response. Genes that were continuously upregulated were primarily involved in nucleic acid replication and protein synthesis, reflecting a gradual host cell reprogramming to support viral replication. Taken together, these findings reveal a temporal shift in BTG cells from an initial burst of immune activity to immune suppression, accompanied by enhanced viral replication. The BTG cell line thus represents a valuable in vitro model for dissecting ISKNV–host interactions and offers new perspectives on the molecular strategies employed by megalocytiviruses in B. taipingensis. Full article

DNA viruses rely extensively on host cellular machinery, including replication factors and transcriptional systems, to persist after infection. These mechanisms make studying and targeting DNA viral proteins challenging, as they also play key roles in mammalian processes. Traditional strategies include CRISPR-mediated gene disruption and small interfering RNA (siRNA) to target host proteins. However, Proteolysis Targeting Chimeras (PROTACs) offer a novel strategy by enabling the selective and rapid degradation of specific viral or host proteins involved in the DNA viral lifecycle. PROTACs are heterobifunctional molecules composed of three key components: a ligand that binds the target protein, a chemical linker, and a ligand that recruits an E3 ubiquitin ligase. By simultaneously binding both the target protein and the E3 ligase, PROTACs form a ternary complex. This proximity enables the E3 ligase to ubiquitinate the target protein, marking it for recognition and subsequent degradation by the intracellular proteasome. This approach represents a promising avenue for targeting previously undruggable proteins and improving therapeutic outcomes in virus-associated malignancies. In this perspective, we describe studies that use PROTACs as tools to modulate host proteins to investigate DNA viral processes with temporal control of host protein expression, as well as the use of PROTACs as antivirals to directly target DNA viral proteins. We also provide a detailed chart summarizing known host-targeting PROTACs and their potential applications across different stages of DNA viral lifecycles, highlighting opportunities for future DNA virus research. Full article