Search Results (212)

Porcine reproductive and respiratory syndrome virus (PRRSV), an enveloped single-stranded positive-sense RNA virus, poses a significant threat to global swine production. Despite the availability of modified live virus and inactivated vaccines, their limited efficacy and safety concerns highlight the urgent need for novel antiviral therapeutics. This study aimed to investigate the molecular mechanisms by which lycopene inhibits PRRSV replication. Initial assessments confirmed that lycopene did not adversely affect cellular viability, cell cycle progression, or apoptosis. Using fluorescence microscopy, flow cytometry, immunoblotting, quantitative real-time PCR (qRT-PCR), and viral titration assays, lycopene was shown to exhibit potent antiviral activity against PRRSV. Mechanistic studies revealed that lycopene suppresses reactive oxygen species (ROS) production, which is critical for PRRSV proliferation. Additionally, lycopene attenuated PRRSV-induced inflammatory responses, as demonstrated by immunoblotting, ELISA, and qRT-PCR assays. These findings suggest that lycopene inhibits PRRSV replication by modulating ROS levels and mitigating inflammation, offering a promising avenue for the development of antiviral therapeutics. This study provides new insights and strategies for combating PRRSV infections, emphasizing the potential of lycopene as a safe and effective antiviral agent. Full article

A novel barna-like virus was found to be associated with field-collected Afrina sporoboliae plant-parasitic nematodes. The positive-sense, single-stranded RNA genome of this virus, named Afrina barna-like virus (AfBLV), comprises 4020 nucleotides encoding four open reading frames (ORFs). ORF 1 encodes a protein product spanning a transmembrane, a peptidase, and VPg domains, whereas an overlapping ORF 2 encodes an RNA-dependent RNA polymerase (RdRP). ORF2 may be expressed via a −1 translational frameshift. In phylogenetic reconstructions, the RdRP of AfBLV was placed inside a separate clade of barna and barna-like viruses related to but distinct from the genera in the Solemoviridae and Alvernaviridae families, within the overall lineage of Sobelivirales. ORF 3 of AfBLV encodes a protein product of 206 amino acids (aa) long with homology to a putative protein encoded by a similarly positioned gene of an uncharacterized virus sequence identified previously as Barnaviridae sp. ORF 4 encodes a 161 aa protein with no significant similarities to sequences in the GenBank databases. AfBLV is the first barnavirus found in a nematode. Sequence comparisons of the AfBLV genome and genomes of other barna-like viruses suggested that a recombination event was involved in the evolution of AfBLV. Analyses of the phylogeny of RdRPs and genome organizations of barna-like and solemo-like viruses support the re-classification of Barnavirus and Dinornavirus genera as members of the Solemoviridae family. Full article

Hepatitis E virus (HEV) is a positive-sense, single-stranded RNA virus that poses a significant public health risk, yet its study is hindered by the complexity of conventional RNA-based reverse genetics systems. These systems require multiple steps, including genome cloning, in vitro transcription, and capping, making them labor-intensive and susceptible to RNA degradation. In this study, we developed a single-step, plasmid-based HEV expression system that enabled direct intracellular transcription of the full-length HEV genome under a cytomegalovirus immediate-early (CMV-IE) promoter. The viral genome was flanked by hammerhead (HH) and hepatitis delta virus (HDV) ribozymes to ensure precise self-cleavage and the generation of authentic 5′ and 3′ termini. This system successfully supported HEV genome replication, viral protein expression, and progeny virion production at levels comparable to those obtained using in vitro-transcribed, capped HEV RNA. Additionally, a genetic marker introduced into the plasmid construct was stably retained in progeny virions, demonstrating the feasibility of targeted genetic modifications. However, plasmid-derived HEV exhibited delayed replication kinetics, likely due to the absence of an immediate 5′ cap. Attempts to enhance capping efficiency through co-expression of the vaccinia virus capping enzyme failed to improve HEV replication, suggesting that alternative strategies, such as optimizing the promoter design for capping, may be required. This plasmid-based HEV reverse genetics system simplifies the study of HEV replication and pathogenesis and provides a versatile platform for the genetic engineering of the HEV genome. Full article

Infectious bronchitis virus (IBV) is a chicken pathogen present in commercial poultry farms worldwide. It is classified within the species Avian coronavirus, genus Gammacoronavirus. As with other members of the family Coronaviridae, it has a single positive-sense RNA genome with 27.6 Kb and presents viral particles with a typical crown-like aspect due to the spike (S) transmembrane glycoprotein. IBV has a remarkable capacity for genetic recombination and mutation, resulting in many genotypes and antigenic variants over evolutionary time. Currently, it is classified into nine genetic types (GI to GIX) and 41 (1 to 41) lineages disseminated worldwide. In South America, IBV was first identified in early commercial poultry production ventures in Brazil in the 1950s. Since then, this virus has been frequently detected in commercial South American poultry farms, being classified into serotypes in the first decades and genotypes more recently. IBVs of the Massachusetts (Mass) serotype were initially detected and vaccine strains of this serotype were used extensively on commercial poultry farms. Other serotypes/genotypes were identified later, with almost all of them classified in the current genetic type I (GI). In addition, five GI lineages (GI-1, -11, -13, -16, and -23) have been associated with the main infectious bronchitis outbreaks in the continent, with some variations in the occurrence according to the countries and the period of time. Molecular epidemiological surveillance of IBV genetic types and lineages is necessary to anticipate potential outbreaks, revealing patterns of viral evolution and dissemination, as well as to guide the selection of appropriate vaccine strains and immunization programs. Full article

The family Alphaflexiviridae comprises plant- and fungus-infecting viruses with single-stranded, positive-sense RNA genomes ranging from 5.4 to 9 kb. Their virions are flexuous and filamentous, measuring 470–800 nm in length and 12–13 nm in diameter. The family includes 72 recognized species, classified into six genera: Allexivirus, Lolavirus, Platypuvirus, Potexvirus (plant-infecting), and Botrexvirus and Sclerodarnavirus (fungus-infecting). The genus Potexvirus is the largest, with 52 species, including Potexvirus ecspotati (potato virus X), an important crop pathogen and plant virology model. The genera are distinguished by genome organization and host range, while species differentiation relies on nucleotide and protein sequence identity thresholds. In this review, we summarize the current knowledge on the genomic structure, conserved genes, and phylogenetic relationships within Alphaflexiviridae, with a particular focus on the replicase and coat protein genes as signature markers. Additionally, we update the model of cellular remodeling driven by the triple gene block proteins, which are essential for virus movement, among other viral functions. Beyond their biological significance, alphaflexiviruses serve as valuable models for studying virus–host dynamics and hold potential applications in plant disease control and biotechnology. This review provides an updated framework for understanding Alphaflexiviridae and their broader impact on plant virology. Full article

Infectious bronchitis virus (IBV) is an enveloped, positive-sense, single-stranded RNA virus belonging to the genus Gammacoronavirus. It primarily infects avian species, causing respiratory and renal disease and irreversible damage to the oviduct, which can lead to high mortality rates in chickens. The lack of rapid and reliable diagnostic tools for on-farm IBV detection hampers timely disease management and control measures. The introduction of DNA biosensors offers a promising approach for the sensitive and specific detection of IBV, facilitating rapid identification and intervention. In this study, an electrochemical DNA biosensor with a multi-walled carbon nanotube (MWCNT)-modified gold electrode was developed for IBV detection. The biosensor targeted the target-specific 5′ untranslated region (5′-UTR) of the IBV genome. Under optimal conditions, the immobilization and hybridization efficiencies were evaluated by cyclic voltammetry (CV) and differential pulse voltammetry (DPV), with methylene blue as a redox indicator. The developed DNA biosensor demonstrated a dynamic detection range from 2.0 × 10⁻¹² to 2.0 × 10⁻⁵ mol L⁻¹, with a limit of detection (LOD) of 2.6 nM and a limit of quantification (LOQ) of 0.79 nM. Validation using a small subset of clinical samples, including crude complementary DNA, and polymerase chain reaction products, showed high recovery rates ranging from 95.41% to 99.55%. While these findings highlight the potential of the proposed DNA biosensor as an innovative diagnostic tool for IBV detection, this study remains a proof of concept. However, further validation using a large number of clinical samples is essential to assess its feasibility, robustness, and practical application in a real-world farm setting Full article

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a positive-sense RNA virus with an unusually large genome of approximately 30 kb. It is highly transmissible and exhibits broad tissue tropism. The third most pathogenic of all known coronaviruses, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is responsible for the clinical manifestation known as coronavirus disease 2019 (COVID-19), which has resulted in the loss of millions of lives on a global scale. This pandemic has prompted significant efforts to develop therapeutic strategies that target the virus and/or human proteins to control viral infection. These efforts include the testing of hundreds of potential drugs and thousands of patients in clinical trials. Although the global pandemic caused by the SARS-CoV-2 virus is approaching its end, the emergence of new variants and drug-resistant mutants highlights the need for additional oral antivirals. The appearance of variants and the declining effectiveness of booster shots are resulting in breakthrough infections, which continue to impose a significant burden on healthcare systems. Computer-aided drug design (CADD) has been widely utilized for predicting drug–target interactions and evaluating drug safety; it is regarded as an effective tool for identifying promising drug candidates to combat SARS-CoV-2. The CADD approach aids in the discovery of new drugs or the repurposing of United States Food and Drug Administration (FDA)-approved drugs, whose safety and side effects are already well established, thus making the process more viable. This review summarizes potential therapeutic agents that target SARS-CoV-2 or host proteins critical for viral pathogenesis, as identified using CADD approaches. Additionally, this study provides insights into the common in silico methods used in CADD and their current applications in the SARS-CoV-2 drug discovery process. Full article

Orthoflavivirus, a genus encompassing arthropod-borne, positive-sense, single-stranded RNA viruses in the Flaviviridae family, represents clinically relevant viruses that pose significant threats to human and animal health worldwide. With warming climates and persistent urbanization, arthropod vectors and the viruses they transmit continue to widen their geographic distribution, expanding endemic zones. Flaviviruses such as dengue virus, Zika virus, West Nile virus, and tick-borne encephalitis virus cause debilitating and fatal infections globally. In 2024, the World Health Organization and the Pan American Health Organization declared the current dengue situation a Multi-Country Grade 3 Outbreak, the highest level. FDA-approved treatment options for diseases caused by flaviviruses are limited or non-existent, and vaccines are suboptimal for many flaviviruses. Understanding the molecular characteristics of the flavivirus life cycle, virus-host interactions, and resulting pathogenesis in various cells and model systems is critical for developing effective therapeutic intervention strategies. This review will focus on the virus-host interactions of mosquito- and tick-borne flaviviruses from the virus replication and assembly perspective, emphasizing the interplay between viral non-structural proteins and host pathways that are hijacked for their advantage. Highlighting interaction pathways, including innate immunity, intracellular movement, and membrane modification, emphasizes the need for rigorous and targeted antiviral research and development against these re-emerging viruses. Full article

Enterovirus 68 (EV-D68) is a non-enveloped virus with a positive-sense single-stranded RNA genome that causes respiratory diseases and acute flaccid myelitis, posing significant threats to human health. However, an effective vaccine remains undeveloped. SIRT1, a nicotinamide adenine dinucleotide (NAD+)-dependent enzyme, plays a key role in cellular metabolism, but its interaction with NAD+ during viral infections is not well understood. In this study, through a metabolomics analysis, we demonstrate that EV-D68 infection influences cellular metabolism. Additionally, we show that NAD+ inhibits EV-D68 infection both in vivo and in vitro. EV-D68 reduces cellular NAD+ levels by regulating the expression of enzymes involved in NAD+ consumption and synthesis. Moreover, the infection increases the expression of sirtuin 1 (SIRT1), which inhibits EV-D68 replication in turn. Mechanistically, SIRT1 suppresses EV-D68 5′UTR-mediated translation, and the antiviral effect of SIRT1 on EV-D68 replication is enhanced by NAD+. Collectively, our findings highlight the critical role of NAD+ metabolism in EV-D68 infection and reveal the antiviral potential of SIRT1, providing valuable insights for the development of antiviral strategies. Full article

The tick-borne encephalitis virus is a pathogen endemic to northern Europe and Asia, transmitted through bites from infected ticks. It is a member of the Flaviviridae family and possesses a positive-sense, single-stranded RNA genome encoding a polypeptide that is processed into seven non-structural and three structural proteins, including the envelope (E) protein. The glycosylation of the E protein, involving a single N-linked glycan at position N154, plays a critical role in viral infectivity and pathogenesis. Here, we dissected the entire glycosylation profile of the E protein using liquid chromatography-tandem mass spectrometry and identified three novel O-linked glycans, which were found at relatively low frequency. One of the O-linked glycans was positioned close to the highly conserved N-linked glycan site, and structural analysis suggested that it may be relevant for the function of the E 150-loop. The N154 site was found to be glycosylated with a high frequency, containing oligomannose or complex-type structures, some of which were fucosylated. An unusually high portion of oligomannose N-linked glycan structures exhibited compositions that are normally observed on proteins when they are translocated from the endoplasmic reticulum to the trans-Golgi network, suggesting disruption of the glycan processing pathway in the infected cells from which the E protein was obtained. Full article

Duck Tembusu virus (DTMUV), a novel positive-sense RNA virus, has caused significant economic losses in the poultry industry of Eastern and Southeast Asia since its outbreak in 2010. Furthermore, the rapid transmission and potential zoonotic nature of DTMUV pose a threat to public health safety. In this study, a 4D-DIA quantitative proteomics approach was employed to identify differentially expressed cellular proteins in DTMUV-infected DF-1 cells, which are routinely used for virus isolation and identification for DTMUV, as well as the development of vaccines against other poultry viruses. One hundred fifty-seven differentially expressed cellular proteins were identified, including 84 upregulated and 73 downregulated proteins at 48 h post-infection, among which CXCL8, DDX3X, and TRPV2 may play crucial roles in viral propagation. Notably, for the upregulated protein TRPV2, the DTMUV replication was inhibited in TRPV2-low-expressing DF-1 cells. In summary, our research represents the application of 4D-DIA quantitative proteomics to analyze the proteomic landscape of DTMUV-infected poultry cells. These findings may provide valuable insights into understanding the interaction mechanism between DTMUV and poultry cells, as well as the identification of disease-resistant host factors in poultry breeding research. Full article

Dengue virus (DENV) is an enveloped, positive sense, single-stranded RNA virus belonging to the Flaviviridae. Translation initiation of the DENV mRNA (vRNA) can occur following a cap-dependent, 5′-3’end-dependent internal ribosome entry site (IRES)-independent or IRES-dependent mechanism. This study evaluated the activity of DENV IRES in BHK-21 cells and the role of the polypyrimidine-tract binding protein (PTB) isoforms PTB1, PTB2, and PTB4 as IRES-transacting factors (ITAFs) for the DENV IRES. The results show that DENV-IRES activity is stimulated in DENV-replicating BHK-21 cells and cells expressing the Foot-and-mouth disease virus leader or Human rhinovirus 2A proteases. Protease activity was necessary, although a complete shutdown of cap-dependent translation initiation was not a requirement to stimulate DENV IRES activity. Regarding PTB, the results show that PTB1 > PTB2 > PTB4 stimulates DENV-IRES activity in BHK-21 cells. Mutations in the PTB RNA recognition motifs (RRMs), RRM1/RRM2 or RRM3/RRM4, differentially impact PTB1, PTB2, and PTB4’s ability to promote DENV IRES-mediated translation initiation in BHK-21 cells. PTB1-induced DENV-IRES stimulation is rescinded when RRM1/RRM2 or RRM3/RRM4 are disrupted. Mutations in RRM1/RRM2 or RRM3/RRM4 do not affect the ITAF activity of PTB2. Mutating RRM3/RRM4, but not RRM1/RRM2, abolishes the ability of PTB4 to stimulate the DENV IRES. Thus, PTB1, PTB2, and PTB4 are ITAFs for the DENV IRES. Full article

N6-methyladenosine (m⁶A) is the most prevalent internal RNA modification. Here, we demonstrate that coxsackievirus B3 (CVB3), a common causative agent of viral myocarditis, induces m⁶A modification primarily at the stop codon and 3′ untranslated regions of its genome. As a positive-sense single-stranded RNA virus, CVB3 replicates exclusively in the cytoplasm through a cap-independent translation initiation mechanism. Our study shows that CVB3 modulates the expression and nucleo-cytoplasmic transport of the m⁶A machinery components—METTL3, ALKBH5 and YTHDFs—resulting in increased m⁶A modifications that enhance viral replication. Mechanistically, this enhancement is mediated through YTHDF-driven stress granule (SG) formation. We observed that YTHDF proteins co-localize with human antigen R (HuR), a protein facilitating cap-independent translation, in SGs during early infection. Later in infection, YTHDFs are cleaved, suppressing SG formation. Notably, for the first time, we identified that during early infection CVB3’s RNA-dependent RNA polymerase (3D) and double-stranded RNA (dsRNA) are stored in SGs, co-localizing with HuR. This early-stage sequestration likely protects viral components for use in late-phase replication, when SGs are disrupted due to YTHDF cleavage. In summary, our findings reveal that CVB3-induced m⁶A modifications enhance viral replication by regulating YTHDF-mediated SG dynamics. This study provides a potential therapeutic strategy for CVB3-induced myocarditis. Full article

SARS-CoV-2 is a spherical, positive-sense, single-stranded RNA virus with a large genome, responsible for encoding both structural proteins, vital for the viral particle’s architecture, and non-structural proteins, critical for the virus’s replication cycle. Among the non-structural proteins, two cysteine proteases emerge as promising molecular targets for the design of new antiviral compounds. The main protease (M^pro) is a homodimeric enzyme that plays a pivotal role in the formation of the viral replication–transcription complex, associated with the papain-like protease (PL^pro), a cysteine protease that modulates host immune signaling by reversing post-translational modifications of ubiquitin and interferon-stimulated gene 15 (ISG15) in host cells. Due to the importance of these molecular targets for the design and development of novel anti-SARS-CoV-2 drugs, the purpose of this review is to address aspects related to the structure, mechanism of action and strategies for the design of inhibitors capable of targeting the M^pro and PL^pro. Examples of covalent and non-covalent inhibitors that are currently being evaluated in preclinical and clinical studies or already approved for therapy will be also discussed to show the advances in medicinal chemistry in the search for new molecules to treat COVID-19. Full article

Omsk Hemorrhagic Fever Virus (OHFV) is an RNA virus with a single-stranded, positive-sense genome. It is classified under the Flaviviridae family. The genome of this virus is 98% similar to the Alkhurma hemorrhagic fever virus (AHFV), which belongs to the same family. Cases of the virus have been reported in various regions of Saudi Arabia. Both OHFV and AHFV have similarities in pathogenic polyprotein targets. No effective and licensed vaccines are available to manage OHFV infections. Therefore, an effective and safe vaccine is required that can activate protective immunity against OHFV. The current study aimed to design a multiepitope subunit vaccine against the OHFV utilizing several immunoinformatic tools. The polyprotein of OHFV was selected and potent antigenic, non-allergenic, and nontoxic cytotoxic T-lymphocyte (CTL), helper T-lymphocyte (HTL), and linear B-lymphocyte (LBL) epitopes were chosen. After screening, eight (8) CTL, five (5) HTL, and six (6) B cell epitopes were joined with each other using different linkers. Adjuvant human beta defensin-2 was also linked to the epitopes to increase vaccine antigenic and immunogenic efficiency. The designed vaccine was docked with Toll-like receptor 4 (TLR4) as it activates and induces primary and secondary immune responses against OHFV. Codon optimization was carried out, which resulted in a CAI value of 0.99 and 53.4% GC contents. In addition, the construct was blindly docked to the TLR4 immune receptor and subjected to conformational dynamics simulation analysis to interpret the intricate affinity and comprehend the time-dependent behavior. Moreover, it was predicted that immune responses to the developed vaccine construct reported formation of strong humoral and cellular immune cells. Therefore, the proposed vaccine may be considered in experimental assays to combat OHFV infections. Laboratory experiments for the above predictions are essential in order to evaluate the effectiveness, safety, and protective properties of the subject in question. Full article