Introduction: Canine distemper virus (CDV) and canine coronavirus (CCoV) are two highly contagious viruses that primarily affect dogs, causing disease in the gastrointestinal and respiratory systems, and sometimes the neurological system. Despite advances in vaccines and antiviral therapies, many viruses remain difficult to treat, underscoring the need for innovative control strategies. This study explores the antiviral activity of two antimicrobial peptides, pantinin-1 and pantinin-2, derived from the venom of the scorpion Pandinus imperator, against CDV and CCoV.

Methods: The peptides were prepared via solid-phase peptide chemical synthesis and assessed for their cytotoxicity in canine fibrosarcoma cells (A-72), and Vero/hSLAM cells, using MTT assay. Non-cytotoxic concentrations of both peptides were tested for antiviral activity primarily through plaque reduction assays and tissue culture infective dose (TCID₅₀), alongside quantification of viral N gene expression levels. Furthermore, peptide conformations were examined in aqueous buffer and in a membrane/mimetic environment (trifluoroethanol (TFE)/H₂O) using circular dichroism (CD) and NMR analyses.

Results: The results show that both pantinins exert antiviral effects within a concentration range from 6 µM to 50 µM against both viruses. Peptides affect viral particles by exerting a virucidal effect and preventing the fusion and entry of the virus into host cells. These results were confirmed by means of qPCR, showing complete suppression of N viral gene expression at the highest concentration of both pantinins (50 μM). In TFE/H₂O, both peptides shift to α-helical structures, which are often associated with membrane interaction and significantly influence antiviral activity.

Conclusion: These findings show that pantinin-1 and pantinin-2 have promising antiviral properties, supporting their potential as therapeutic agents against CDV and CCoV infections.

Further studies are needed to explore their mechanism, optimise antiviral potency, and assess safety in animal models.

Apoptosis, also known as programmed cell death type I (PCD I), is a phenomenon responsible for the proper development and functionality of animal organisms. It enables the elimination of old, damaged, malfunctioning, oncogenic, and/or infected cells. As an essential determinant of organismal health, apoptosis is subject to multi-level regulation. PCD I can be induced via two molecular pathways: the extrinsic and the intrinsic. The latter is mitochondria-dependent – the key event in this signaling cascade is the permeabilization of the outer mitochondrial membrane (OMM), leading to the release of cytochrome c, apoptosome formation, the proteolysis of caspase-9, and, finally, the activation of caspase-3, an effector enzyme of apoptosis. Bid is a member of the Bcl-2 family, which comprises proteins responsible for regulating mitochondrial integrity. Its active form, truncated Bid (tBid), can promote OMM permeabilization, partly by recruiting Bax protein to the OMM, resulting in Bax oligomerization into pores that perforate the membrane. As Bid is cleaved to tBid downstream of surface receptor activation, it serves as a link integrating the extrinsic and intrinsic apoptotic pathways. Our study aimed to determine the influence of the ectromelia virus Moscow strain (ECTV-MOS) infection of L929 fibroblast on Bid activation and the consequent events, including colocalization of Bax with the OMM, caspase-3 activation, and changes in the structure and integrity of the cell membrane, measured cytometrically using an annexin–propidium iodide kit. We used Western blot to determine the level of Bid and active caspase-3, confocal microscopy to examine the Bax-OMM colocalization, and flow cytometry to assess the percentage of apoptotic cells in control as well as in ECTV-infected and/or staurosporine (STS)-treated fibroblasts. ECTV-MOS infection did not activate Bid, but counteracted STS-induced apoptosis by decreasing intracellular Bid levels, Bax-OMM colocalization, and caspase-3 activation. The results suggest that ECTV-MOS actively inhibits mitochondria-dependent PCD I.

This study provides a comprehensive and clear investigation into the critical role of host glutamine metabolism in the replication cycle of Avian Leukosis Virus Subgroup J (ALV-J). We hypothesized that ALV-J reprograms host cell metabolism to support its replication. Using both an HD11 macrophage cell line and a chicken infection model, we demonstrated through qPCR and western blot analysis that ALV-J infection significantly upregulates key genes involved in glutamine metabolism, particularly glutaminase (GLS) and the solute carrier transporters SLC1A5 and SLC38A2. In vitro, pharmacological inhibition of GLS with the specific inhibitor BPTES, or culturing under glutamine-deprived conditions, substantially reduced viral replication (as measured by p27 antigen levels) and progeny virion production (via viral titers). Conversely, supplementing the culture medium with exogenous glutamine significantly enhanced ALV-J propagation in a dose-dependent manner. Consistent with cellular findings, in vivo experiments showed that ALV-J infection enhances glutamine metabolic activity—evidenced by increased enzyme activity and metabolite levels—in target organs such as the bursa of Fabricius and the spleen. Importantly, therapeutic administration of BPTES effectively suppressed viral loads in these tissues and alleviated virus-induced pathological damage. Our results conclusively demonstrate that ALV-J exploits host glutamine metabolism to facilitate its replication and propose that targeting this pathway with inhibitors such as BPTES represents a novel and promising therapeutic strategy against ALV-J infection and associated diseases. Further investigation into the specificity and safety of glutamine-targeting approaches will help advance their translational potential in poultry health.

Goose astrovirus type 2 (GAstV-2) has recently emerged as a causative agent of fatal gout in goslings, causing significant economic losses to the waterfowl industry. This study aimed to isolate and characterize a novel GAstV-2 strain to understand its genetic evolutionary characteristics.A virus strain was isolated from clinical samples of diseased geese collected from a farm in Taizhou, Jiangsu province, and the virus was isolated by inoculating chicken liver cancer cells. The virus was identified by electron microscopy and RT-PCR. The complete genome was sequenced and analyzed using bioinformatic tools for phylogenetic relationship and genetic variation.A novel GAstV-2 strain, designated as GAstV/Goose/China/TZ04/2025, was successfully isolated. Typical astrovirus-like particles were observed under an electron microscope. The complete genome was determined to be 7018 in length. Phylogenetic analysis based on the complete genome and ORF2 gene revealed that the isolate clustered within the GAstV-2 clade but formed a distinct branch. It shared 97.55% - 99.86% nucleotide identity with other known GAstV-2 strains. Notably, several unique amino acid substitutions were identified in the capsid protein.This is the first report on the isolation and genomic characterization of a novel GAstV-2 variant. The findings expand our knowledge of the genetic diversity of GAstV-2 and provide valuable information for developing diagnostic tools and vaccines against this virus.

Introduction
Herpesvirus (HV) infections in cetaceans are increasingly recognized as indicators of health disturbances due to their association with immunosuppression, stress, and co-infections. However, its full clinical and ecological significance remains underexplored.

Our research in this field compiles and analyzes findings from stranded cetaceans in the Western Mediterranean, offering critical insights into herpesvirus tropism and pathology, emergence in new hosts, and its relevance in One Health frameworks.

Methods
Over 1,000 post-mortem tissue samples from 49 cetaceans, representing five different species, stranded between 2010 and 2022 were analyzed using conventional PCR, sequencing, and histopathology. HV RNA detection was employed to assess viral replication.

Results

• Ecological expansion: an overall prevalence of 80.85% was reported in a 2021 cohort of 47 stranded cetaceans in the Valencian Community, which was the highest recorded to date. Additionally, Alphaherpesvirus (AHV) was identified in a Mediterranean-stranded humpback whale (Megaptera novaeangliae)—the first such report in the region and only the second globally. These findings suggest both wide circulation and an expanded host range.
• Tropism: HV was detected in diverse tissues—skin, genital mucosa, and notably, the central nervous system (CNS). Gammaherpesvirus (GHV) sequences were associated with CNS inflammation and meningitis, representing the first evidence of neurotropism in odontocetes.
• Clinical pathology: HV-associated lesions ranged from proliferative to ulcerative and inflammatory changes. Some animals exhibited histological lesions with active HV replication in the absence of overt clinical signs.
• Genetic diversity and replication: most sequences belonged to GHV, while AHV showed greater genetic heterogeneity. RNA detection confirmed active replication, particularly in juveniles and neonates—supporting age-related susceptibility.

Conclusions
Our integrated assessment identifies HV as a complex, multi-organ pathogen with underrecognized impacts on cetacean health and conservation. Integrating HV surveillance within a One Health framework will enhance our capacity to detect early signs of marine ecosystem disruption through sentinel species like cetaceans.

Bovine coronavirus (BCoV) represents a significant etiological agent of enteric disease, exhibiting widespread endemicity and imposing substantial economic burdens on the livestock industry through its pronounced pathogenicity in neonatal calves. Within the betacoronavirus lineage A, several species including human coronaviruses (HKU1, OC43), equine coronavirus (ECoV), murine hepatitis virus (MHV), canine respiratory coronavirus (CRCoV), and porcine hemagglutinating encephalomyelitis virus (PHEV) demonstrate phylogenetic relatedness to BCoV. As an enveloped RNA virus with a positive-sense, single-stranded genome, BCoV expresses the hemagglutinin–esterase (HE) glycoprotein, which functions as an auxiliary receptor-binding protein and potentially influences viral evolution through modifications in host range determination and tissue tropism.

This investigation analyzed fecal specimens (n=188) obtained from diarrheic calves to assess HE gene variability. Total RNA was extracted following standardized protocols, and HE-specific amplification was performed using targeted primers yielding 497-bp amplicons. Electrophoretic visualization confirmed successful amplification, with molecular detection revealing BCoV prevalence of 15.42% (29/188 samples). Bidirectional Sanger sequencing was conducted on selected positive isolates, and comparative sequence analysis was performed using BLAST algorithms against established betacoronavirus databases. Molecular characterization identified distinct amino acid polymorphisms in isolates designated HE1, HE10, HE30, and HE40. Phylogenetic reconstruction demonstrated that isolates HE1, HE10, and HE40 exhibited monophyletic clustering, whereas isolate HE30 displayed divergent evolutionary positioning, clustering proximally with SARS-CoV-2 variants of Chinese, Israeli, and American origin.

Our investigation into the effects of the identified mutations in the HE gene is currently ongoing.

Viruses are among the simplest pathogens and rely on host cell metabolic machinery for multiplication. However, viruses also represent complex macromolecular assemblies and can therefore be analyzed as biothermodynamic systems. This means that virus–host interactions represent processes that proceed in accordance with the laws of physics and chemistry. The biothermodynamic methodology has been extensively applied in science and engineering to analyze microorganisms, which include bacteria, yeasts, filamentous fungi, and algae. Gibbs energy represents the driving force of metabolism and multiplication of microorganisms. In this research, the computational and theoretical methodology of biothermodynamics is applied to analyze virus particles, which include the atom counting method, Patel–Erickson–Battley model, and biosynthesis reactions. Based on the determined chemical and thermodynamic properties of virus particles, a mechanistic model of virus–host interactions is developed, based on chemical and nonequilibrium thermodynamics. Virus–host interactions at the cell membrane (antigen-receptor binding) and in the cytoplasm (virus multiplication) are analyzed. Antigen-receptor binding represents a chemical reaction that is driven by Gibbs energy. Virus multiplication can be analyzed as a chemical reaction in which precursors like amino acids are combined to form new virus particles. This process is performed by the metabolic machinery, which is also used by the host cell for reparation. This means that the processes of virus replication and host cell reparation are competitive chemical reactions. According to phenomenological equations of nonequilbrium thermodynamics, the reaction with a more negative Gibbs energy change will dominate in the competition. Therefore, a more negative Gibbs energy of biosynthesis contributes to the ability of viruses to hijack the metabolism of their host cells and multiply. Furthermore, the methodology of biothermodynamics is applied to analyze virus evolution.

Early and precise identification of illnesses linked to the bovine leukemia virus (BLV) is crucial for maintaining food safety and efficient herd management in nations like Japan, where milk and beef are staples of diets. Enzootic bovine leukosis (EBL), a B-cell lymphoma caused by BLV, emerges in approximately 5% of infected cattle after a prolonged asymptomatic phase. However, current diagnostic tools, such as serological assays and PCR detection of proviral DNA from whole blood, often lack the sensitivity and specificity needed to distinguish EBL from non-EBL cases, especially in early clinical stages. This diagnostic gap emphasizes the need for reliable, non-invasive biomarkers that reflect disease progression with greater precision. Given the increased turnover of malignant cells in EBL, we investigated the potential of plasma-derived circulating cell-free DNA (cfDNA) as a novel biomarker. Proviral loads (PVL) in whole blood and plasma were quantified using qPCR targeting the BLV LTR and pol regions in cattle at different clinical stages. Although PVL in whole blood was generally higher in EBL cases, significant overlap with non-EBL animals limited its diagnostic reliability. In contrast, BLV-derived cfDNA in plasma was significantly elevated in clinically diagnosed EBL cattle and effectively distinguished between disease states. These findings highlight cfDNA as a promising tool for liquid biopsy-based approaches in veterinary virology, with the potential to enhance early diagnosis, surveillance, and control of BLV-associated malignancies in livestock populations.

Influenza A H5N1 virus represents a global zoonotic threat, capable of infecting a wide range of hosts, including (besides birds) non-human mammals and, occasionally, humans. In recent years, the virus has exhibited unprecedented geographic expansion and genetic diversification, with numerous documented cases of infection in non-avian species, particularly those associated with clade 2.3.4.4b. The primary aim of this study was to evaluate the potential of the H5N1 virus to adapt to human hosts. In pursuit of this goal, all available nucleotide sequences of the genes encoding Hemagglutinin (HA) and Neuraminidase (NA) from the H5N1 strain—isolated from birds, non-human mammals, and humans and accessible via the GISAID platform—were analysed. Evolutionary dynamics and adaptive potential were assessed across the resulting datasets. The results reveal a significant expansion of clade 2.3.4.4b, along with the presence of mutations known to promote infection and replication in mammals. However, no distinct genetic signature was identified that would indicate spillover or confirmed adaptation to the human host. These findings suggest that, although the H5N1 virus has infected numerous mammalian species in recent years and several human cases have been reported, there is currently no evidence of sustained human-to-human transmission, and birds still remain the main hosts. Rather, humans appear to be incidental hosts, acquiring infections from strains typically circulating in avian or bovine populations, depending on geographic region, sanitary conditions, and lifestyle factors.

In recent decades, there has been a significant increase in epizootic events in aquatic species. Moreover, a notable number of pathogens typically associated with human infections have been reported in these environments, some of which are causative agents of mortality in aquatic animals. This alarming trend underscores the urgent need for comprehensive research and mitigation strategies.
In this context, we present a study focused on Callinectes sapidus reovirus 1 (CsRV1), a pathogenic virus spreading along the Atlantic coast of the Americas that may influence the distribution of its host populations, the blue crab Callinectes sapidus. CsRV1 has a double-stranded RNA genome, organized into 12 segments, and is characterized by high mutation rates, segment recombination and reassortment.
Our study investigates the genetic variability of CsRV1, aiming to understand how it is evolving along the Atlantic coast of the Americas, its area of origin. To achieve this goal, we conducted phylodynamic reconstruction using all available genomes and segments from the NCBI Virus database. Molecular dating based on whole-genome data suggests that CsRV1 emerged approximately 40 years ago, with a notable increase in genetic diversity and viral population size occurring around a decade ago.
These results not only provide an overview of the genetic variability of CsRV1 in the Americas but may also prove valuable should the virus be detected in C. sapidus specimens from other geographic areas, given that its host is a highly invasive alien species. In this context, we are currently screening C. sapidus specimens, primarily collected from Sardinia, but also from other areas of the Mediterranean Sea.
Moreover, these findings underscore the importance of monitoring aquatic pathogens, particularly when invasive alien species such as C. sapidus may serve as vectors for previously undetected pathogens that pose potential threats to native species and local ecosystems.