Feline infectious peritonitis (FIP) is a fatal, immune-mediated disease caused by feline coronavirus (FCoV). Although several antiviral therapies have been validated for FIP treatment, safer and more cost-effective options are still needed. CK301, a novel nucleoside analog, may represent a practical alternative to current therapies. This study evaluated the efficacy and safety of CK301 in cats with effusive FIP. In this open-label trial, 150 cats with naturally occurring effusive FIP were randomly assigned to receive oral CK301 tablets (Patent WO2023197791) at low, medium, or high doses; placebo; or subcutaneous GS-441524. Treatment outcomes were assessed based on survival, clinical improvement, viral clearance, and adverse events. After 56 days of treatment, medium- and high-dose CK301 demonstrated clinical outcomes and survival comparable to those of GS-441524. However, high-dose CK301 was associated with an increased risk of drug-induced liver injury. Subcutaneous GS-441524 was associated with injection-site reactions, including ulceration, crusting, and subcutaneous masses, whereas oral CK301 avoided these complications. In conclusion, medium-dose oral CK301 (12.5 mg/kg) significantly prolonged survival, highlighting its potential as a therapeutic option for effusive FIP. It offers a convenient, non-invasive alternative to injectable treatments, provided liver function is closely monitored for potential transaminase elevations.

Parasitic diseases are among the common health concerns in dogs and cats, with ascaridoid nematodes representing one of the major groups of zoonotic parasites. Three ascaridoids-Toxocara canis, Toxocara cati, and Toxascaris leonina-commonly infect these hosts. Effective diagnostic techniques are important for systematics, epidemiology, and clinical management. In this study, a multiplex PCR (mPCR) assay was developed for the simultaneous and specific identification of three ascaridoids. Specific primers targeting conserved mitochondrial regions were designed. Sensitivity was evaluated using serially diluted standard DNA samples, and specificity was assessed by testing cross-reactivity against a Madin-Darby canine kidney cell line, Ancylostoma caninum, Dipylidium caninum, Trichinella spiralis, and Escherichia coli. The reliability of the assay was validated using 60 clinical samples collected from dogs and cats. The experimental results demonstrated that the detection limit of the assay was 10² copies/µL for a single ascaridoid and 10 ³ copies/µL for the simultaneous detection of three ascaridoids. Furthermore, the assay exhibited good specificity and reliability. The assay provides a practical tool for diagnosing and monitoring major ascaridoid infections in dogs and cats, with potential applications in source tracing of human toxocariasis.

[This corrects the article DOI: 10.3389/fvets.2025.1560273.].

[This corrects the article DOI: 10.3389/fvets.2025.1641963.].

Developing novel antiviral drugs has always been a significant forefront in biological medicine research. Antiviral drugs can be extracted, purified, and synthesized from various biological sources and by different methods. However, they are less explored in veterinary medicine for animal viruses. This research primarily selected feline calicivirus (FCV) to screen the novel antiviral drug against animal viruses. A preliminary screening from a natural product library was conducted, with subsequent assessments to ascertain their toxicity levels and antiviral capabilities. The results showed that decitabine and alprostadil were effective in reducing FCV replication. The decitabine (5-aza-2 '-deoxycytidine) was selected for antiviral mechanism investigation. Decitabine has been proven to modulate gene expression through its demethylating effect. Thus, we carried out further experiments and found that decitabine inhibited the FCV by enhancing the transcription of the feline Retinoic acid-inducible gene I (RIG-I) gene. Moreover, we also validated the same antiviral effect and mechanism of decitabine against the canine influenza virus (CIV). In summary, this study unveils the antiviral role of decitabine against FCV and CIV and provides evidence and novel insights into the demethylation drug-mediated antiviral effect for animal RNA viruses.

Recent reports have highlighted the increasing frequency of influenza A virus (IAV) spillover events from other species to dogs and cats. IAV, particularly the H3 subtype, exhibits a broad host range and a propensity for interspecies transmission, as exemplified by the sustained circulation of H3N2 and H3N8 canine influenza viruses in dog populations. This raises concerns about the potential role of companion animals as intermediate hosts in influenza virus transmission. To evaluate the susceptibility of dogs and cats to the prevalent H3 subtype influenza viruses, we experimentally inoculated groups of both species with three prevalent influenza viruses: H3N2 avian influenza virus (AIV), H3N8 avian influenza virus, and H3N2 swine influenza virus (SIV). Results showed that while all inoculated dogs exhibited seroconversion to all three viruses at 7, 14, and 21 days post-inoculation (dpi), they displayed no clinical signs, viral shedding, or evidence of viral replication in their organ tissues. In contrast, despite the cats did not exhibit apparent clinical signs, all inoculated cats exhibited seroconversion to all viruses at 7, 14 and 21 dpi, sustained nasal viral shedding for approximately one week, and demonstrated viral replication in their lungs, trachea, and nasal turbinate. Our findings underscore the higher susceptibility of cats compared to dogs to H3 subtype influenza viruses. These results emphasize the critical need for enhanced surveillance of cats within the influenza virus transmission network.

Influenza A virus (IAV) remains a major global health threat. Its M2 protein plays crucial roles in viral fusion, transportation, assembly, and release. Recent studies have shown that IAV impairs host autophagy flux to enhance viral replication. However, the precise mechanisms by which IAV M2 manipulates host cellular autophagy during virus replication remain unclear. In this study, we analysed cellular transcriptional responses of cells to IAV M2 overexpression and identified RAB GTPase protein RAB33B as a key factor. RAB33B was significantly up-regulated by IAV M2 and promoted IAV replication by enhancing autophagy. We further found that autophagy regulates the interaction of IAV M2, RAB33B, and LC3, facilitating M2 membrane trafficking through autophagic-like vesicles. In addition, ATG16L1 (an effector of RAB33B) and TBC1D25 (a GTPase-activating protein for RAB33B) contributed to IAV M2-induced autophagy, thereby affecting viral replication. Collectively, our findings reveal a novel mechanism in which RAB33B is essential for IAV M2 trafficking to the plasma membrane, facilitating viral replication through enhanced autophagy. These insights shed new light on the autophagy-based cellular transport mechanisms of IAV M2 and highlight potential antiviral targets.

Lumpy skin disease (LSD), caused by the Lumpy skin disease virus (LSDV), poses a significant threat to the global cattle farming industry. Viral proteins evolved the ability to escape host immune responses. Here, we demonstrate that LSDV infection inhibits interferon-β (IFN-β) production in Madin-Darby bovine kidney (MDBK) cells, even in the presence of Sendai virus (SEV) inducers. Through further investigation, we identify multiple genes at both ends of the LSDV genome that strongly inhibit IFN-β or NF-κB activation of the promoter. Notably, ORF142 selectively inhibits IFN-β promoter activity but fails to inhibit NF-κB promoter activity. Subsequently, we reveal that ORF142 negatively regulate the cGAS/STING-mediated IFN-I production. Overexpression of ORF142 suppresses IFN-β and interferon-stimulated response element (ISRE) activity, while the absence of ORF142 upregulates the transcription levels of IFN-β and interferon-stimulated genes (ISGs) in MDBK cells. Mechanistically, ORF142 interacts with STING to facilitate autophagy-lysosomal degradation by recruiting NBR1, thereby impeding the activation of downstream signaling molecules, such as IRF3, and inhibiting IFN-I production. In conclusion, our findings elucidate the immune evasion mechanism of ORF142 encoded by LSDV, offering insights for the development of antiviral drugs or attenuated live vaccines to mitigate LSD.

Feline herpesvirus-1 (FHV-1) is a leading cause of feline viral rhinotracheitis (FVR), which mainly presents upper respiratory tract symptoms. Vaccination is the most effective strategy for controlling FHV-1. Prior to the initiation of this study, China does not have domestically produced commercially available FHV-1 vaccines using field strain as antigenic component and most corresponding imported vaccines contained feline viral rhinotracheitis, calicivirus, and panleukopenia (FVRCP) antigens. However, the protective efficacy of these vaccines against the prevalent FHV-1 strains in China remains unclear. In the present study, a total of 12 cats were randomly divided into 3 groups, which were vaccinated with FHV-1 field vaccine (Group 1 [an inactivated vaccine developed by ourselves using the Chinese field strain FHV-1 2020GD02]) and FVRCP vaccine (Group 2) and PBS (Group 3) as control, respectively. These animals received two vaccinations with a 21-day interval and were challenged with 2020GD02 at 21 days after the second vaccination. Clinical signs, serological responses, viral shedding, and histopathological changes were used to estimate protective efficacy of the two vaccines. Compared to Group 2, animals in Group 1 produced higher level FHV-1 antibody titers during immune processes. After challenge, Group 3 developed typical FVR. In contrast, animals in both Groups 1 and 2 showed significantly fewer clinical signs, viral shedding, and pathological changes, but could not provide complete protection. Our results provided a reference for further FHV-1 vaccines development in China.

Introduction:The bovine respiratory disease complex poses a significant threat to the cattle industry, necessitating a multifaceted approach to address its occurrence. The syndrome is caused by various pathogens such as bovine respiratory syncytial virus (BRSV), bovine parainfluenza virus type 3 (BPIV3), bovine viral diarrhea virus (BVDV), bovine adenovirus type 3 (BAV3), Mycoplasma bovis (Mb), and infectious bovine rhinotracheitis virus (IBRV). The confluence of these pathogens causes substantial economic losses to the cattle industry. Although preventive and control measures have been implemented, containment of bovine respiratory diseases continues to present a formidable challenge, highlighting the need for innovative diagnostic and intervention strategies. Methods:In this study, we designed specific primers targeting six conserved pathogen genes (N of BRSV, M of BPIV3, 5'UTR of BVDV, Hexon of BAV3, oppF of Mb, and gB of IBRV). Subsequently, we established a multiplexed fluorescent real-time quantitative PCR (qPCR) assay for simultaneous detection of these pathogens. Results:The developed method exhibited high specificity and sensitivity, with the lowest detection limits for plasmid DNA standards of BRSV, BPIV3, BVDV, BAV3, Mb, and IBRV being 70.1, 40.4, 15.1, 74.4, 69.6, and 4.99 copies/μL, respectively. The coefficients of variation determined by the assay established in this study were <4%, and the amplification efficiency was 93.84%-111.60%, which showed the reliability and stability of the method. Discussion:The detection rates for BRSV, BPIV3, BVDV, BAV3, Mb, and IBRV were 7.59% (17/224), 11.61% (26/224), 8.04% (18/224), 22.32% (50/224), 27.23% (61/224), and 8.04% (18/224), respectively. All 224 cows were cases of natural disease. Fifty-six diseased cattle were infected with a mixture of two or more of the six pathogens at a mixed infection rate of 25% (56/224). Therefore, this study successfully developed a highly efficient, rapid, specific, and sensitive multiplex qPCR method to detect major pathogens associated with bovine respiratory diseases. This advancement is expected to significantly influence the future of the cattle industry and serve as a valuable reference for subsequent research in this field.

ABSTRACT As companion animals, dogs are susceptible to various subtypes of influenza A virus (IAV), with the H3N2 and H3N8 subtypes of canine influenza virus (CIV) stably circulating among canines. Compared to the H3N8 CIV, the H3N2 CIV is more widely prevalent in canine populations and demonstrates increased adaptability to mammals, potentially facilitating cross-species transmission. Therefore, a comprehensive elucidation of the mechanisms underlying H3N2 CIV adaptation to mammals is imperative. In this study, we serially passaged the GD14-WT strain in murine lungs, successfully establishing a lethal H3N2 CIV infection model. From this model, we isolated the lethal strain GD14-MA and identified the key lethal mutations PA(S184N) and PB2(E627K). Moreover, the GD14-ma[PA(S184N)+PB2(E627K)] strain exhibited markedly enhanced pathogenicity in dogs. Viral titers in lung tissues from infected dogs and mice showed that GD14-ma[PA(S184N)+PB2(E627K)] does not increase its pathogenicity to mice and dogs by upregulating viral titers compared to the GD14-WT strain. Notably, sequence alignments across all H3N2 IAVs showed an increasing prevalence of the PA (S184N) and PB2 (E627K) mutations from avian to human hosts. Finally, single-cell RNA sequencing of infected mouse lung tissues showed that GD14-ma[PA(S184N)+PB2(E627K)] effectively evaded host antiviral responses, inducing a robust inflammatory reaction. Considering the recognized role of the PB2 (E627K) mutation in the mammalian adaptation of IAVs, our findings underscore the importance of ongoing surveillance for the PA (S184N) mutation in H3N2 IAVs. IMPORTANCE Since the 21st century, zoonotic viruses have frequently crossed species barriers, posing significant global public health challenges. Dogs are susceptible to various influenza A viruses (IAVs), particularly the H3N2 canine influenza virus (CIV), which has stably circulated and evolved to enhance its adaptability to mammals, including an increased affinity for the human-like SAα2,6-Gal receptor, posing a potential public health threat. Here, we simulated H3N2 CIV adaptation in mice, revealed that the synergistic PA(S184N) and PB2(E627K) mutations augment H3N2 CIV pathogenicity in dogs and mice, and elucidated the underlying mechanisms at the single-cell level. Our study provides molecular evidence for adapting the H3N2 CIV to mammals and underscores the importance of vigilant monitoring of genetic variations in H3N2 CIV.

BackgroundAnemoside B4 (AB4), a pentacyclic triterpenoid saponin extracted from the traditional Chinese medicinal herb Pulsatilla chinensis, has shown anti-inflammatory and immunomodulatory effects in both preclinical and clinical studies. However, pharmacokinetic and safety data in dogs remain limited. This study aimed to evaluate the pharmacokinetics, bioavailability, and safety of AB4 in healthy Beagle dogs.MethodsIn the single-dose pharmacokinetic study, 40 dogs received subcutaneous AB4 at 10, 20, or 40 mg/kg, or an intravenous bolus at 20 mg/kg. Plasma concentrations were measured using a validated HPLC–MS/MS method to determine pharmacokinetic parameters, bioavailability, dose proportionality, and sex-related differences. In the repeated-dose study, 10 dogs received 20 mg/kg subcutaneously once daily for 7 consecutive days to evaluate drug accumulation and fluctuation. In the target animal safety study, 32 dogs were randomly assigned to receive 1× (20 mg/kg), 3× (60 mg/kg), or 5× (100 mg/kg) doses of AB4, and saline as a control, via daily subcutaneous injection for 7 days. Routine clinical examinations, hematology, serum biochemistry, gross necropsy, and histopathology were assessed.ResultAB4 exhibited rapid elimination, high absolute bioavailability, and dose-proportional pharmacokinetics in the 10–40 mg/kg range. No evidence of accumulation after repeated dosing. Within the dose range of 20–100 mg/kg, AB4 demonstrated good safety, with no observable toxicity or adverse effects. No significant effects were observed on physiological parameters. Histopathological analysis revealed no consistent or target-organ specific lesions.DiscussionThese findings provide fundamental pharmacokinetic and safety data to support the rational clinical use of AB4 in veterinary medicine and lay the groundwork for future clinical applications.

Bovine viral diarrhea-mucosal disease (BVD-MD), caused by bovine viral diarrhea virus (BVDV), is a major infectious disease affecting the cattle industry. The nonstructural protein NS4B of BVDV has been shown to induce autophagy and antagonize the expression of the host innate immune sensor MDA5. However, the precise mechanism underlying NS4B-mediated suppression of MDA5 remains unclear. In this study, we demonstrate that NS4B interacts with the host protein CCDC50, as confirmed by co-immunoprecipitation and indirect immunofluorescence assays. Stable overexpression of CCDC50 significantly promoted MDA5 degradation and enhanced BVDV replication, whereas CCDC50 knockdown markedly impaired NS4B-mediated degradation of MDA5 in HEK293T cells. Interestingly, CCDC50 also suppressed the production of type I interferons (IFN-I) and interferon-stimulated genes (ISGs) during BVDV infection. Furthermore, the translocation of CCDC50 reduced MDA5 protein levels through the autophagy pathway. Mechanistically, we found that CCDC50 interacts with MDA5 and facilitates NS4B-mediated autophagic degradation of MDA5. These findings uncover a novel mechanism by which BVDV NS4B hijacks the host protein CCDC50 to subvert antiviral innate immunity by promoting MDA5 autophagic degradation and suppressing IFN-I signaling.

BackgroundCanine pneumonia is a serious respiratory disease often associated with Canine Infectious Respiratory Disease (CIRD). Current treatment strategies primarily rely on antibiotics and corticosteroids; however, the emergence of antibiotic resistance and potential side effects from prolonged corticosteroid use limit the effectiveness of these therapies in clinical practice. These challenges highlight the urgent need for alternative treatments. Anemoside B4 (AB4), derived from the traditional Chinese medicine Pulsatilla, has shown promise in preclinical studies for modulating inflammatory responses and improving clinical symptoms of pneumonia. Therefore, AB4 may offer a valuable alternative treatment option for canine pneumonia in veterinary medicine.MethodsA prospective, randomized controlled trial was conducted at the Veterinary Drug Research and Evaluation Center of South China Agricultural University. Seventy-two dogs with mild-to-moderate pneumonia were randomly assigned to one of three groups: AB4, placebo, or Chuanxinlian injection (CXL). The primary outcome was the effect of AB4 on comprehensive clinical scoring of canine pneumonia; secondary outcomes included recovery times for primary symptoms and efficacy assessments. Additionally, AB4′s safety in clinical applications was evaluated.ResultsThe AB4 group demonstrated significantly lower composite clinical scores on Days 7 and 14 compared to the placebo group (p = 0.033 and p = 0.000, respectively). Significant differences in recovery times for fever and dyspnea were observed between the AB4 and placebo groups (p = 0.041 and p = 0.024, respectively). Moreover, the cure rate and overall efficacy on Day 14 were significantly higher in the AB4 group than those in the placebo group (p = 0.001 and p = 0.009, respectively).ConclusionThese findings suggest that AB4 may be a promising treatment option for canine pneumonia, potentially serving as an alternative to traditional therapies. Further research is needed to explore its clinical potential in veterinary medicine.

Canine influenza virus (CIV) is a respiratory pathogen that causes fever, coughing, and sneezing in dogs and is continuously circulating in canine populations. Tetherin is an antiviral host restriction factor mediated by interferon, capable of inhibiting the release of enveloped viruses from infected cells. The antiviral mechanism of tetherin is mainly due to its unusual topology, which includes a short N-terminal cytoplasmic tail (CT), a transmembrane (TM) domain, a coiled-coil extra-cellular region (CC), and a C-terminal glycosyl-phosphatidylinositol anchor (GPI). Previous studies have found that canine tetherin has the ability to limit the release of CIV, but its main antiviral domain remains unclear. In the present study, the potential CT, TM, CC, and GPI domains of canine tetherin were predicted through systemic bioinformatic analysis, and mutational variants of canine tetherin based on the four domains were constructed. Confocal microscopy demonstrated that the CT, TM, and CC domains are critical for the cell membrane localization of canine tetherin. The results of in vitro CIV infection experiments showed that the TM region is a critical functional domain of canine tetherin in limiting the replication of CIV. Our study will help better understand the antiviral activity of canine tetherin and the role of the structural domains of canine tetherin in inhibiting the replication of CIV.

Tetherin is an interferon-induced-expressing transmembrane protein that utilizes a unique topology to restrict the release of enveloped viruses from the surface of the cell membrane. N-linked glycosylation plays an important role in protein post-translational modifications. To investigate the role of glycosylation in the antiviral activity of canine tetherin, its potential glycosylation sites were predicted and mutated, and the effects of glycosylation site mutations or treatment with a glycosylation inhibitor on the ability of canine tetherin to restrict H3N2 canine influenza virus (CIV) replication were examined. Mutations in the glycosylation sites of canine tetherin (N72A, N99A, and N72,99A) lead to changes in its intracellular distribution and weakened or even lost antiviral activity against H3N2 CIV. Similarly, the subcellular localization of tetherin after tunicamycin treatment was altered, and its antiviral activity was weakened. Colocalization analysis revealed that the colocalization of canine tetherin and H3N2 CIV protein was weakened under the condition of impaired glycosylation. These results indicate that canine tetherin maintains its localization in the cell membrane through glycosylation and exerts its antiviral activity. This study provides new insights into the antiviral mechanisms of host restriction factors and offers a theoretical basis for developing small-molecule anti-influenza strategies targeting glycosylation modifications.

Melatonin improves chronic stress-induced hippocampal damage and depression-like behaviors, but the mechanism needs further study. This study was to explore the mechanism of melatonin inhibiting microglia pyroptosis. In virtro experiments, melatonin improved corticosterone-induced the ultrastructure and microstructure damage of HAPI cells by inhibiting pyroptosis, thereby increasing cell survival rate. Protein-protein interaction network and molecular autodocking predicted that Cathespin B might be the target of melatonin inhibition of NLRP3-mediated pyroptosis. Melatonin inhibited corticosterone-induced Cathespin B expression. Both Cathepsin B inhibitor CA-074Me and NLRP3 knockout inhibited the HAPI cells pyroptosis. Similarly, melatonin inhibited Cathepsin B agonist Pazopanib-induced activation of Cathepsin B/NLRP3 signaling pathway and HAPI cells pyroptosis. In vivo studies, melatonin inhibited chronic restraint stress (CRS)-induced activation of Cathepsin B/NLRP3 signaling pathway and alleviated hippocampal microglia pyroptosis in rats. Inhibition of microglia pyroptosis improved CRS-induced depression-like behaviors of rats. In addition, inhibition of Cathepsin B and NLRP3 alleviated hippocampal pyroptosis. Melatonin inhibited Pazopanib-induced activation of Cathepsin B/NLRP3 signaling pathway and hippocampal pyroptosis. These results demonstrated that melatonin could alleviate CRS-induced hippocampal microglia pyroptosis by inhibiting Cathepsin B/NLRP3 signaling pathway, thereby improving depression-like behaviors in rats. This study reveals the molecular mechanism of melatonin in the prevention and treatment of chronic stress-related encephalopathy.

IntroductionBovine torovirus (BToV), Bovine enterovirus (BEV), Bovine norovirus (BNoV), Bovine coronavirus (BCoV), Bovine rotavirus (BRV), and Bovine viral diarrhea virus (BVDV) are significant pathogens causing diarrhea in calves, characterized by their high prevalence and challenging prevention and control measures.MethodsWe analyzed 295 calf diarrhea samples, amplifying the M gene from BToV-positive samples, the 5’UTR gene from BEV-positive samples, the RdRp gene from BNoV-positive samples, the VP7 gene from BRV-positive samples, the S gene from BCoV-positive samples, and the 5’UTR gene from BVDV-positive samples. Subsequent homology analysis and phylogenetic tree construction were performed.ResultsThe overall viral positive rate in Guangdong Province was 21.36%. Specific detection rates were as follows: Foshan City at 50.00% (18/36), Guangzhou City at 43.90% (36/82), Huizhou City at 21.21% (7/33), Yangjiang City at 2.08% (1/48), Meizhou City at 1.39% (1/72), and Heyuan City at 0.00% (0/24). The detection rates for BToV, BEV, BNoV, BCoV, BRV, and BVDV were 0.34% (1/295), 6.10% (18/295), 0.68% (2/295), 1.36% (4/295), 10.85% (32/295), and 2.03% (6/295), respectively. Notably, the highest overall virus detection rate was observed in the Guangzhou-Foshan region, with BRV and BEV showing the highest detection rates among the six viruses. This study marks the first report of BToV and BNoV in Guangdong Province. Phylogenetic analysis revealed that the BToV strain belonged to type II, sharing genetic similarities with epidemic strains from various provinces in China. The BEV strains were categorized into E and F types, with the F type being the predominant strain in Guangdong Province and exhibiting the closest genetic relationship to strains from Heilongjiang and Guangxi. The BNoV strains, along with Hebei strains, were identified as GIII.2 subgenotype. BCoV strains showed the highest genetic similarity to strains from Sichuan. All BRV strains were classified under the G6 subtype and had the closest genetic relationship with human rotavirus strains. BVDV strains were identified as subtype 1b, closely related to the Beijing strain. In conclusion, this study investigated the prevalence and evolutionary characteristics of diarrhea-associated viruses in calves in specific areas of Guangdong Province, providing a valuable reference for establishing effective prevention and control measures in cattle farms.