The global duck industry faces substantial challenges due to the emergence of novel duck reovirus (NDRV) infections, which are pathologically characterized by splenomegaly, hemorrhagic manifestations, and necrotic lesions in affected birds. Surviving ducklings often suffer from severe growth retardation. Here, we investigated an outbreak of hepatic necrosis in commercial ducklings from Shandong Province, China. After excluding other potential pathogens, an NDRV strain was isolated and designated SD416. This strain demonstrated infectivity in both chicken and duck embryos and induced syncytial formation in Vero cells. Full-genome sequencing revealed a 23,420-bp dsRNA genome consisting of 10 segments, displaying significant genetic divergence from other Chinese duck reovirus isolates. Phylogenetic analysis indicated that SD416 is genetically distinct from known DRV strains, particularly within the M2 and S3 genes. Furthermore, its σC protein-coding sequence exhibited notably high genetic variability compared to strains from other genotyping clusters. To evaluate the age-dependent pathogenicity of SD416, 1-, 7-, and 14-day-old ducklings were intramuscularly inoculated with 0.1 mL of allantoic fluid containing the virus at a titer of 1 × 10⁶ TCID₅₀/0.1 mL. The strain exhibited pronounced age-related virulence: 1-day-old ducklings developed severe hepatic and splenic lesions resulting in mortality, whereas ducklings inoculated at 7 and 14 days of age exhibited only splenic pathology and survived throughout the study. The SD416 strain was comprehensively characterized in terms of tissue tropism, pathogenesis, genomic structure, and evolutionary relationships. Our findings reveal a unique genomic and virulence pattern associated with this novel isolate, providing important insights into the biology of NDRV. These results underscore the need for enhanced surveillance and development of targeted intervention strategies to limit the spread of this virus in global waterfowl production systems.

OASL, a distinctive member of the 2'-5'-oligoadenylate synthetase (OAS) family, is a well-characterized interferon-stimulated gene (ISG). In mammals and certain avian species, OASL exhibits dual functions in antiviral defense and innate immune regulation. However, the biological role of chicken OASL (chOASL) remains poorly understood, largely due to the natural absence of RIG-I in chickens. Infectious bronchitis virus (IBV) belongs to Gammacoronavirus genus and causes substantial economic losses in the global poultry industry. To explore whether chOASL is involved in the host defense against IBV, this study aimed to comprehensively characterize its expression dynamics and antiviral function. Firstly, the anti-chOASL polyclonal antibody was generated using prokaryotically expressed recombinant protein. Using this validated tool, we demonstrated that endogenous chOASL was significantly induced by Poly(I:C) and QX-IBV in various chicken cells. Furthermore, we mapped the baseline tissue distribution of chOASL protein for the first time and proved that chOASL was markedly upregulated, particularly in immune organs and IBV target tissues during IBV infection. Importantly, overexpression and knockdown assays verified that chOASL could suppress IBV replication. In conclusion, this study not only provides a reliable tool for detecting chOASL but also establishes its specific expression profile and direct antiviral activity against IBV, laying a solid foundation for further elucidating antiviral mechanisms of chOASL.

Avian reovirus (ARV) is a major pathogen causing viral arthritis, tenosynovitis, malabsorption syndrome, and immunosuppression in chickens, leading to severe economic losses in the global poultry industry. The nonstructural protein p17 is vital for ARV replication and modulates diverse host signaling pathways by interacting with multiple host factors. In our previous work, nucleolar GTP‑binding protein 2 (GNL2), a conserved nucleolar GTPase governing 60S ribosomal subunit maturation and nucleocytoplasmic transport, was screened as a potential binding partner of ARV p17 using yeast two‑hybrid assay. In the present study, the physical interaction between p17 and GNL2 was validated by coimmunoprecipitation (Co‑IP), glutathione S‑transferase (GST) pull‑down, and laser confocal microscopy. Mechanistically, the central region of p17 (amino acids 19–61) directly binds to the GTPase domain of GNL2. Notably, ARV infection or ectopic expression of p17 significantly upregulates GNL2 expression. Functional assays revealed that GNL2 overexpression markedly promotes ARV replication at both RNA and protein levels, whereas GNL2 knockdown remarkably restricts viral proliferation. Furthermore, GNL2 enhances p17‑mediated cellular autophagosome formation, thereby constructing a favorable intracellular milieu for efficient ARV propagation. Collectively, these findings demonstrate that GNL2 serves as a key host dependency factor hijacked by ARV p17 to remodel host cellular functions. This study provides new insights into the molecular pathogenesis of ARV and offers potential targets for developing antiviral strategies against ARV infection.

In 2025, an infectious bronchitis virus (IBV) strain of the GVIII genotype was identified and isolated from a poultry farm in Hebei Province, China, representing the first documented detection of this IBV genotype within the country to date. Nucleotide homology analysis revealed that the S1 gene of this isolate shares 62.2%-96.9% sequence identity with previously reported GVIII-type strains from other countries. Comprehensive phylogenetic analysis of the complete genome indicated that this strain originated from a single recombination event: Its S gene may have been derived from the GVIII-1 lineage strain IBV/Ck/USA/CA/21-1883, while the remaining genomic regions were likely acquired from the GI-19 lineage strain SD. Pathogenicity evaluation demonstrated that infected flocks exhibited reduced body weight gain, with 40% of chickens showing signs of diarrhea. Necropsy findings included hemorrhagic lesions at the proventriculus-gizzard junction and within the bursa of Fabricius. Moreover, 85% of infected hens developed severe oviduct hypoplasia, and 90% showed a reduction in follicular number. The isolate described in this study constitutes the first detection of a GVIII genotype IBV strain in China, although its route of introduction remains undetermined. The emergence of this strain highlights the necessity for sustained molecular surveillance of IBV and the development of genotype-matched vaccines.

Avian infectious bronchitis virus (IBV) belongs to the genus Gammacoronavirus (family Coronaviridae), causes severe multi-system disease in chickens, inflicting major global economic losses. The molecular interplay between IBV and host metabolic networks remains poorly understood. Through integrated transcriptomic, metabolomic, and lipidomic profiling of oviduct tissues from specific-pathogen-free (SPF) chickens infected with the IBV QXL strain, we demonstrate tripartite metabolic reprogramming: (1) redirected glucose flux through the pentose phosphate pathway (PPP) to fuel nucleotide synthesis, (2) rewired lipid metabolism to prioritize de novo membrane biogenesis over fatty acid β-oxidation, and (3) orchestrated glycerophospholipid remodeling. This integrated analysis revealed a coordinated upregulation of fatty-acid biosynthesis genes and accumulation of specific glycerophospholipids and eicosanoids. Mechanistically, IBV co-opts the Warburg effect and PPP activation while uniquely suppressing fatty acid β-oxidation to channel fatty acids toward lipid droplets (LDs) biogenesis. Phosphatidylserine (PS) overproduction (e.g. 2.55-fold increase in PS(22:0/22:6)) and phospholipase A2 (PLA2)-mediated lysophospholipids (Lyso-PLs) and eicosanoids generation (e.g. 7.09-fold increase in prostaglandin E2 (PGE2)) emerged as critical regulators of membrane dynamics and inflammatory signaling. This process was centrally coordinated by the significant activation of peroxisome proliferator-activated receptor (PPAR) (e.g. 1.74-fold increase in ACSL1) and transforming growth factor-beta (TGF-β) (e.g. significant increase in p-SMAD2) signaling pathways, directly linking lipid remodeling to immunomodulation. Functionally, targeting acetyl-CoA carboxylase (ACC) or glucose-6-phosphate dehydrogenase (G6PD), alongside TGF-β pathway modulation, synergistically curtailed viral replication in vitro. Our findings delineate a critical PPAR-TGF-β cross-talk that governs lipid remodeling during infection and identify host metabolic nodes that are potentially targetable for antiviral intervention.

Infectious coryza (IC), caused by Avibacterium paragallinarum (Av. paragallinarum), imposes a substantial economic burden on global poultry production. This study established and standardized comparative challenge models employing infraorbital sinus injection and intranasal inoculation to evaluate the pathogenicity of Av. paragallinarum serovars A, B, and C in SPF chickens. Infraorbital sinus injection leads to rapid onset, stable incidence, and more severe disease progression. The clinical symptom scores at 1 days post-infection (dpi) were higher in the infraorbital sinus injection group compared to the intranasal inoculation group. Additionally, the clinical symptoms in the 2019/JS31 (serovar B) and 2020/JS80 (serovar C) groups were more severe than those in the 2019/HB64 (serovar A) group. The disease progression in the intranasal inoculation group was more prolonged, with slightly higher clinical symptom scores at 7 dpi compared to the infraorbital sinus injection group. The trends in gross lesions and histopathological lesions were consistent with the clinical symptoms. Bacterial shedding results indicated that the infraorbital sinus injection group exhibited higher bacterial shedding than the intranasal inoculation group, demonstrating a dose-dependent relationship. The 2020/JS80 (serovar C) group had slightly higher bacterial shedding than the 2019/HB64 (serovar A) and 2019/JS31 (serovar B) groups. In all groups, bacterial shedding peaked at 5 dpi. This study provides a basis for investigating the mechanisms of pathogenesis in IC and the disease mechanisms associated with different serovars.

Mycoplasma gallisepticum (MG) is a major pathogen of poultry that causes substantial economic losses to the global poultry industry. Vaccination with live attenuated vaccines is currently a core strategy for controlling MG infection in chickens. In China, the officially approved MG vaccine strains for poultry use include F-36, TS-11, and 6/85. Following vaccination, effective methods are required to evaluate vaccine strain colonization and differentiate wild-type from vaccine strain infections. To address this unmet need, we developed a set of rapid, specific duplex quantitative PCR (qPCR) methods based on MGB probes. This method can discriminate between wild-type MG and all three currently used major vaccine strains, and users can select the corresponding specific duplex combination according to the vaccine strain adopted in field production. First, we compared wholegenome sequences of wild-type MG and vaccine strains, and successfully identified strain-specific polymorphic sites. Next, we designed and optimized specific primers and MGB probes targeting these polymorphic sites. Using the developed primer-probe sets, we established three independent duplex qPCR assays, each of which enables simultaneous detection of one specific vaccine strain and wild-type MG. Validation experiments confirmed that all three assays have high specificity, sensitivity, and reproducibility. When applied to DNA extracted from clinical chicken choanal cleft swab samples, this method successfully differentiated between wild-type MG infection and vaccine strain exposure. In conclusion, this novel assay provides a valuable technical tool for MG surveillance, MG eradication programs, and vaccine efficacy monitoring.

Infectious bronchitis is a highly contagious acute disease in chickens caused by the infectious bronchitis virus (IBV). The GI-19 lineage of IBV has been shown to undergo continuous evolution, with increasing variance in antigenicity and pathogenicity in China. The aim of this study was to assess the efficacy of the only registered QX vaccine (i.e., QXL87) in China against diverse GI-19 clades. A total of 1503 S1 sequences from China were identified as belonging to the IBV GI-19 based on phylogenetic analysis, including sequences from GenBank and clinical samples sequenced by our team (Key Laboratory of Animal Infectious Diseases of Ministry of Agriculture, China) from 2018 to 2022. Within the GI-19 lineage, five clades were identified and QXL87 vaccine belonged to clade 3. Clade 2 and clade 4 were found to be prevalent from 2018 to 2022. One strain was randomly selected from each of the four clades distinct from the QXL87 vaccine strain to serve as challenge strains for testing the cross-protective efficacy of the QXL87 vaccine. The amino acid homology of the S1 protein between these strains and the QXL87 vaccine strain ranged from 94.6 to 95.0

Mycoplasma synoviae (MS) is a prevalent pathogen in poultry, causing substantial economic losses. Vaccination is a crucial strategy for controlling MS infections. In China, a temperature-sensitive live vaccine, Vaxsafe®MS (MS-H strain), was introduced to address the rising prevalence of MS since 2017. Accurate differentiation between the MS-H vaccine strain and field strains is essential for the effective application of the vaccine and the eradication of MS in poultry. In this study, a duplex TaqMan real-time quantitative PCR (qPCR) method was first developed based on a single nucleotide polymorphism (SNP) at position 367 of the obg gene. However, some reisolated strains of the MS-H vaccine strain may exhibit reverse mutations at this site, potentially leading to false-positive results field strains detection. To address this issue, a comprehensive comparison of 24 MS genomes was conducted, identifying 18 SNPs unique to the MS-H lineage. Subsequently, an (A/G) SNP at position 131 of the MSH_02330 gene was selected through sequencing. Based on this SNP, a novel duplex qPCR method was established, characterized by high sensitivity, specificity, and repeatability. This method eliminates the influence of obg revertant strains, enabling accurate differentiation of the MS-H vaccine strain from MS field strains. It can be applied to both pure cultures and clinical samples, serving as a valuable tool for evaluating the immunogenicity of the MS-H vaccine and advancing the prevention and control of MS infections.

Avian reovirus (ARV) typically induces viral arthritis or tenosynovitis in chickens. The ARV structural protein σB plays a crucial role in viral replication and regulates cellular signaling pathways through interactions with host proteins. In this study, we demonstrate that both ARV infection and ARV σB protein can activate intracellular Wnt signaling pathways and induce inflammatory responses based on qPCR and Western blot analyses in HD11 cells. Interestingly, ARV infection can inhibit β-catenin ubiquitination and upregulate its protein expression levels. To further investigate the mechanism of this phenomenon, the quantity of ARV replication and the expression of inflammation cytokine IL-1β were both significantly increased when overexpression of Wnt14 protein. Conversely, shRNA-mediated knockdown of endogenous Wnt14 expression substantially suppressed ARV replication and virus-induced inflammatory responses. Furthermore, the inflammatory response was concomitantly attenuated in parallel with the suppression of ARV replication in the condition of pretreated with Wnt inhibitor. Finally, the direct interaction of ARV σB and Wnt14 protein was confirmed by using immunoprecipitation, glutathione S-transferase (GST)-pulldown assay. We further observed the colocalization of σB and Wnt14 protein by laser scanning microscopy techniques. The cellular Wnt signaling pathway regulated by ARV may mediated through this direct interaction between σB protein and Wnt14. In summary, our research provides new insights into the functional role of σB protein as well as elucidating pathogenic mechanisms associated with ARV infection-particularly its relationship with inflammatory responses.

Infectious coryza (IC) is a respiratory disease in poultry caused by Avibacterium paragallinarum (Av. paragallinarum). The disease caused growth retardation in broilers and reduced egg production in laying hens, resulting in significant economic losses to the global chicken industry. The biofilm is an important virulence factor for many bacterial pathogens, yet there is a paucity of research on the biofilm of Av. paragallinarum. This study aimed to construct a random mutant library of Av. paragallinarum using the Tn5-Kan transposon to identify genes involved in biofilm formation. A total of approximately 3000 mutants were obtained, and 38 of them demonstrated a reduction in biofilm formation of 70-90 % by crystal violet staining. The transposon insertion sites were further determined by chromosome walking, and 17 functional genes related to biofilm formation were identified. According to the functional analysis of the mutated genes, 14 mutants with mutated genes associated with energy metabolism, cell membrane formation, gene transcription and translation, and material transmembrane transport were screened to further explore their biological characteristics and pathogenicity in vivo and in vitro. The results indicated that the growth performance, resistance to disinfectants, adhesion and invasion ability to DF-1 cells and pathogenicity of the 14 mutants were reduced. The 14 mutants displayed increased sensitivity to antibiotics but did not show significant changes in hemagglutination titer or antiserum bactericidal ability. It is noteworthy that the M-76 mutant exhibited a marked reduction in pathogenicity. Following challenge, the experimental chickens did not present any clinical symptoms or pathological changes for a period of seven days, and the respiratory tract bacterial shedding was also the lowest. This indicates that a deficiency in biofilm formation reduces the pathogenicity of Av. paragallinarum. This study will contribute to our understanding of the molecular mechanism of biofilm formation of Av. paragallinarum and further study the pathogenesis of Av. paragallinarum.

Avian encephalomyelitis virus (AEV), a picornavirus, primarily infects the central nervous system of 1 to 2-week-old young chickens but not pullets. When wild-type AEV undergoes serial passaging in chicken embryos, it becomes to be embryo-adapted and can cause avian encephalomyelitis in chickens of all ages following intracutaneous infection through parenteral routes. This study was conducted to explore whether an outbreak of AEV in 95-day-old chickens was linked to inadvertent embryo adaptation of the AEV vaccine and its association with vaccination method. In this study, an AEV strain AEV/JS202201 was isolated from the flocks of chickens that had been shortly after vaccinated with the AEV vaccine combined with the avian pox vaccine by the wing-web method. Whole-genome sequencing was performed on the isolated AEV/JS202201 and the immunized VACCINE X strain. The results showed that the length of AEV/JS202201 and VACCINE X strain was determined to be 7,032 bp and 7,034 bp, respectively (both excluding the poly A tail). Compared with VACCINE X strain, one mutation, T24A, were found at the VP4 in the isolated AEV/JS202201 strain. Multiple sequence alignment revealed that no other AEV strains exhibited this mutation. Animal regression experiment confirmed that AEV/JS202201 could infect layer pullets and caused typical pathological changes in brain tissue, with a higher morbidity rate (4/10) and more severe clinical symptoms in chickens immunized via the wing-web method compared to those immunized orally (2/10). In summary, this study found a potential virulence-related mutation in the VP4 protein of AEV and emphasized that the oral vaccine method is safer than the wing-web method.

Avibacterium paragallinarum (Av. paragallinarum), the causative agent of infectious coryza, is a significant pathogen responsible for substantial economic losses in the poultry industry. Current preventive strategies rely primarily on inactivated vaccines, which have limitations such as vaccine failure and limited cross-protection between serotypes. This study aimed to develop an attenuated strain of Av. paragallinarum as a potential live vaccine candidate. Using the Tn5-Kan transposon, we constructed a transposon mutant library and identified a mutant strain, designated 2019/HB64-40, which harbored a disrupted ksgA gene encoding a critical enzyme involved in ribosomal RNA methylation. Compared with the wild-type strain, the 2019/HB64-40 strain presented significantly reduced biofilm formation, lower hemagglutination titres, and impaired growth. Pathogenicity assessments in chickens demonstrated that the mutant strain displayed significantly attenuated virulence, characterized by fewer clinical symptoms and reduced bacterial shedding. Furthermore, following challenge, all unimmunized chickens presented severe clinical signs of infectious coryza at 2 dpi, with symptoms beginning to ameliorate by 5 dpi, culminating in a mean clinical sign score of 2.1. In contrast, only one chicken (1/10) in the immunized group displayed mild facial swelling and nasal discharge, with a mean clinical sign score of 0.1. The immunized group receiving the 2019/HB64-40 strain demonstrated 90% immunoprotection, highlighting the potential of this attenuated strain as a live vaccine candidate. While cross-serotype protection was not evaluated, the results suggest effective homologous protection and colonization capacity, underscoring its promising application in the prevention and treatment of infectious coryza.

Background Severe Fever with Thrombocytopenia Syndrome (SFTS) is an emerging infectious disease that poses a significant threat to public health, given its high fatality rate and potential for person-to-person transmission. Caused by the SFTS virus (SFTSV), a novel bunyavirus first identified in central and eastern China, SFTS has drawn increasing attention in recent years. Methods To gain an updated and in-depth insight into the epidemiological features of SFTS in eastern China, this study collected all confirmed SFTS cases in the year of 2024 from Yangzhou, Jiangsu Province. A total of 33 laboratory-confirmed SFTS cases were included in the analysis; after obtaining their complete viral genome sequences through sequencing, an epidemiological investigation was conducted. Additionally, a phylogenetic tree was constructed using MEGA-X software. Results Our findings revealed that the number of SFTS cases in Yangzhou showed an overall upward trend over the study period, with only a slight decline in the past three years. The laboratory confirmation rate stood at approximately 64.4%. Most SFTS cases were sporadic rather than clustered. Notably, the mortality rate exhibited a positive correlation with age-rising as patients’ age increased. Conclusions In summary, this study systematically analyzed laboratory-confirmed SFTS cases in Yangzhou. The results suggest that the diagnostic and reporting criteria for SFTS need further optimization and standardization. Such improvements would facilitate a more accurate understanding of the disease’s epidemiological characteristics and provide scientific evidence to support SFTS prevention and control efforts.

The QXL87 live attenuated vaccine strain for infectious bronchitis represents the first approved QX type (GI-19 lineage) vaccine in China. This strain was derived from the parental strain CK/CH/JS/2010/12 through continuous passage in SPF chicken embryos. To elucidate the molecular mechanism behind its attenuation, whole-genome sequencing was conducted on both the parental and attenuated strains. Analysis revealed 145 nucleotide mutations in the attenuated strain, leading to 48 amino acid mutations in various proteins, including Nsp2 (26), Nsp3 (14), Nsp4 (1), S (4), 3a (1), E (1), and N (1). Additionally, a frameshift mutation caused by a single base insertion in the ORFX resulted in a six-amino-acid extension. Subsequent comparison of post-translational modification sites, protein structure, and protein–protein binding sites between the parental and attenuated strains identified three potential virulence genes: Nsp2, Nsp3, and S. The amino acid mutations in these proteins not only altered their conformation but also affected the distribution of post-translational modification sites and protein–protein interaction sites. Furthermore, three potential functional mutation sites—P106S, A352T, and L472F, all located in the Nsp2 protein—were identified through PROVEAN, PolyPhen, and I-Mutant. Overall, our findings suggest that Nsp2, Nsp3, and S proteins may play a role in modulating IBV pathogenicity, with a particular focus on the significance of the Nsp2 protein. This study contributes to our understanding of the molecular mechanisms underlying IBV attenuation and holds promise for the development of safer live attenuated IBV vaccines using reverse genetic approaches.

Infectious bronchitis (IB) is a highly contagious acute viral disease that leads to substantial economic losses in the poultry industry. Previous research conducted in our laboratory has indicated that Nsp2 may serve as a key virulence factor within the IBV genome, as evidenced by its pronounced divergence between the field strain and its attenuated counterpart. Understanding the interaction between Nsp2 and host proteins is crucial to elucidating the role of the Nsp2 protein in the pathogenesis and proliferation of IBV. Currently, much remains to be uncovered regarding the host proteins that interact with the IBV Nsp2 protein. In this study, 10 host proteins, including COX1, COX3, NFIA, ITGA1, ATP1B1, ATP1B3, ABCB1, ISCA1, DNAJA1, and IREB2, were screened to interact with IBV Nsp2 through yeast two-hybrid experiments and molecular docking simulations. Furthermore, the interaction of Nsp2 with ATP1B3, DNAJA1, and ISCA1 proteins was further validated through co-immunoprecipitation and confocal experiments. The GO, KEGG, and PPI databases revealed that the host proteins interacting with Nsp2 are primarily associated with ATPase activation, Fe-S cluster binding, ion homeostasis, and innate immune regulation. The examination of the expression levels of these Nsp2-interacting host proteins during IBV infection demonstrated the significant downregulation of COX3, COX1, ATP1B1, and ATP1B3, while NFIA, DNAJA1, and IREB2 showed significant upregulation. Moreover, our study identified that IBV enhances viral replication by upregulating DNAJA1 expression, although the underlying mechanism requires further investigation. These findings provide valuable insights into the potential role of the Nsp2 protein in the pathogenesis of IBV.

Avian reovirus (ARV), which commonly induces viral arthritis or tenosynovitis and immunosuppression in chickens, is associated with the nonstructural protein p17 that plays a crucial role in viral replication and regulates cellular signaling pathways through its interaction with cellular proteins. In our previous study, we identified the host protein IFN-γ-inducible protein-16 (IFI16) as an interacting partner of ARV p17 through yeast two-hybrid screening. In the current study, we further confirmed the interaction between IFI16 and p17 protein using coimmunoprecipitation, glutathione S-transferase (GST)-pulldown assay, and laser confocal microscopy techniques. Additionally, we found that the amino acid of p1761-119 is responsible for mediating the interaction with the HINa and HINb domains of IFI16. Interestingly, we observed a significant increase in IFI16 expression upon ARV infection or p17 protein exposure. Moreover, the replication of ARV was found to be largely influenced by the quantity of IFI16 protein. Overexpression of IFI16 led to a significant decrease in ARV replication, while knockdown of the IFI16 expression led to the contrary result. Additionally, our findings demonstrate that IFI16 plays a crucial role in the induction of inflammatory cytokines IFN-β and IL-1β during ARV infection as confirmed by qRT-PCR and ELISA analyses. In conclusion, our study provides novel insights into the functional role of p17 protein and the pathogenic mechanism underlying ARV infection, particularly its association with inflammatory response. Furthermore, it offers new perspectives for identifying potential therapeutic targets against ARV infection.

Avian reovirus (ARV) is highly prevalent in healthy poultry flocks and has been linked to viral arthritis/tendonitis, dwarf syndrome, chronic respiratory disease, and immunosuppression in avian species, resulting in significant economic losses within the poultry industry. The non-structural protein p17 encoded by ARV induces cellular autophagy and facilitates viral proliferation, playing a pivotal role in viral pathogenesis. To further elucidate the pathogenic mechanism basis of ARV p17 protein function, we employed a yeast two-hybrid system to identify Phosphoribosyl pyrophosphate synthetase 2 (PRPS2) as an interacting host protein with p17. In this study, we validated the interaction between PRPS2 and p17 using laser confocal microscopy, coimmunoprecipitation, and GST-Pulldown assays. Moreover, our findings demonstrate that the C-terminal region of PRPS2 is responsible for its binding to the p17 protein. Intriguingly, ARV infection significantly upregulated PRPS2 expression levels. Additionally, PRPS2 was shown to have a substantial impact on ARV replication; overexpression of PRPS2 increased ARV replication while knockdown of PRPS2 resulted in decreased quantities of ARV particles. Furthermore, our findings suggest that this process involves cellular apoptosis as a potential mechanism underlying these observations. Overall, this research provides valuable insights into elucidating the function of the p17 protein and sheds light on the pathogenic mechanism involving ARV-induced cellular apoptosis while offering novel perspectives for exploring therapeutic targets against ARV.

The NF-κB pathway is a critical signaling involved in the regulation of the inflammatory and innate immune responses. Previous studies have shown that Pseudorabies Virus (PRV), a porcine alpha herpesvirus, could lead to the phosphorylation and nucleus translocation of p65 while inhibiting the expression of NF-κB-dependent inflammatory cytokines, which indicated that there may be unknown mechanisms downstream of p65 that downregulate the activation of NF-κB signaling. Here, we found that PRV DNA polymerase factor UL42 inhibited TNFα-, LPS-, IKKα-, IKKβ-, and p65-mediated transactivation of NF-κB signaling, which demonstrated UL42 worked either at or downstream of p65. In addition, it was found that the DNA-binding activity of UL42 was required for inhibition of NF-κB signaling. Importantly, it was revealed that UL42 could induce the ubiquitination degradation of p65 by upregulating the suppressor of cytokine signaling 1 (SOCS1). Additionally, it was found that UL42 could promote the K6/K29-linked ubiquitination of p65. Finally, knockdown of SOCS1 attenuated the replication of PRV and led to a significant increase of the inflammatory cytokines. Taken together, our findings uncovered a novel mechanism that PRV-UL42 could upregulated SOCS1 to promote the ubiquitination degradation of p65 to prevent excessive inflammatory response during PRV infection.

Pseudorabies virus (PRV), an alpha herpesvirus, induces significant economic losses to the swine industry and infects multiple kinds of animals. Therefore, it is of great importance to explore anti-PRV compounds. In this study, to explore the anti-PRV compounds, a library of natural compounds was screened through a cell-based ELISA assay, and it was discovered that bufalin, a Na+/K+-ATPase inhibitor, had a robust inhibitory effect on PRV replication. A time-of-addition experiment and temperature-shift assay showed that bufalin significantly inhibited the entry stage of PRV. NaCl- or KCl-treatment showed that NaCl could enhance the inhibitory effect of bufalin on PRV replication, whereas there was no significant effect under the treatment of KCl. Meanwhile, it was also found that bufalin possessed antiviral activity against other alpha herpesviruses, including human herpes simplex virus type 1 (HSV-1) and chicken Marek's disease virus (MDV). Finally, it was found that bufalin could decrease the viral load in multiple tissues, and reduce the morbidity and mortality in PRV-challenged BALB/c mice. Overall, our findings demonstrated that bufalin has the potential to be developed as an anti-PRV compound.