Veterinary Research最新文献

Upon RNA virus infection, nuclear translocation of activated transcriptional factors via the RNA-sensing signal pathway is a key event in the interferon (IFN)-mediated antiviral response, and a specific target of viral immune evasion. Foot-and-mouth disease virus (FMDV) causes an acute vesicular disease in cloven-hoofed animals and poses a serious economic risk to the dairy industry. FMDV VP4, one of the structural proteins, is an internal protein of the viral capsid and is known to play an important role in cell entry. Here, we demonstrate a novel molecular mechanism by which VP4 inhibits karyopherin (KPNA)-mediated antiviral immune responses. VP4 and IRF3 specifically interacted with the nuclear localization signal (NLS) binding site on the KPNA4 molecule, and VP4 inhibited the interaction between KPNA4 and IRF3 via competitive binding with higher affinity. Thus, VP4 inhibited nuclear translocation of IRF3 without affecting dimerization and phosphorylation of IRF3. Consequently, VP4 significantly enhanced the replication of RNA and DNA viruses by suppressing IFN production through inhibition of the IRF3-mediated type I IFN signaling pathway. Taken together, these results suggest that VP4 negatively regulates host type I IFN signaling by inhibiting the nuclear translocation of IRF3 and provide a critical implication for better understanding the pathogenesis of FMDV.

The antemortem diagnosis of aspergillosis in birds remains a complex challenge. A variety of diagnostic methods are currently available, including direct detection of Aspergillus components, antibody-based assays, and nonspecific markers such as serum protein electrophoresis (SPE), but their diagnostic performances remain limited. The objective of this study was to assess and compare the performances of several diagnostic approaches, specifically, galactomannan index measurement, beta-D-glucan assay, 3-hydroxybutyrate quantification, SPE, and mannoprotein detection tests. A secondary objective was to develop a predictive model of aspergillosis incorporating optimal test thresholds identified in the first phase, combined with clinical signs. A total of 101 serum and 31 plasma samples were collected from 118 aquatic bird of various species in France. Birds were classified into three categories: control individuals (n = 88), suspected aspergillosis cases (n = 7), and confirmed cases (n = 23). While individually considered tests displayed limitations in specificity, predictive modeling revealed that elevated levels of 3-hydroxybutyrate above 0.52 mmol/L (Se = 96%, Sp = 51%) combined with beta-globulins above 6.90 g/L (Se = 78%, Sp = 51%), in conjunction with concurrent respiratory clinical signs, were significantly associated with the occurrence of aspergillosis. A multivariable logistic model combining these variables achieved excellent diagnostic performance, with AUCs up to 0.98 and sensitivity above 98%. These findings suggest that these parameters, particularly when considered alongside clinical signs, may serve as more reliable indicators for detecting aspergillosis in birds.

Streptococcus suis is a zoonotic pathogen that poses a significant threat to both the swine industry and human health. This bacterium utilizes a type VII secretion system (T7SS) to translocate effector proteins that mediate bacterial competition and contribute to virulence. However, the functions of T7SS effectors in S. suis remain poorly understood. In this study, we identified and characterized LXG-T2, a T7SS-secreted toxin from S. suis virulent strain WUSS351. Bioinformatics analysis revealed that LXG-T2 harbors a C-terminal glycine zipper motif, a structural feature commonly associated with membrane-disrupting toxins. Functional assays demonstrated that LXG-T2 exhibits strong bactericidal activity against E. coli and provides S. suis with a competitive advantage. Furthermore, the LXG-T2 has the capacity to compromise the integrity of bacterial membranes, as evidenced by the observed increase in membrane permeability and depolarization in target cells. Moreover, LXG-T2 exhibited cytotoxic effects on host cells and promoted S. suis survival in a murine infection model. Collectively, our findings establish LXG-T2 as a T7SS effector mediated membrane disruption to enhance both bacterial competition and virulence. This work not only reveals a novel mechanism by which S. suis manipulates microbial communities, but also highlights the significance of T7SS effectors as key mediators of pathogenesis in Gram-positive bacteria.

Newcastle disease (ND), caused by virulent strains of the Newcastle disease virus (NDV), is a highly contagious disease that poses significant economic burdens on the global poultry industry. The DNA damage response (DDR) is a critical cellular mechanism that detects and repairs genomic damage to maintain cellular integrity. While viral infections are known to modulate DDR pathways to either inhibit or enhance viral replication, the interaction between NDV and host DDR remains largely underexplored. Here, we demonstrate that NDV infection induces significant DNA damage in DF-1 cells and activates DDR signaling, primarily via the ataxia-telangiectasia mutated (ATM) kinase pathway, in a manner dependent on active viral replication. Pharmacological inhibition of ATM kinase, but not ataxia telangiectasia and Rad3-related (ATR) kinase, significantly suppresses NDV replication, alleviates virus-induced G1-phase cell cycle arrest, and modulates the host immune response. Moreover, short interfering RNA (siRNA)-mediated knockdown of Chk2 markedly reduced viral M gene expression and progeny production, indicating that Chk2 is required for efficient NDV replication. These findings suggest that NDV exploits the ATM-Chk2 DDR pathway to establish a replication-favorable environment. Our study provides new insights into NDV pathogenesis and highlights potential targets for antiviral interventions.

Restrictions on antibiotics use have increased interest in the gut microbiota relationship to host health, particularly in enteric infections. The present field study, performed on two farms with endemic swine dysentery (SD) infection, characterises the faecal microbiota in 102 faecal samples from 13 diseased and 13 non-diseased pigs by shotgun metagenomic sequencing. The samples were collected during four samplings, which allowed us to monitor the animals before, during and after the clinical disease to investigate the role of the gut microbiota in disease outcome, assess the impact of infection on microbial composition and evaluate the microbiota evolution following recovery. Samples collected before disease demonstrated that SD susceptible pigs had lower microbial diversity, with significantly lower abundance of Treponema rectale, Prevotella spp. or Ruminiclostridium E compared with SD resistant pigs, which remained healthy. Marked alterations in microbial species composition and their functional profiles were evident during clinical disease. Brachyspira hyodysenteriae, Dysosmobacter sp. BX15, Acetivibrio ethanolgignens and Mucispirillum sp. 910586745 were significantly increased in abundance, which was associated with an increase of functions such as Bacteroides capsular polysaccharide transcription antitermination proteins or pterin carbinolamine dehydratase. No changes in the microbiota were observed after the disease when compared with non-diseased pigs, thus evidencing a restoration of the microbiota composition after therapeutic treatment and recovery. The study demonstrates that the microbiota may play a relevant role in SD disease outcome and evidences the changes that occur during clinical disease do not persist over time after pig therapeutic treatment.

Avian pathogenic Escherichia coli (APEC) is an extraintestinal pathogenic Escherichia coli that primarily causes avian colibacillosis, leading to localized or systemic infections. Its persistent survival within host macrophages is a critical component of the systemic infection process. In this process, outer membrane vesicles (OMVs) secreted by APEC play an important role; however, the interactions between OMVs and macrophages and their effects on bacterial intracellular survival remain unclear. This study demonstrated that APEC-derived OMVs induced intracellular Ca²⁺ release and reactive oxygen species (ROS) accumulation in macrophages, thereby activating all three branches of the protein kinase RNA-like ER kinase (PERK), inositol-requiring enzyme 1 (IRE1), and activating transcription factor 6 (ATF6) pathways of the unfolded protein response (UPR), leading to sustained endoplasmic reticulum stress (ERS). Additionally, OMVs disrupted the acidic environment of lysosomes, inhibiting autophagosome-lysosome fusion and leading to abnormal accumulation of autophagy marker proteins LC3-II and p62, resulting in ERS-mediated autophagy flux blockade (incomplete autophagy). Importantly, this ERS-dependent autophagy dysfunction significantly impaired the ability of macrophages to clear pathogens, thereby promoting intracellular proliferation and survival of APEC. Intervention with ERS inhibitors effectively alleviated ERS and restored autophagosome-lysosome fusion, significantly reducing APEC intracellular survival within macrophages (p < 0.01) and bacterial loads in chick tissues (trachea: p < 0.001, lungs: p < 0.001, liver: p < 0.05, spleen: p < 0.001). These results indicated that APEC triggers ERS in host macrophages by secreting OMVs, thereby causing autophagy flux blockade, escaping host immune clearance, achieving sustained intracellular survival, and ultimately leading to systemic infection. This study provides new insights into the role of OMVs in regulating the innate immune response of macrophages during APEC infection.

Porcine deltacoronavirus (PDCoV) is an enteropathogenic virus that causes severe diarrhea in pigs, particularly suckling piglets, and exhibits cross-species transmission with zoonotic potential. The S1 subunit of the viral spike (S) protein mediates cell entry and elicits neutralizing antibodies, making it an ideal target for diagnostics, prophylaxis, and therapeutics. Here, we produced two monoclonal antibodies (mAbs), B8F10 and G10C2, by immunizing BALB/c mice with a recombinant PDCoV S1 protein fused to a human IgG Fc fragment, expressed in an insect baculovirus system and purified using Protein A/G magnetic beads. Both mAbs can specifically recognize native PDCoV S protein in indirect immunofluorescence assays and western blot analyses under denaturing conditions, indicating their binding to linear epitopes. Isotyping classified both as IgG1/κ, and sequence analysis revealed distinct heavy-chain complementarity-determining regions (CDRs) but identical light-chain CDRs. Using truncated S1 proteins coupled with Pepscan ELISA, the B8F10 and G10C2 epitopes were precisely mapped to ³⁴²LETNFMCT³⁴⁹ and ⁴⁹¹VINNTVVG⁴⁹⁸, respectively. These epitopes can be recognized by both mAbs and swine PDCoV antiserum, confirming their immunodominance. While global PDCoV strain alignments revealed high conservation of these epitopes, a V491A mutation within the G10C2-binding site abolished mAb binding. Structural analysis confirmed both epitopes are surface-exposed on S1. These findings demonstrate the diagnostic potential of these mAbs and the epitopes' suitability as vaccine targets. Their high conservation suggests broad applicability, whereas the V491A mutation may represent an immune escape mechanism. These epitopes could serve as diagnostic markers, immunotherapeutics, or the foundation for epitope-based vaccines to induce protective immunity.

Autogenous vaccines are a critical tool in aquaculture for managing bacterial diseases when commercial vaccines are unavailable or ineffective. To improve vaccine efficacy, this study explored how different cultivation conditions influence virulence gene expression in two major fish pathogens, Aeromonas salmonicida subsp. salmonicida and Aeromonas hydrophila. Isolates were cultured in nutrient-rich (tryptic soy broth [TSB]) and nutrient-limited (Mueller-Hinton broth [MH]) media, with and without supplementation of 1% fetal bovine serum (tryptic soy broth supplemented with 1% FBS [TSB1] and Müller-Hinton supplemented with 1% FBS [MH1]), to mimic environmental and host-like conditions. Total RNA was sequenced using the Oxford Nanopore MinION platform, and gene expression was quantified using featureCounts and Salmon, followed by differential expression analysis with DESeq2. Results revealed that culture conditions significantly shaped transcriptomic profiles. TSB1 promoted the highest and most consistent expression of classical virulence genes such as aerA, exeC, and fliP, due to serum-derived host signals. In contrast, MH induced higher expression of genes linked to motility and early host interaction, including flpI and exeB, despite overall lower transcriptional activity. These findings highlight the complementary expression of virulence factors under distinct nutritional conditions. Heatmaps and principal component analysis (PCA) confirmed clustering of expression profiles across media types. In relation to our findings, TSB1 is therefore recommended as the primary medium for bacterin production in autogenous vaccine development. However, combining cultures grown in both TSB1 and MH may capture a broader antigen repertoire, enhancing immune recognition and protection. This transcriptomics-based strategy presents as a rational framework for designing next-generation autogenous vaccines in aquatic veterinary medicine.

Cathelicidin CATH-2 has been reported to exert potent anti-inflammatory activity in different species though neutralizing stimuli such as lipopolysaccharide (LPS) and lipoteichoic acid (LTA). CATH-2 has been shown to inhibit Streptococcus suis (S. suis)-induced activation of dendritic cells and macrophages by binding to LTA. However, the exact mechanism of this prophylactically anti-inflammatory activity remains unclear. Therefore, we investigated the anti-inflammatory activity and mechanism of CATH-2 in mice peritoneal macrophages pretreated with CATH-2 followed by S. suis infection. The results showed that CATH-2 pretreatment significantly reduced S. suis-induced transcription and secretion of interleukin (IL)-1β, IL-6, and IL-12. CATH-2 also downregulated NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) expression and apoptosis-associated speck-like protein containing a CARD (ASC) oligomerization, and inhibited the maturation of IL-1β, suggesting that CATH-2 inhibits NLRP3 activation. In addition, CATH-2 significantly inhibited S. suis-induced phosphorylation of p65 and extracellular signal-regulated kinase (ERK). Further study showed that CATH-2 inhibited S. suis-induced reactive oxygen species (ROS) by upregulating the expression of ROS scavenging genes including catalase (CAT) and superoxide dismutase 1 (SOD1). Mechanistically, transcriptome analysis revealed that CATH-2 regulated the protein kinase B (ATK)/mammalian target of rapamycin (mTOR) pathway, which was evident by the downregulation of phosphorylated (p)-ATK and p-mTOR induced by CATH-2. Notably, CATH-2 induced autophagy and autophagic flux. Inhibition of mTOR using rapamycin enhanced the CATH-2-induced autophagic efficacy, demonstrating that CATH-2 induces mTOR-dependent autophagy. However, inhibition of autophagy using 3-methyladenine (3-MA) reversed the reduction in the expression of p-p65, p-ERK, and IL-1β induced by CATH-2. Our study reveals that CATH-2 inhibits the nuclear factor kappa-B (NF-κB)/NLRP3-mediated inflammatory response through the induction of mTOR-dependent autophagy during S. suis infection, which provides new insight into the anti-inflammatory pathways of antimicrobial peptides.

Prion diseases are fatal and contagious brain disorders caused by a pathogenic prion protein (PrP^Sc) derived from the benign prion protein (PrP^C). To date, there are no therapeutic substances to completely block prion diseases. Thus, the development of a therapeutic substance is necessary, and the identification of a novel biomarker of prion disease is the first essential step to develop new drugs. In the present study, we carried out a metagenomic analysis to identify microbiome biomarkers for prion disease using next-generation sequencing and bioinformatics tools in intraperitoneally prion-infected mice. In addition, we evaluated the protective effects of epigallocatechin-3-gallate (EGCG), a potent microbiome changer, in prion-infected mice by western blotting and survival analysis. We found a total of 14 differentially abundant taxa between prion-infected and control mice. In addition, we found that prion diseases caused altered microbiome networks and upregulation of DNA repair-related pathways. Furthermore, we observed the protective effect of the microbiome changer EGCG against prion disease in prion-infected mice. Given previous reports of microbiome alterations in prion diseases, we further validated these associations and demonstrated the protective effects of a microbiome-modulating compound.