Pegiviruses are generally regarded as non-pathogenic viruses with controversial clinical significance. Here, we describe an avian pegivirus (partridge pegivirus, ParPgV) associated with field outbreaks of encephalitis in red-legged partridges (Alectoris rufa). Next-generation sequencing identified ParPgV in brain tissues, revealing two distinct avian-origin pegiviruses. Histopathology and electron microscopy revealed encephalitic lesions, neuronal degeneration, and viral particles within neurons. Field surveillance demonstrated widespread vertical transmission across multiple partridge flocks. Experimental inoculation of red-legged partridges, grey partridges, and specific-pathogen-free chickens demonstrated viral neurotropism and systemic distribution with differences in humoral immune response. Infected red-legged partridges developed cerebellar atrophy detectable by MRI. Detection of negative-strand RNA replication intermediates confirmed active viral replication across different experimental hosts, and RNAscope in situ hybridization and immunohistochemistry further confirmed viral RNA and antigen in neural and lymphoid tissues. Here, we show experimental evidence supporting an association between a pegivirus and encephalitis, and suggest underappreciated neuropathogenic potential.

Spatiotemporal pathogenesis and tissue tropism of a virulent goatpox virus Mukteshwar strain in the goats.

Won't you be my neighbor? Control of the immune response by stromal and immune cell microenvironments within the lymph node.

Production and characterization of novel murine monoclonal antibodies against programmed death-ligand 1 with potential diagnostic application in immunohistochemistry.

Central and peripheral immune tolerance depend on distinct transcriptional programs orchestrated by autoimmune regulator (AIRE) and FOXP3, respectively. AIRE promotes the expression of peripheral tissue antigens in medullary thymic epithelial cells for negative selection of autoreactive T cells, whereas FOXP3 enforces the immune-suppressive program of regulatory T cells. Although their immunological roles are well established, the molecular mechanisms by which AIRE and FOXP3 engage the genome and regulate transcription have long been unclear. Recent structural, biochemical and genomic work has revealed an unexpected shared principle: both FOXP3 and AIRE form homomultimers that function as chromatin organizers. Despite functioning in different immunological contexts and possessing distinct modes of genome interaction, both proteins leverage and reinforce pre-existing chromatin landscapes to coordinate broader gene expression programs. In this Review, we summarize recent advances and emerging mechanistic insights into FOXP3 and AIRE, focusing on their multimerization, interactions with repetitive DNA and enhancers and roles as architectural regulators that shape transcriptional programs essential for immune tolerance.

Objective: To explore the mechanism of vitamin K2 (VitK2) regulating matrix Gla protein (MGP) in inhibiting tumor growth in inflammation-associated colorectal cancer (CAC) model mice. Methods: Twenty-six C57BL/6 male mice, 6-8 weeks old and weighing 20-25 g, were divided into 4 groups according to the random number table method (each group received different treatments): normal group (treated with olive oil gavage and physiological saline intraperitoneal injection) (n=5), model group [treated with olive oil gavage, azoxymethane (AOM) intraperitoneal injection, and dextran sulfate sodium (DSS) solution drinking treatment)] (n=5), 30 mg group (treated with 30 mg·kg-1·d-1 of vitamin K2 gavage, AOM intraperitoneal injection, and DSS solution drinking treatment) (n=8), and 60 mg group (treated with 60 mg·kg-1·d-1 of vitamin K2 gavage, AOM intraperitoneal injection, and DSS solution drinking treatment) (n=8). The mice were sacrificed at the end of the 12th week, and the number and length diameter of colon tumors in each group were compared. The expression level of nuclear proliferation antigen (Ki-67) in colon tissues was assessed by immunohistochemistry (IHC), while the levels of MGP protein and Smad1/5 pathway-associated proteins were determined by Western blotting (WB). Additionally, the expression of MGP mRNA was quantified using real-time quantitative PCR (RT-qPCR). According to different treatment methods, colon cancer epithelial cell line SW480 cells were divided into control group [treated with dimethyl sulfoxide (DMSO)], 100 μmol/L group (treated with 100 μ mol/L VitK2), 200 μmol/L group (treated with 200 μ mol/L VitK2), 400 μmol/L group (treated with 400 μ mol/L VitK2), empty vector group (SW480 cells were transfected with empty plasmid) and MGP overexpressing group (SW480 cells were transfected with MGP overexpressing plasmid). After 48 hours of treatment, the expression of MGP protein, changes in the Smad1/5 pathway, and cell proliferation at different time points (24, 48, 72, and 96 hours) after treatment were detected. Results: All mice in the model group, 30 mg group, and 60 mg group developed colorectal tumors, with a tumorigenesis rate of 100% (17/17). There was no significant difference in the number of colon tumors between the 30 mg and 60 mg groups and the model group (both P>0.05), but the long diameter of tumors in the 30 mg or 60 mg groups was smaller than those in the model group (both P<0.05). There was no significant difference in tumor number or long diameter between the 30 mg and 60 mg groups (both P>0.05). The Ki-67 protein expression levels in the 30 mg and 60 mg groups were both lower than those in the model group (both P<0.05). Meanwhile, the MGP protein expression levels and pSmad1/5 protein expression levels in the 30 mg and 60 mg groups were both higher than those in the model group (all P<0.05). There was no statistically significant difference in MGP mRNA levels between the 30 mg and 60 mg groups and the model group (both P>0.05). Furthermore, there was no statistically significant difference in the expression levels of Ki-67 protein, MGP protein, pSmad1/5 protein, or MGP mRNA between the 30 mg and 60 mg groups (all P>0.05). In the cell experiments, the MGP and pSmad1/5 protein expression levels in the 100 μmol/L group showed no statistically significant difference compared to the control group (both P>0.05). The MGP and pSmad1/5 expression levels in the 200 μmol/L and 400 μmol/L groups were both higher than those in the control group (both P<0.001). After 72 and 96 hours, the cell proliferation capacity in the 200 μmol/L and 400 μmol/L groups was both lower than that in the control group (both P<0.001). The pSmad1/5 protein expression level in the MGP overexpression group was higher than that in the empty vector group (P<0.001). After 24, 48, 72, and 96 hours, the cell proliferation capacity in the MGP overexpression group was lower than that in the empty vector group (all P<0.001). Conclusion: VitK2 can inhibit the growth of CAC model mice by promoting MGP expression and activating Smad1/5 pathway.

Long coronavirus disease (COVID) (post-acute sequelae of severe acute respiratory syndrome coronavirus 2 infection) is characterized by persistent or new health issues weeks or months after acute COVID-19. Cardiac involvement, including myocarditis, is a notable complication that can occur even after an initially mild infection. To review current literature (2022-2025) on myocarditis in Long COVID, focusing on incidence, pathophysiological mechanisms, clinical presentation, diagnosis, management, prognosis, and future directions. Post-COVID myocarditis remains relatively uncommon but is significantly more frequent than pre-pandemic rates. Myocardial injury in Long COVID is hypothesized to result from multiple mechanisms: persistent viral antigen or RNA in cardiac tissue causing chronic immune activation, immune-mediated damage (including autoimmunity such as anti-heart antibodies), microvascular endothelial dysfunction with impaired perfusion, and maladaptive inflammatory responses. Patients may present with typical myocarditis symptoms (exertional chest pain, dyspnea, palpitations) or atypical features like exercise intolerance, orthostatic tachycardia, or unexplained fatigue. Management is supportive and guided by myocarditis severity: exercise restriction for 3-6 months in confirmed cases, guideline-directed heart failure therapy if ventricular dysfunction is present, anti-inflammatory or immunosuppressive therapy in specific scenarios (eg, corticosteroids in fulminant cases or overlapping multisystem inflammatory syndrome), and therapies targeting pericardial involvement (nonsteroidal anti-inflammatory drugs, colchicine) if present.

African buffalo are long-term carriers of foot-and-mouth disease virus (FMDV), particularly the South African Territories serotypes. Detecting low viral loads in carrier animals remains challenging. This study aims to detect FMDV antigen in formalin-fixed tissues of carrier buffalo using immunohistochemistry (IHC) to identify potential sites of viral persistence. Polyclonal antibodies targeting structural FMDV proteins were used to detect FMDV in formalin-fixed tissues from African buffalo (n = 15) culled in the Kruger National Park. The results were scored and compared with those from polymerase chain reaction (PCR). Viral antigen was consistently associated with the vasculature of the coronary band and interdigital skin as well as in the lungs. It was also associated with the germinal centres of the palatine tonsils. Detection outside these areas was rare, indicating a distinct tissue tropism favouring the vasculature in carrier animals. All buffalo tested IHC positive, including some PCR-negative cases, suggesting IHC may offer increased sensitivity when viral loads are low. The results revealed a novel association of viral antigen with vascular tissue and demonstrated the potential for superior sensitivity over PCR. Further research is needed to validate this assay, prove repeatability and clarify how carriers contribute to disease persistence and transmission.

Hypofibrinolysis is prevalent in obesity and type 2 diabetes mellitus (T2D) and may be involved in their pathophysiology. Metabolic bariatric surgery (MBS) markedly improves both obesity and T2D, but whether these improvements are related to fibrinolysis is not known. In this study, biomarkers of fibrinolysis were assessed before and after MBS in patients with and without T2D. A single center prospective cohort study with 2-year follow-up. The participants were adults with obesity with (n = 28) or without T2D (n = 33) undergoing MBS preceded by 2 weeks of a low-calorie diet (LCD). The plasma concentrations of plasminogen activator inhibitor 1 activity (PAI-1act), tissue plasminogen activator activity (tPAact) and antigen (tPAag), plasmin-antiplasmin complexes (PAP) and fibrinogen were determined at baseline, after LCD and at 6 weeks, 1 and 2 years after MBS. After LCD and prior to MBS, PAI-1act decreased and PAP increased significantly. Two years after MBS, PAI-1act decreased from baseline 29.8 (63) to 3.8 (121) while tPAact increased (0.08 (390) to 0.23 (150) IU), and PAP increased (544 (54) to 814 (60) ng/mL). tPAag and fibrinogen decreased (16.4 (29) to 9.7 (51) ng/mL and 4.15 (23) to 3.82 (20) g/L, respectively); p < 0.001 for all; (geometric means, except tPAact which is arithmetic mean, coefficient of variation (%CV)). There were no differences in fibrinolysis between groups at baseline or after surgery. Multivariate analysis showed that reduction in body weight and fat mass had the most important influence on fibrinolysis. Fibrinolysis is significantly improved two years after MBS, as also indicated earlier after the two-week LCD. The lack of differences between patients with vs. without T2D suggests that fat mass reduction rather than improvement of glucose control is most important for improved fibrinolysis after MBS. In summary, our results support hypofibrinolysis as a mechanism in the pathophysiology in obesity.

The elimination of obscuring pigments and (or) artefacts that interfere with accurate interpretation of immunohistochemistry results is crucial towards improving specificity of antibody binding, ensuring clear results that are reliable and reproducible while forestalling false-positive or false-negative results. Extensive deposits of hemozoin pigment in tissues from malaria infected subjects constitute a hinderance towards accurate assessment of tissue and cellular antigen conformations in immunohistochemistry. Consequently, the present study aimed to establish a rapid and effective hemozoin depigmentation method for formalin fixed paraffin embedded (FFPE) mouse malarial tissues subsequent to immunohistochemistry. FFPE spleens from malarial mice with severe infections (≥ 42%) due to Plasmodium berghei ANKA parasite were subjected to depigmentation employing three (3) methods. The first and second method involved tissue incubation in 1% and 10% aqueous ammonium hydroxide solutions respectively while the third method employed saturated ethanolic picric acid solution. Optimal hemozoin pigment removal without distortion of tissue architecture or loss of antigenic epitopes was achieved upon incubation in saturated ethanolic picric acid at room temperature (26 °C) for 15 min. In contrast, both solutions of ammonium hydroxide utilized demonstrated unsatisfactory pigment removal, tissue deterioration and frequent instances of dislodged tissue(s). Saturated ethanolic picric acid solution ensured rapid and efficient depigmentation of hemozoin from tissues of malaria infected mice while preserving tissue morphology and antigenicity. Saturated ethanolic picric acid solution thus represents an effective approach for the clearance of hemozoin pigment from malarial tissue sections without compromising tissue integrity or antigenicity in immunohistochemistry.

The DC1 conundrum: Do type I dendritic cells make or break allograft tolerance?

African swine fever (ASF) is a devastating viral disease that affects domestic pigs (Sus scrofa domesticus) and European wild boar (Sus scrofa scrofa). A previous study indicated that wild boar were more susceptible to ASF than pigs, with shorter incubation periods and earlier onset of the disease. This follow-up study aims to compare the pathogenesis and progression of the disease between the two subspecies during the early and late stages of infection following intranasal inoculation with the highly virulent genotype II strain ‘Armenia 2007’. Histopathological changes and viral antigen distribution by immunohistochemistry (IHC) were assessed over time following intranasal inoculation. Viral antigen and histological changes were detected earlier in wild boar than in pigs. In wild boar, the medial retropharyngeal (MRPLN) and submandibular lymph nodes were among the earliest sites of virus replication from 3 days post-infection (dpi), with widespread dissemination occurring by 5 dpi. In pigs, the viral antigen was first detected in the MRPLN at 5 dpi. At the humane endpoint, which occurred at 6 dpi in wild boar and 9 dpi in pigs, virus antigen and histopathological scores were lower in wild boar than in pigs, even though the appearance and increase of viral antigen in tissues, onset and development of lesions and humane endpoint occurred earlier in the former. The lower severity and lesser extent of lesions in wild boar suggest lower tolerance of tissue damage prior to reaching the humane endpoint, demonstrating their greater susceptibility to and lower resistance against ASFV.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13567-025-01701-x.

Hepatitis D virus (HDV) is a defective RNA virus requiring hepatitis B virus (HBV) for replication, and co-infection of them significantly accelerates liver inflammation, fibrosis, and hepatocarcinogenesis. Recent studies have reported that the prevalence of HDV infection is extremely high in certain regions, such as Mongolia, exceeding 50% among individuals infected with HBV. To clarify the potential presence of HDV infection in Japan, we retrospectively examined 95 surgically resected cases for HBV-related hepatocellular carcinoma (HCC) at Tokushima University Hospital between 2003 and 2023. Formalin-fixed, paraffin-embedded liver tissues were subjected to immunohistochemistry using an antibody against large and small delta antigens of HDV. Plasma samples from 12 patients were additionally tested by ELISA. As a result, none of the HBV-related HCC cases showed immunoreactivity for HDV antigens in either tumor or non-neoplastic tissues, and all available plasma samples were negative by ELISA. These findings suggest that HDV infection is either absent or extremely rare among patients with HBV-related HCC in this cohort. Our results align with prior seroprevalence estimates and provide histopathological evidence supporting minimal contribution of HDV to liver carcinogenesis in Japan, contrasting sharply with patterns in hyperendemic regions. J. Med. Invest. 73 : 52-54, February, 2026.

Domestic cat hepatitis B virus (DCHBV), shares similarities with human hepatitis B virus (HBV) which is associated with liver disease. We report the first case of cholangiocarcinoma in a 17-year-old spayed female cat infected with DCHBV and positive for feline immunodeficiency virus. The patient presented with vomiting, anorexia, and an elevated globulin level. Ultrasonography revealed multiple hypoechoic hepatic lesions, and histopathology confirmed cholangiocarcinoma. The tumor exhibited CK7 positivity and HepPar-1 negativity, confirming biliary origin. Quantitative PCR detected DCHBV in the spleen and ascitic fluid, while immunohistochemistry and RNA in situ hybridization revealed viral antigen and mRNA in both tumor and non-tumor liver. The presence of a viral antigen and mRNA in neoplastic tissue suggests a potential role for DCHBV in hepatobiliary carcinogenesis.

The B4GALNT2 gene has become an important target for xenotransplantation because its inactivation reduces the antigenicity of porcine tissues. The growing use of organ-source pig models has led to an increased demand for the rapid creation of these animals. However, the physiological role of this gene in pigs remains poorly understood. In 2015, after generating pigs lacking B4GALNT2 expression, researchers observed a third allele for this gene. Subsequently, another gene, described as B4GALNT2-like, was found in the porcine genome. We have collected data over four years since the production of our first line of xenotransplantation pigs lacking B4GALNT2 and B4GALNT2-like expression. In this study, we were able to show that pig cells expressing B4GALNT2-like also reacted to Dolichos biflorus agglutinin (DBA) lectin, which recognizes GalNAc epitopes. Additionally, we demonstrated that B4GALNT2/B4GALNT2-like knockout embryos were able to be carried to term in females with the same genotype. We hypothesized that the presence of both genes in the porcine genome might have occurred due to duplication, inversion, and reinsertion of part of the Phospho1-B4GALNT2 segment. Finally, the pig B4GALNT2-like gene showed greater similarity to the human, bovine, and murine B4GALNT2 genes than the original pig B4GALNT2. These data clarify the importance of targeting both B4GALNT2 and B4GALNT2-like for xenotransplantation studies and have improved our knowledge about their genomic structure and role in pig reproduction.

Rabies remains a critical global health concern, particularly in endemic regions where timely access to postexposure prophylaxis (PEP) is often limited. The effectiveness of PEP relies heavily on rabies immune globulin (RIG), yet plasma-derived products continue to face persistent issues of limited supply, variable potency, and high cost. These constraints have intensified the demand for recombinant monoclonal antibodies (mAbs) that provide consistent quality and scalable production. Here, we describe the development of a fully human mAb, H81L90, directed against the rabies virus glycoprotein (RABV-G). The antibody was isolated from a phage-display library constructed using peripheral blood mononuclear cells (PBMCs) from vaccinated donors. H81L90 exhibited strong, specific binding to native RABV-G with nanomolar affinity as determined by surface plasmon resonance (SPR) analysis. In cell-based neutralization assays, H81L90 efficiently blocked infection by the ERA-enhanced green fluorescent protein (EGFP) strain, achieving complete viral inhibition at low microgram concentrations. Protective efficacy was subsequently evaluated in a murine challenge model, where a single intramuscular injection of H81L90 conferred full survival when administered before or at the time of viral exposure and retained measurable activity at reduced doses postexposure. Histopathological assessment revealed substantially lower viral antigen in hippocampal tissue from treated animals, indicating suppression of early neuroinvasion. Collectively, these data establish H81L90 as a potent, fully human antibody with both preventive and postexposure prophylactic potential, supporting its continued development as a next-generation biologic to complement or replace current RIG formulations in rabies PEP.

Bridging discovery and therapy in precision medicine-Phage display as a catalyst for nanobody innovation: A review.

Whole-mount X-gal staining is a classical histochemical method for detecting β-galactosidase (LacZ) expression in fixed tissues, providing spatial resolution of gene activity in situ. In cancer research, LacZ serves as a versatile reporter for monitoring gene activation, tracing cell lineage in genetically engineered models, and assessing cellular responses to oncogenic signaling within the tumor microenvironment. Here, we present an optimized protocol for the in situ visualization of LacZ+ cells in the skin of Ptch1+/- SKH-1 mice-a genetically defined model of basal cell carcinoma (BCC) characterized by constitutive Hedgehog (Hh) pathway activation. In this model, LacZ expression faithfully reports Hh signaling and allows direct visualization of emerging and established BCC lesions. Importantly, our protocol preserves tissue integrity and antigenicity, enabling seamless integration with downstream immunostaining or multispectral immunofluorescence. When combined with immune markers-including those for regulatory T cells, cytotoxic T lymphocytes, myeloid subsets, and cytokine expression-this approach permits high-resolution spatial profiling of immune architecture in relation to LacZ+ tumor foci. This method is particularly suited for studying how oncogenic signaling pathways such as Hh shape the immune landscape during tumor initiation, progression, or therapeutic response. Overall, the protocol offers a versatile platform for coupling gene expression mapping with immune contexture analysis in preclinical models of skin cancer.

BackgroundLong-term antigen-specific data in PMN among Chinese populations remain limited. This study evaluated six target antigens and their clinical significance during extended follow-up.MethodsWe retrospectively analyzed 132 treatment-naïve PMN patients diagnosed by biopsy (2010–2018) and followed for a median of 62.9 months. Renal tissue expression of PLA2R, THSD7A, NELL-1, PCDH7, EXT1, and EXT2 was assessed by immunohistochemistry, and serum anti-PLA2R antibodies were measured by ELISA. Associations between antigen profiles and 5-year outcomes (remission, renal survival, malignancy) were evaluated.ResultsPLA2R was the predominant antigen (84.1%), followed by THSD7A (5.3%) and NELL-1 (0.76%); no PCDH7, EXT1, or EXT2 positivity was detected. PLA2R-negative patients were more often female (71.4% vs. 36.0%, P = 0.003), with better renal function and more frequent C1q deposition (38.1% vs. 13.5%, P = 0.016). Serum anti-PLA2R antibodies were detected in 55.3% of patients and strongly correlated with tissue PLA2R positivity (AUC = 0.851; optimal cutoff ≥17.47 RU/mL). Baseline antibody titers were not associated with remission (P = 0.573). During 5-years follow-up, 42.4% achieved CR, 36.4% PR, and 21.2% had NR, with an estimated 5-year renal survival rate of 81.95%. No malignancy events were observed among the seven THSD7A-positive patients or the single NELL-1–positive patient in this cohort. Statistical power for rare antigen subgroups was limited.ConclusionsThis >5-year Chinese PMN cohort provides the first comprehensive analysis of six target antigens. PLA2R remains predominant, while PLA2R-negative patients distinct immunopathologic features yet favorable long-term outcomes. A population-specific anti-PLA2R cutoff showed good diagnostic performance for predicting tissue antigen deposition. Rare antigens were infrequent and their malignancy associations require cautious interpretation. These findings provide long-term antigen-specific data supporting antigen-guided, population-adapted precision management of PMN.

Zika virus (ZIKV) is a neurotropic flavivirus linked to severe neurodevelopmental defects following prenatal exposure. While the mechanisms by which ZIKV spreads within the central nervous system remain incompletely understood, extracellular vesicles (EVs) have emerged as potential mediators of intercellular communication and viral dissemination. Here, we demonstrate that EVs derived from ZIKV-infected neural cells encapsulate full-length viral genomes capable of establishing productive in vivo infection, independent of free virions. Primary cortical neurons, astrocytes, and mouse brain microvascular endothelial cells (MBECs) from neonatal mice were infected with ZIKV at a low multiplicity of infection (MOI 0.1). EVs were isolated and treated with acid glycine buffer and RNAase to exclude residual virions or free RNA. RNA sequencing, RT-qPCR, and droplet digital PCR (dd-PCR) analyses revealed that EVs—particularly those derived from neurons and MBECs—encapsulated ZIKV RNA, including full-length viral genomes. These EVs were able to transfer viral RNA to A549 cells in vitro, and its intracranial injection into neonatal mice resulted in productive infection, confirmed by detection of ZIKV capsid protein, viral RNA, and viral antigen in brain tissue. Our findings demonstrate that EVs from ZIKV-infected neural cells can serve as vehicles for genome transfer and initiate infection, even in the absence of detectable virions. The persistence of EVs-packaged genomes post-viremia could explain clinical observations of prolonged ZIKV RNA within the nervous tissue or delayed transmission. Understanding this pathway provides new insights into ZIKV neuropathogenesis and opens potential avenues for therapeutic intervention, for example targeting EVs biogenesis or cargo sorting.