Search Results (417)

Epstein–Barr virus (EBV) contributes to human immunodeficiency virus (HIV)-associated diffuse large B-cell lymphoma (DLBCL) pathogenesis. We retrospectively analyzed clinical features and outcomes of EBV-positive (n = 32) and -negative (n = 71) cases. EBV status was determined using in situ hybridization. Immunological parameters, histological subtype, systemic B symptoms, plasma EBV DNA, and response to therapy were examined. Survival was compared using Kaplan–Meier analysis. Factors associated with overall survival (OS) and progression-free survival (PFS) in EBV-positive patients were examined using Cox regression models. EBV-positive cases showed a higher proportion of non-germinal center B cell subtypes and B symptoms, higher circulating EBV viral loads, and lower CD4⁺ T-cell counts at diagnosis than EBV-negative cases. OS did not differ significantly within the full cohort but was shorter in EBV-positive cases with high International Prognostic Index scores or CD4⁺ T-cell counts ≥ 50 cells/$\mathsf{\mu }$L. Concurrent infections and elevated plasma EBV DNA levels remained associated with unfavorable survival outcomes. EBV-positivity in HIV-associated DLBCL is related to more aggressive clinical and immunological features and independently predicts poorer survival outcomes in high-risk subgroups. Plasma EBV DNA may reliably indicate EBV status and serve as a prognostic biomarker. Integrating these features into clinical decision-making may enhance risk stratification and inform individualized treatments. Full article

Plant mitochondrial genomes are important for respiration, organelle evolution, and phylogenetic inference, yet remain poorly characterized in Clausena lansium, an economically important Rutaceae fruit tree. Here, we generated the complete mitogenome of C. lansium cultivar ‘Ji Xin’ by integrating Oxford Nanopore and Illumina sequencing data, followed by comparative analyses of repeats, plastid-derived mitochondrial sequences, collinearity, RNA editing, and phylogenetic placement. The mitogenome consisted of two circular molecules totaling 499,621 bp, and encoded 56 genes, including 33 protein-coding genes, 20 tRNA genes, and three rRNA genes. Repeat analysis identified 206 SSRs, 21 tandem repeats, and 35 dispersed repeats, with dispersed repeats strongly enriched on mtChr1. We detected 21 plastid-derived mitochondrial fragments totaling approximately 9.23 kb, indicating limited but detectable plastid-to-mitochondrion DNA transfer. Comparative analyses revealed fragmented synteny between C. lansium and related Sapindales mitogenomes, suggesting lineage-specific structural rearrangement. Based on leaf rRNA-depleted lncRNA sequencing data, 402 candidate C-to-U RNA editing sites were detected in 31 mitochondrial PCGs, most of which were nonsynonymous. Phylogenetic reconstruction placed C. lansium within Rutaceae, close to Limonia and Citrus. These results provide a mitochondrial genomic resource for C. lansium and contribute to understanding mitogenome evolution in Rutaceae. Full article

Non-tuberculous mycobacteria (NTM) are increasingly recognized as opportunistic pathogens, yet accurate species identification remains challenging. In this study, two clinical isolates, 19-857^T and 20-525, obtained from patients with suspected pulmonary tuberculosis in Beijing, were characterized using a polyphasic approach integrating phenotypic, biochemical, chemotaxonomic, and genomic analyses. Phylogenetic analysis based on 16S rRNA gene sequences placed both strains within the genus Mycobacterium, with highest similarity to M. peregrinum DSM 43271^T and M. arcueilense DSM 46715^T. However, whole-genome comparisons via average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) yielded values of 85.8–94.1% and 29.1–54.3%, respectively, against their closest relatives—well below the species delineation thresholds (95–96% for ANI; 70% for dDDH)—while the two isolates shared 98.8% ANI and 90.1% dDDH, confirming their co-assignment to a single novel species. Genomic analysis further revealed a repertoire of genes potentially associated with virulence, including glycopeptidolipid biosynthesis, mce operons, ESX secretion systems, and mycobactin synthesis, alongside antimicrobial resistance determinants against rifampicin, tetracyclines, and β-lactams. Mobile genetic element analysis also uncovered extensive insertion sequences and transposon-related elements with differential distribution between the two strains. Based on the combined evidence, we propose Mycobacterium shunyiense sp. nov., with strain 19-857^T (=CGMCC 38906^T) as the type strain. This discovery expands the understanding of NTM diversity and provides genomic reference data for accurate clinical identification. Full article

Ngueak Pla Mo is a Thai medicinal crude drug derived from the pharmacopoeial Acanthus species A. ebracteatus Vahl and A. ilicifolius L.; however, morphological similarity and the occurrence of non-pharmacopoeial substitutes such as A. montanus (Nees) T. Anderson may complicate authentication and quality control. This study aimed to develop an integrated molecular–chemical approach for differentiating pharmacopoeial Ngueak Pla Mo species from A. montanus. Twenty authenticated Acanthus samples were analyzed using ITS2 DNA barcoding, including nucleotide variation, pairwise genetic distance, barcode gap, haplotype network, phylogenetic analysis, and predicted ITS2 secondary structure. Representative samples were further examined by untargeted LC–QTOF/MS metabolite profiling combined with hierarchical clustering, PCA, PLS-DA, VIP analyses, fold-change evaluation, and specificity scoring. ITS2 analyses clearly separated A. montanus from A. ebracteatus and A. ilicifolius, supported by diagnostic sequence variation, positive barcode gaps, distinct haplotypes, phylogenetic clustering, and secondary-structure differences, whereas A. ebracteatus and A. ilicifolius could not be reliably differentiated from each other at the species level using ITS2. LC–QTOF/MS profiling revealed species-consistent metabolite patterns and prioritized gallic acid, catechol, 3,4-dihydroxybenzoic acid, trigonelline, and ursolic acid as preliminary candidate markers for A. montanus discrimination. These results indicate that combining ITS2 barcoding with metabolite profiling provides complementary evidence for Ngueak Pla Mo authentication and supports future quality-control development. Full article

Background/Objectives: Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies because of aggressive tumor biology, pervasive therapeutic resistance, rapid adaptive reprogramming, and a profoundly immunosuppressive tumor microenvironment. This review summarizes recent advances in KRAS-targeted therapies, DNA damage response (DDR)-directed strategies, immune-redirection platforms, and tumor microenvironment modulation. Methods: We conducted a narrative review of peer-reviewed publications, clinical trial reports, trial registries, and selected conference data addressing emerging therapeutic strategies for PDAC. Results: KRAS-targeted therapies, including KRAS G12D-selective and multi-selective RAS(ON) inhibitors, have demonstrated substantial preclinical and emerging clinical activity while revealing diverse mechanisms of adaptive resistance. DDR-directed approaches are expanding beyond BRCA-mutated disease toward functional homologous recombination deficiency, replication stress, and synthetic lethality. In parallel, bispecific antibodies, T-cell engagers, and CAR-T-cell therapies have generated preliminary evidence of antitumor activity, although stromal exclusion, antigen heterogeneity, and immune suppression remain major barriers. These advances indicate that therapeutic resistance in PDAC is a dynamic process involving interconnected oncogenic, genomic, stromal, metabolic, and immune mechanisms. Conclusions: Future progress will require biologically informed treatment frameworks integrating complementary therapeutic modalities according to baseline tumor biology and treatment-induced adaptive states, supported by longitudinal biomarkers and rational clinical trial design. Full article

The COVID-19 pandemic underscored the need to better characterize immune and molecular responses following SARS-CoV-2 infection and vaccination. Beyond antibody and cellular immunity, COVID-19 involves oxidative stress and DNA damage, affecting repair mechanisms and metabolic adaptation linked to immune resilience. Here, we present a multidimensional analysis of 20 individuals who recovered from mild COVID-19, integrating clinical features with humoral and cellular immune responses, T cell and myeloid phenotypes, oxidative stress, DNA damage, and metabolomic and lipidomic profiles. Although most individual parameters fell within physiological ranges, network modeling revealed structured associations spanning multiple biological domains. A central finding was a coherent cluster organized around vaccine dose number, linking anti-Spike antibody titers, oxidative stress, bioenergetic signatures, and granulocyte activation. Higher vaccination was associated with stronger humoral responses, lower oxidative stress, and a more balanced myeloid–metabolic profile, suggesting a potential protective role extending beyond antibody induction. Additional associations linked symptom patterns to T cell differentiation states, anti-nucleocapsid responses to systemic inflammation, and anaerobic signatures to DNA damage markers, revealing interconnections between immunometabolism, clinical expression, and genomic stress. Despite the small sample size, these findings offer a preliminary systems-level perspective on mild COVID-19 recovery and illustrate the value of integrative exploratory frameworks in infectious disease research, laying the groundwork for validation in larger longitudinal cohorts. Full article

Redox homeostasis is essential for intestinal and systemic health and is regulated by antioxidant defense systems and redox-sensitive signaling pathways such as the nuclear factor erythroid 2-related factor 2 (Nrf2) and the nuclear factor ‘kappa-light-chain-enhancer’ of activated B-cells (NF-κB). Disturbances in this balance promote oxidative stress, chronic inflammation, and disease progression. Increasing evidence indicates that microbiota-derived metabolites act as key modulators of redox biology by shaping host gene expression through receptor-mediated signaling, metabolic regulation, and chromatin-associated mechanisms, including histone modifications, DNA methylation, and changes in chromatin accessibility. This review discusses how major classes of microbiota-derived and microbiota-modulated metabolites, including short-chain fatty acids (SCFAs), secondary bile acids, tryptophan-derived metabolites, polyphenol metabolites, hydrogen sulfide, and lipid mediators, influence redox-sensitive signaling and epigenetic regulation. We highlight their effects on intestinal barrier integrity and immune cell function, with particular emphasis on macrophage polarization and T-cell differentiation. Finally, we consider the emerging translational relevance of the microbiota–metabolite–epigenetic axis, while emphasizing that biomarker development and therapeutic applications require further mechanistic validation and clinical studies. Full article

Alpha-enolase (ENO1) is a multifunctional protein frequently overexpressed in solid tumors, where elevated levels are associated with aggressive behavior and poor prognosis. Beyond its canonical glycolytic role, ENO1 participates in immunoregulatory processes through distinct subcellular pools. Intracellular ENO1 shapes tumor-associated metabolic programs, while surface-exposed ENO1 functions as a plasminogen receptor and can engage innate immune signaling pathways. Post-translational modifications—particularly citrullination and phosphorylation—generate structurally altered epitopes that expand ENO1 antigenicity and enable adaptive immune recognition, including coordinated humoral and T-cell responses in cancer patients. These determinants of ENO1 immunogenicity have downstream consequences within the tumor microenvironment: immune-accessible ENO1 modulates myeloid cell recruitment, dendritic cell maturation, and macrophage polarization, while ENO1-dependent metabolic and signaling programs contribute to immune suppression and escape through multiple interconnected axes. Together, these mechanisms position ENO1 at the interface between tumor metabolism and immune regulation. Preclinical evidence demonstrates that ENO1-directed strategies—including antibody-based targeting, DNA vaccination, and vaccines incorporating post-translationally modified ENO1 peptides—can generate productive antitumor immunity and synergize with checkpoint blockade, supporting the rationale for ENO1 as an immunotherapeutic target. This review synthesizes current evidence within an integrated framework linking ENO1 dysregulation to its immunological consequences in cancer and discusses translational implications for ENO1-centered immunotherapy and immunoprevention. Full article

Feline tritrichomonosis caused by Tritrichomonas foetus is an important cause of chronic or intermittent large-bowel diarrhea in cats. Nested PCR is useful for detecting low parasite burdens in fecal samples, but gel-based endpoint interpretation requires post-amplification handling and may increase the risk of amplicon contamination. This study describes a one-tube nested real-time PCR-HRM assay for qualitative detection of T. foetus DNA in feline fecal extracts. The assay was adapted from a previously described single-tube nested PCR workflow by integrating real-time fluorescence monitoring and high-resolution melting analysis as a closed-tube endpoint. Analytical evaluation included post-extraction matrix-matched sensitivity testing, an exclusivity panel, matrix blank assessment, a checkerboard carryover experiment, and within-platform repeatability assessment of HRM temperatures. Method agreement was assessed using 147 fecal samples from diarrheic cats and compared with gel-based single-tube nested PCR and outer-primer PCR. Both nested methods detected 16/147 samples, whereas outer-primer PCR detected 5/147 samples. All 16 nested-positive amplicons were confirmed by Sanger sequencing and showed 99.51–100.00% identity to T. foetus reference sequences. Across 95 target-associated dual-peak HRM reactions, the low-temperature melting domain showed a mean Tm of 76.18 ± 0.24 °C, and the high-temperature melting domain showed a mean Tm of 81.57 ± 0.42 °C. However, because no independent reference standard was available, diagnostic sensitivity and specificity were not determined. The assay should therefore be interpreted as a preliminary analytical evaluation and method-agreement study of a closed-tube alternative to gel-based nested PCR. Full article

Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of aggressive non-Hodgkin lymphoma and is characterized by pronounced molecular heterogeneity that is not always fully captured by standard histopathological assessment. Circulating tumor DNA (ctDNA) is increasingly regarded as a promising liquid-biopsy biomarker that enables non-invasive molecular tumor profiling, assessment of tumor burden, dynamic monitoring of treatment response, and detection of measurable/minimal residual disease (MRD). Modern analytical platforms, ranging from PCR-based assays to next-generation sequencing approaches, including CAPP-Seq and PhasED-Seq, have substantially expanded the possibilities of molecular monitoring in DLBCL. This review summarizes current data on the biological characteristics of ctDNA, contemporary methods for its analysis, concordance between ctDNA and tumor-tissue mutational profiles, and the clinical significance of baseline ctDNA levels, early molecular response, post-treatment MRD status, and molecular surveillance during remission. Special attention is given to ctDNA monitoring in patients receiving novel immunotherapies, including CAR-T cell therapy, bispecific antibodies, and antibody–drug conjugates. Emerging multi-omic approaches integrating genomic, epigenomic, and fragmentomic data are discussed as promising future directions. Key limitations of clinical implementation include insufficient standardization of preanalytical and analytical workflows, the confounding effect of clonal hematopoiesis of indeterminate potential, variability across technological platforms, and the lack of completed prospective randomized interventional studies demonstrating improved outcomes when therapy is modified according to ctDNA status. Overall, ctDNA is currently a highly informative prognostic biomarker in DLBCL; however, its full implementation as a predictive tool for treatment selection requires further harmonization, prospective validation, and confirmation in interventional clinical trials. Full article

Molecular characterization of the transferred DNA (T-DNA) insertion sites is required for the safety assessment of genetically modified (GM) crops, yet conventional PCR-based methods are labor-intensive and limited in their ability to resolve complex structural variations. We present T-DNA Analyzer, an integrated bioinformatics pipeline that transforms long-read sequencing data (PacBio HiFi or Oxford Nanopore) into a comprehensive insertion site report. The pipeline implements a host-derived read filter that subtracts host-homologous vector regions to eliminate false-positive chimeric read calls; a multi-segment fusion detection algorithm that resolves complex T-DNA integration architectures; and a deletion gap gene impact analysis that identifies genes affected by host genome deletions at the integration site. Validation on maize and cotton datasets demonstrated that the host-derived filter excluded 86.4% of false-positive reads while retaining all true chimeric reads, and the fusion detection algorithm successfully reconstructed a two-copy tandem T-DNA repeat within a single long read. T-DNA Analyzer provides automated, reproducible molecular characterization designed to support regulatory molecular characterization and is freely available as open-source software. Full article

Background: Cigarette smoking promotes persistent systemic alterations in COPD, yet the interplay among genetic susceptibility, oxidative stress, immune dysregulation, impaired control of persistent viral replication, and long-term outcomes remains incompletely understood. Methods: We conducted an observational study in 102 patients aged ≥70 years with severe-to-very-severe COPD undergoing pulmonary rehabilitation. Current smokers (n = 38) were compared with never/former smokers (n = 64). Analyses included Chr15q25 genotyping (rs16969968), oxidative stress biomarkers (tail intensity, 8-OHdG, MDA, and bilirubin), hematological and immunological parameters, α7nAChR expression, TTV load as a surrogate marker of immune competence, latent virus prevalence, and five-year survival assessed by multivariable Cox regression. Results: Current smokers exhibited significantly higher DNA damage (tail intensity, p = 0.001; 8-OHdG, p = 0.002), lower bilirubin levels (p = 0.031), increased neutrophil and CD4⁺ T-cell counts (p = 0.031 and p = 0.028, respectively), altered α7nAChR expression on CD4⁺ T cells (p = 0.030), and higher TTV load (p = 0.002) than never/former smokers. The rs16969968 AA genotype was more frequent among current smokers. In survival analyses, an elevated WBC count was independently associated with increased mortality risk (HR 1.12, 95% CI 1.01–1.23; p = 0.035), whereas higher bilirubin levels showed a protective association. TTV load, smoking status, and FEV1 were not independently associated with mortality. Conclusions: In severe-to-very-severe COPD, smoking is associated with a distinct biological profile characterized by enhanced oxidative DNA damage, systemic inflammation, immune remodeling, reduced antioxidant defenses, and impaired control of persistent viral replication. WBC and bilirubin emerged as the biomarkers most consistently associated with long-term outcomes. These findings support integrated biological profiling as a tool for risk stratification and precision-guided rehabilitation in advanced COPD. Full article

Sepsis is a life-threatening syndrome characterized by a dysregulated host response to infection and progressive organ dysfunction. Although early antimicrobial therapy, source control, hemodynamic resuscitation, and organ support remain the foundations of care, these approaches do not directly reverse the cellular mechanisms that connect systemic inflammation to multi-organ failure. Mitochondrial dysfunction has emerged as a central mechanism linking impaired oxygen utilization, oxidative and nitrosative stress, immune-cell metabolic reprogramming, inflammatory amplification, and organ injury. During sepsis, inflammatory mediators, nitric oxide, microcirculatory abnormalities, calcium dysregulation, and metabolic stress converge on mitochondria, impairing oxidative phosphorylation and promoting mitochondrial reactive oxygen species/reactive nitrogen species (ROS/RNS) generation. When mitochondrial quality-control programs, including fission, fusion, mitophagy, and mitochondrial biogenesis, fail to restore network integrity, damaged mitochondria accumulate and become persistent sources of oxidative stress and danger signals. Mitochondrial damage-associated molecular patterns, particularly mitochondrial DNA, oxidized mitochondrial DNA, cardiolipin, ATP, and N-formyl peptides, activate innate immune pathways such as TLR9-MyD88-NF-kappaB, the NLRP3 inflammasome, and cGAS-STING signaling. In parallel, mitochondrial metabolism shapes macrophage activation, neutrophil function, T-cell competence, pyruvate-lactate handling through the pyruvate dehydrogenase complex, and the transition between hyperinflammation and immunosuppression. Clinical translation remains challenging because sepsis is biologically heterogeneous and mitochondrial dysfunction is dynamic, tissue-specific, and influenced by disease stage. This review synthesizes current knowledge on mitochondrial dysfunction in sepsis, emphasizing oxidative and nitrosative stress, mitochondrial quality control, mitochondrial damage-associated molecular pattern (DAMP) signaling, immunometabolism, organ-specific injury, candidate biomarkers, clinical translational strategies for mitochondria-targeted therapy, and future approaches based on multi-omics and artificial intelligence-assisted patient stratification. We argue that future therapeutic development should move beyond nonspecific antioxidant supplementation toward time-sensitive, phenotype-informed, and biomarker-guided mitochondrial medicine. Full article

Background: PURA-related neurodevelopmental disorder (PURA-NDD; OMIM #616158) is a rare autosomal dominant condition caused by pathogenic variants in the PURA gene encoding Purα, a multifunctional protein involved in DNA replication, transcriptional regulation, and RNA transport. Since its initial description in 2014, PURA-NDD has been increasingly recognized as a distinct clinical entity with early onset and a broad phenotypic spectrum. The clinical presentation is characterized primarily by neonatal hypotonia, global developmental delay, intellectual disability, feeding difficulties, and epilepsy, along with additional features such as respiratory insufficiency, movement disorders, hypersomnolence, and variable dysmorphic traits. Despite a relatively recognizable core phenotype, marked inter-individual variability often limits the ability to establish a definitive clinical diagnosis based on phenotype alone. This underscores the essential role of molecular genetic testing in the differential diagnosis of rare neurodevelopmental disorders. Patients and methods: We report a cohort of eight individuals (four males and four females) aged 17 months to 15.5 years with PURA-related neurodevelopmental disorder (PURA-NDD), evaluated using a genotype-first diagnostic strategy supported by comprehensive genomic testing, including next-generation sequencing (NGS) panels and whole-genome sequencing (WGS). Patients were referred for the evaluation of nonspecific neurodevelopmental features, including neonatal hypotonia, respiratory distress, and epilepsy, in the absence of a definitive clinical diagnosis. Results: Molecular analysis identified eight heterozygous variants in PURA, of which four (50%) were novel: c.311T>G p.(Met104Arg), c.406_407del p.(Gln136Glyfs64), c.515A>C p.(Gln172Pro), and c.885delinsGC p.(His296Profs21). The remaining variants included previously reported missense and frameshift changes associated with PURA-NDD, as well as one variant previously reported in ClinVar. Conclusions: Our findings not only confirm the core clinical features of PURA-related neurodevelopmental disorder but also contribute to a more comprehensive delineation of its phenotypic spectrum. The detailed characterization of our cohort broadens the range of recognized clinical manifestations and further highlights the marked phenotypic heterogeneity of PURA-NDD. In addition, the identification of both novel and previously reported pathogenic variants expands the mutational spectrum of PURA and underscores the importance of integrating clinical, molecular, and bioinformatic data for accurate variant interpretation. Although genotype–phenotype correlations remain incompletely understood, emerging evidence suggests potential associations between variant type or location and clinical severity, warranting further investigation in larger cohorts. The recognition of characteristic neonatal features may facilitate earlier diagnosis and implementation of supportive multidisciplinary management. Overall, this study illustrates how genomic technologies not only improve diagnostic yield in rare disorders but also refine disease definition, enhance the understanding of underlying pathogenic mechanisms, and support the development of more precise genotype–phenotype correlations. Further studies involving larger cohorts and long-term follow-up are needed to better define the full clinical and molecular spectrum of PURA-NDD. Full article

Rosmarinic acid (RA), a naturally occurring polyphenolic compound, has demonstrated promising anticancer activity; however, its combinatorial potential with conventional chemotherapeutic agents remains incompletely characterized. This study investigated the cytotoxic, pro-apoptotic, oxidative stress-associated, and cytokine-associated effects of RA alone and in combination with doxorubicin (DOX) in 4T1 murine breast cancer cells and HaCaT human keratinocyte cells as a non-cancerous control model. Cellular viability, apoptosis, cell cycle progression, oxidative stress, mitochondrial function, cytokine responses, and apoptosis-associated molecular alterations were evaluated using complementary cellular and molecular approaches. In addition, three-dimensional (3D) tumor spheroid experiments were performed to assess treatment responses under physiologically relevant tumor-like conditions. Results demonstrated that RA synergistically enhanced DOX-induced cytotoxicity in 4T1 cells while exhibiting comparatively lower toxicity toward HaCaT cells. Combination treatment significantly increased apoptotic cell death, mitochondrial depolarization, intracellular reactive oxygen species (ROS) accumulation, apoptotic DNA fragmentation, and G₂/M-phase accumulation. N-acetylcysteine (NAC)-mediated rescue experiments partially reversed ROS elevation and treatment-associated cytotoxicity in both monolayer and 3D spheroid models. Furthermore, the RA+DOX combination markedly disrupted spheroid integrity and reduced spheroid viability compared with monotherapies. Collectively, these findings indicate that RA enhances the anticancer activity of DOX and support further investigation of this combination strategy in breast cancer models. Full article