Gedgaudienė, Greta

The human gut microbiome comprises bacteria, viruses, and fungi, yet the fungal component (mycobiome) remains poorly characterized. Here, we investigated gut fungal composition and fungal–bacterial interactions in healthy monozygotic and dizygotic twins. Fungal communities showed substantially higher inter-individual variability than bacterial communities. Zygosity, age, and shared environment had no major influence on fungal abundance, similarity, or dominant genera. Candida was the most abundant genus (mean 5.2% in 161 individuals), followed by Geotrichum (3.7% in 132), whereas Saccharomyces was detected less frequently (0.8% in 92). Most bacterial genera were negatively correlated with Candida and Geotrichum, with stronger negative associations observed at higher bacterial abundances (up to rho = −0.6 for Alistipes). Network analysis revealed complex negative correlations among Bacteroides, Prevotella, and Candida. Overall, our findings reveal a highly variable gut mycobiome independent of host zygosity, pointing to a competitive bacterial-fungal interplay as a key regulator of fungal homeostasis in humans.

Late diagnosis and the lack of effective early detection techniques contribute to the poor prognosis of pancreatic ductal adenocarcinoma (PDAC). To address this challenge, we develop ¹H NMR-based metabolomics-AI platforms employing customized multilayer support vector machine (SVM), AutoGluon, and Tabular Foundation Model (TabPFN) frameworks. These platforms integrate serum metabolomic profiles-including small-molecule metabolites and lipoproteins-with clinical/biochemical parameters (age, CA19-9) and Activin A, derived from 902 participants (424 high-risk controls and 478 PDAC cases). Our TabPFN-based algorithm, PanMETAI, outperform state-of-the-art models. In the Taiwanese training and validation cohort, the model achieved an impressive AUC of 0.99 (95% CI: 0.98-0.99). Its robustness is further confirmed in a Lithuanian external validation cohort (n = 322), which yields an AUC of 0.93 (0.90-0.95). Notably, it identifies key signature patterns that improve early-stage (I/II) PDAC diagnosis and perform well with small sample sizes (n = 50). TabPFN-PanMETAI offers a rapid, accurate, and non-invasive tool for early PDAC detection, with strong potential for clinical application.

Introduction: Patient-derived organoid culture is a powerful and versa-tile system for investigating the molecular mechanisms underlying a wide range of gastrointestinal diseases, including inflammatory bowel disease(IBD) [1].In the context of IBD, intestinal organoids derived from patients retain key genetic, epigenetic, and disease-specific features, offering a unique plat-form to model individual responses to inflammatory triggers [2, 3].This approach also shows promise in predicting personalized responses to biologic and small-molecule therapies, advancing precision medicine strategies for IBD [4].Although intestinal organoids represent a major advancement in personalized ex vivo cellular modeling, the detailed effects of differentiation and inflammation on the cellular landscape remain poorly understood. Aims & Methods: In this study, we analyzed differentiated and undifferentiated intestinal epithelial organoids derived from both an IBD patient with active ulcerative colitis and a healthy donor (non-IBD) (n = 4). Inflammation was induced by treating organoids with the pro-inflammatory cytokines TNF-α (10 ng/mL) and IFN-γ (10 ng/mL). Organoids were dissociated using TrypLE Express Enzyme supplemented with Y27632, and viable cells were enriched by fluorescence-activated cell sorting (FACS) using7-AAD staining. Cell viability and concentration were assessed using the Countess 3 automated cell counter and Trypan Blue staining. Single-cell suspensions were then processed using the 10x Genomics GEM-X Universal 3’ Gene Expression application, and libraries were sequenced on the NovaSeq X Plus platform. Primary data processing was performed using Cell Ranger (10X Genomics), followed by downstream visualization in Loupe Browser (10X Genomics) and an in-house pipeline. Results: The mean number of recovered cells per sample was 1,957, with an average of 142,015 reads per cell and 27,127 genes detected across all samples. Single-cell transcriptome analysis revealed five distinct clusters in undifferentiated organoids (both treated and untreated), and six to seven clusters in differentiated organoids (treated and untreated). The majority of cells expressed the epithelial marker EPCAM, confirming their epithelial origin. Inflammatory stimulation led to a significant upregulation of TNF-α and IFN-γ gene expression (p < 0.05) in both differentiated and undifferentiated samples. Furthermore, differentiated organoids displayed a significant increase in specific epithelial subtypes, including Paneth cells, enteroendocrine cells, and proliferative cells (p < 0.05), compared to their undifferentiated counterparts. Conclusion: To conclude, single-cell transcriptomic profiling of patient-derived intestinal organoids revealed a dynamic and heterogeneous cellular landscape shaped by both differentiation and inflammatory signals. The observed upregulation of inflammatory markers underscores the organoid model’s utility in mimicking inflammatory responses. Additionally, the potential expansion of distinct epithelial subpopulations in differentiated conditions highlights the model’s ability to faithfully recapitulate the complexity of the intestinal epithelium.

Introduction: Epithelial barrier dysfunction is a hallmark of inflammatory bowel diseases (IBD), contributing to increased permeability, microbial translocation, and sustained mucosal inflammation1,2. Targeting barrier restoration has emerged as a promising therapeutic strategy3.Gallic acid, a dietary polyphenol with known anti-inflammatory and antioxidative properties, has shown potential in modulating mucosal re -sponses4.To investigate its impact on epithelial integrity under inflammatory stress, we utilized intestinal organoids as a physiologically relevant ex vivo model system. Aims & Methods: The aim of this study was to investigate the potential of gallic acid to restore intestinal epithelial barrier function under inflammatory conditions, using colonic epithelial organoids and integrating transcriptomic and permeability analyses. The study was approved by the Kaunas Regional Biomedical Research Ethics Committee (protocol no. BE-2-31). Inflamed differentiated colonic epithelial organoids were established from a patient with ulcerative colitis(UC) and non-IBD control individual by exogenous stimulation with TNF-αand IFN-γ (10 ng/mL each) for 24 hours. For gallic acid (GA) screening, or-ganoids were pre-treated with metabolite (100 µM) for 1 hour prior to inflammation induction and co-treated during the induction period. Subsequently, organoids were either processed for mRNA sequencing(1×75 bp, NextSeq 550, Illumina) or subjected to a FITC-Dextran (4kDa)assay. RNA-seq data was processed, analyzed, and visualized using nf-core/RNAseq pipeline and RStudio. Images for permeability assay analysis were acquired using an inverted fluorescence microscope (Zeiss AxioObserver 7), processed with ImageJ, and analyzed and visualized in RStudio. Results: Principal component analysis of transcriptomic profiles from both UC and non-IBD organoids revealed clear sample clustering driven by stimulation. Differential gene expression and Gene set enrichment(GSEA) analyses indicated a more pronounced cell response to both treatments in UC organoids compared to non-IBD organoids. Inflammation activated immune (CXCL9/10/11, CX3CL1), interferon (IFI44, SOCS1, NOS2),and defense genes (GBP4, GBP5, OAS2), while downregulating metabolic genes (FABP6, CFTR, HMGCS2) in both groups. Gene Ontology-based GSEA confirmed immune pathway enrichment and lipid metabolism suppression in inflamed cultures. In total, 3,148 and1,479 DEGs were identified in inflamed UC and non-IBD organoids, respectively. GA treatment modulated 2,144 and 384 genes (e.g., LYPD8, MKI67,MT-ATP6, CXCL14, CCL22) in these groups, suggesting potential to rebalance inflammation and metabolism. GA co-treatment enhanced epithelial integrity in UC organoids via upregulation of cell junction, peptide hormone processing, and immune-related pathways. In non-IBD organoids, GA promoted cell adhesion pathways, supporting epithelial cohesion. Functional permeability analysis confirmed GA’s protective role, as sup-ported by transcriptomic data. Mean percentages of FITC-Dextran-permeable organoids were 16.02% (untreated), 86.08% (inflamed), and 42.58% (inflamed + GA). A trend toward statistical significance (Wilcoxon p = 0.125) was observed in both untreated vs. inflamed and inflamed vs. inflamed +GA comparisons. Conclusion: In conclusion, gallic acid modulates inflammatory and metabolic pathways during colonic inflammation and enhances epithelial barrier integrity.

Fecal microbiota transplantation is an effective treatment method for recurrent Clostridioides difficile infection. Widely used enteric tube and colonoscopy methods demonstrate excellent efficacy and safety results. Recent data suggest that new fecal microbiota transplantation methods using oral capsules may provide a less invasive approach. In this study, we aimed to compare primary fecal microbiota transplantation efficacy as well as short- and long-term safety of two different administration routes: oral capsules and enteric tube.

Šio tyrimo tikslas buvo įvertinti uždegimo mediatorių ir mikroRNR molekulių tinkamumą kepenų ląstelių karcinomos neinvazinei diagnostikai, gydymo veiksmingumui bei ligos prognozei vertinti. Atlikta uždegimo mediatorių, tokių kaip IL-6, IL-8, IL-10, FABP2, LPS, VEGF, ir mikroRNR transkriptomo analizė. Tyrimo rezultatai patvirtino potencialią IL-6 ir IL-8 vertę kaip biožymenų, padedančių patikimai vertinti kepenų ląstelių karcinoma sergančių pacientų išgyvenamumo, kepenų funkcijos dekompensacijos ir gydymo veiksmingumo prognozę. Nustatyta, kad mikroRNR profiliai reikšmingai skiriasi, lyginant naujai diagnozuota liga sergančius pacientus ir sveikus asmenis. Tam tikros mikroRNR molekulės gali būti svarbios kaip biožymenys kepenų ląstelių diagnostikai, pacientų išgyvenamumui ir gydymo veiksmingumui vertinti. Išskirtina viena mikroRNR (hsa-miR-215-5p), kuri reikšmingai padeda prognozuoti pacientų išgyvenamumą ir yra susijusi su šios ligos gydymu.

Introduction: Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer, predominantly diagnosed in patients with chronic liver diseases. The absence of specific biomarkers and the typically asymptomatic progression of the disease lead to delayed diagnosis.Patients with liver disease, including HCC, exhibit alterations in the gut microbiome and increased bacterial permeability, factors that may influence disease progression and patient outcomes. In recent years, there has been a growing interest in exploring circulating microbiome signatures as potential indicators of liver disease stutus, providing a new avenue for understanding HCC.Aims & Methods: Aims: To identify the signatures of the circulating blood microbiome associated with hepatocellular carcinoma, response to treatment and disease prognosis.Methods: This study conducted a post-hoc analysis of the SORAMIC clinical trial utilizing peripheral blood plasma samples. For this analysis, blood plasma samples from a cohort of 71 patients with HCC were examined(25 treated with Sorafenib and 46 with SIRT/Sorafenib). Peripheral blood samples were collected at two time points: baseline (prior to the initiation of anti-tumor therapy) with all 71 patients included, and post-therapy (7 to 9 weeks after treatment) with 39 patients included. Additionally, peripheral blood samples were obtained from 52 control subjects, who were endoscopically diagnosed with colonic diverticulosis, to compare against the patient group. The composition of the circulating blood microbiome was assessed through 16S rRNA gene sequencing, targeting the V1-V2 hypervariable regions. Subsequent analyses focused on identifying associations with potential novel biomarkers for diagnosis, prognosis, and response to treatment.Results: Patients with HCC had significantly diminished β-diversity of the circulating blood microbiome as compared to controls. Significant differences were also observed in between microbial communities of these two cohorts based on Bray-Curtis dissimilarity index. Patients with HCC had significantly higher levels of Proteobacteria and Bacteroidetes and lower levels of Firmicutes and Campylobacterota. Differential abundance analysis revealed that families of Gordoniaceae, Cryomorphaceae, Burkholderiaceae, Comamonadaceae, Enterobacteriaceae were significantly enriched in patients with HCC. At the genus level, patients with HCC had higher levels of Gordonia, Fluviicola, Burholderia, Limnohabitans and Escherichia/Shigella and lower levels of Corynebacterium. HCC patients, who responded to treatment with SIRT/Sorafenib showed an increase in blood bacterial diversity, enrichment of families Burkholderiales incertae sedis,Arcobacteraceae, Peptostreptococcaceae, Moraxellaceae, genera Sphaero-tilus, Aliacrobacter, Rhodoluna, Giesbergeria, Microbacterium, Rombout-sia, Arcicella, Pseudorhodoferax and depletion in families Gordoniaceae,Cryomorphaceae and genera Cloacibacterium, Gordonia, Limnohabitans,Fluviicola after the treatment. Levels of Acinetobacter, Enhydrobacter and Micrococcus in the blood before the HCC treatment were associated with patient prognosis.Conclusion: Patients with HCC exhibit a unique circulating blood microbiome profile characterized by reduced bacterial diversity. Response to treatment is linked to an increase in α-diversity and changes in the circulating microbiome profile. Specific members of the circulating micro-biome show potential prognostic capabilities, suggesting their utility as biomarkers for predicting disease progression and treatment outcomes in HCC.

Objective: Gastric cancer (GC) is a leading cause of mortality globally, underscoring the urgent need for innovative non-invasive monitoring tools. Molecular profiling indicates that liquid biopsy assays could address many diagnostic and prognostic challenges associated with GC. Materials and Methods: Blood samples from 13 control subjects (CON), 13 atrophic gastritis (AG) patients, and 33 GC patients were collected, with multiple samples from GC patients taken at different time points. Total cfDNA was isolated and very deep targeted sequencing was performed (NovaSeq 6000, Illumina). Results: In the CON, 53.8% of subjects had detectable genetic variants in APC, ERBB2, FAT1, and MUC16 genes, mainly low (60%) or moderate (30%) impact. In the AG, 23.1% had detectable genetic variants in APC, ERBB4, FAT1, KMT2C, MUC16, PIK3CA, and TRRAP genes, most moderate (20.0%) or modifier (53.3%) impact. In the GC, 51.5% of patients had detectable genetic variants in APC, CDH1, EGFR, ERBB2, ERBB4, FAT1, FAT4, KMT2C, KRAS, MUC16, PIK3CA, PTEN, SPEN, TRRAP genes, with high (4.0%), moderate (37.4%), and modifier (58.6%) impact. Mutational cfDNA profiles overlapped by different proportions in GC group comparing the time points: before vs. after the operation – 50.0%, before the operation vs. control visit – 12.5%, after the operation vs. control visit – 12.0%, and all three time points – 12.5%. Detection of somatic plasma cfDNA variants was significantly associated with tumor size (48.3% vs. 82.1%, T1-T2 vs. T3-T4 respectively, p = 0.007). Conclusions: This study illuminates plasma cfDNA dynamics from health to malignancy and links cfDNA mutational profiles with diverse clinical characteristics.

Prophages can have major clinical implications through their ability to change pathogenic bacterial traits. There is limited understanding of the prophage role in ecological, evolutionary, adaptive processes and pathogenicity of Helicobacter pylori, a widespread bacterium causally associated with gastric cancer. Inferring the exact prophage genomic location and completeness requires complete genomes. The international Helicobacter pylori Genome Project (HpGP) dataset comprises 1011 H. pylori complete clinical genomes enriched with epigenetic data. We thoroughly evaluated the H. pylori prophage genomic content in the HpGP dataset. We investigated population evolutionary dynamics through phylogenetic and pangenome analyses. Additionally, we identified genome rearrangements and assessed the impact of prophage presence on bacterial gene disruption and methylome. We found that 29.5% (298) of the HpGP genomes contain prophages, of which only 32.2% (96) were complete, minimizing the burden of prophage carriage. The prevalence of H. pylori prophage sequences was variable by geography and ancestry, but not by disease status of the human host. Prophage insertion occasionally results in gene disruption that can change the global bacterial epigenome. Gene function prediction allowed the development of the first model for lysogenic-lytic cycle regulation in H. pylori. We have disclosed new prophage inactivation mechanisms that appear to occur by genome rearrangement, merger with other mobile elements, and pseudogene accumulation. Our analysis provides a comprehensive framework for H. pylori prophage biological and genomics, offering insights into lysogeny regulation and bacterial adaptation to prophages.

Colonic epithelial barrier dysfunction is one of the early events in ulcerative colitis (UC) and microRNAs (miRNAs) participate in its regulation. However, cell type-specific miRNome during UC is still unknown. Thus, we aimed to explore miRNA expression patterns in colon tissue and epithelial cells at active and quiescent UC.