Search Results (1,279)

This study reports, for the first time, changes in the microbiome community associated with anhydrobiosis in two tardigrade species of the genus Paramacrobiotus. To identify bacteria linked to the anhydrobiosis phenomenon and to track microbiome changes under anhydrobiotic stress, next-generation sequencing of bacterial 16S rRNA genes was conducted. Microbiome profiling was performed across various developmental and physiological stages of tardigrades, including: eggs; active adult specimens (both before and after 7, and 120 days of anhydrobiosis, referred to as short- and long-term anhydrobiosis, respectively); specimens in the desiccated tun stage; dead specimens following long-term anhydrobiosis (no dead specimens were observed after short-term anhydrobiosis); and the culture medium. It was shown that the microbiome community varied among stages, with high stage-specificity. Several bacterial genera were identified that may assist the host during anhydrobiosis, potentially through biofilm formation and by supporting stress-protective mechanisms such as heat shock protein expression and trehalose synthesis in eggs and tuns. These findings reveal that microbiota may contribute to anhydrobiotic survival in tardigrades, providing novel insights into host–microbe interactions under extreme environmental stress. Full article

Pancreatic ductal adenocarcinoma (PDAC) is the most complex and aggressive disease with the worst rates of unresectable or metastatic disease at diagnosis, resistance to systemic therapy, and case fatality rate (CFR) among leading cancers. In non-metastatic disease, neoadjuvant treatment with modern chemotherapeutic regimens followed by surgical resection and/or adjuvant mFOLFIRINOX has significantly improved oncological outcomes. However, recurrence rates remain alarmingly high, while immune checkpoint inhibitors (ICIs) or molecularly targeted therapy have not yet demonstrated clinical benefits. Comprehensive genomic profiling through NGS-based approved assays such as TruSight Oncology 500 (TSO500) could guide targeted therapy. Rapidly evolving mRNA cancer vaccines and circulating tumor DNA (ctDNA)-based prediction of minimal residual disease (MRD) and recurrence risk hold great promise towards the realization of rational combination therapy to improve recurrence-free survival (RFS) and overall survival (OS). More recently, single-cell multiomics (SC MO), spatial proteomics and transcriptomics (SPT), artificial intelligence (AI), and systems biology have revolutionized cancer research, enabling holistic tumor microenvironment (TME) analysis. In this comprehensive review, we describe the latest advances and unmet needs in the standard of care of PDAC. Moreover, we discuss the expectations of ongoing randomized clinical trials of adjuvant mRNA vaccine-based therapy and ctDNA MRD testing as prognostic biomarkers, towards personalized treatment to improve RFS and OS in a medium-term perspective. With a longer perspective, we explore how harnessing SC MO, SPT, AI, and systems biology can reveal the 3D spatial organization of interacting cancer, immune, and stromal cells. Multi-dimensional TME-, TSO500- and ctDNA-based framework of dynamic biomarkers are of paramount importance to achieve an optimal patient-specific perioperative multimodal treatment combining precision immunotherapy, targeted drugs, and modern chemotherapy, translated into future practice-changing clinical trials, that could eliminate MRD towards recurrence prevention. Full article

Background: Yes-associated protein 1 (YAP1), a key effector of the Hippo signaling pathway and mechanosensitive transcriptional coactivator, plays a complex role in osteoarthritis (OA) and cartilage regeneration. While YAP1 is essential for tissue homeostasis, its dysregulation has been implicated in both inflammatory and degenerative joint pathologies. However, its precise function remains ambiguous. Methods: We silenced YAP1 with small interfering RNA (siYAP1) in two human-cell-based models relevant to OA pathogenesis and cartilage repair: (1) IL-1β (10 ng/mL)-stimulated articular chondrocytes in monolayer and pellet cultures, and (2) TGF-β1 (10 ng/mL)-induced chondrogenesis in MSC pellet cultures. Outcome measures comprised YAP1 nuclear localization; inflammatory/catabolic markers in chondrocytes (IL6, IL8, ADAMTS5, MMP13); and, in MSC pellets, chondrogenic or hypertrophic markers (COL2A1, ACAN, RUNX2, MMP13, COL10A1) together with glycosaminoglycan (GAG) deposition. Statistical significance was assessed using an ANOVA or Friedman test with post hoc correction (Tukey or Dunn’s test, respectively); p < 0.05 was considered significant. Results: In human chondrocytes, siYAP1 reduced IL-1β-induced nuclear YAP1 localization and suppressed pro-inflammatory mediators IL6 and IL8, indicating an anti-inflammatory effect. YAP1 silencing also downregulated ADAMTS5 expression in 2D monolayers but not in 3D pellet cultures, suggesting reduced regulatory influence in the three-dimensional environment. Notably, MMP13 expression was paradoxically increased following YAP1 knockdown, underscoring the complexity of YAP1’s role in catabolic regulation. In MSC chondrogenesis, siYAP1 enhanced TGF-β1-induced chondrogenesis by increasing COL2A1 and ACAN expression and promoting GAG deposition on day 21. Additionally, it reduced hypertrophic markers RUNX2 and MMP13 on day 7, though COL10A1 remained elevated compared to negative siRNA, indicating only partial suppression of hypertrophic differentiation. Nuclear YAP1 levels were increased by day 21 despite reduced mRNA, suggesting post-transcriptional regulation or enhanced nuclear translocation. Conclusions: These findings demonstrate that YAP1 knockdown exerts context-specific anti-inflammatory and pro-chondrogenic effects while partially mitigating hypertrophy. However, divergent outcomes, namely elevated MMP13 in chondrocytes and upregulated COL10A1 in MSCs, indicate that YAP1 silencing does not uniformly suppress inflammation or hypertrophy. YAP1 represents a potential therapeutic target for OA, but its modulation requires careful consideration of cellular context, siRNA delivery method, and timing to optimize outcomes for cartilage repair and joint preservation. Full article

Background: Otitis media with effusion (OME) is a highly prevalent pediatric condition and a leading cause of conductive hearing loss in children. Its pathogenesis remains uncertain, and diagnostic and therapeutic challenges make management difficult. Objectives: This review evaluates current evidence on the middle ear microbiome in pediatric OME, focusing on the diagnostic value of 16S ribosomal ribonucleic acid (16S rRNA) gene sequencing and its potential clinical implications. Methods: A literature review was conducted using the PubMed database, including studies published between 2006 and 2026. Eligible studies involved pediatric patients with OME and examined the sources and characteristics of microbiota potentially involved in disease pathogenesis. Microbiome analysis was performed using next-generation sequencing (NGS) techniques. Results: Growing evidence indicates that OME is associated with microbial dysbiosis and biofilm formation rather than a sterile inflammatory process. The most frequently detected genera include Haemophilus, Moraxella, Streptococcus, and Alloiococcus, although substantial variability exists between studies. Pathogens are believed to reach the middle ear through the Eustachian tube from two main reservoirs: the nasopharynx and the adenoids. The potential role of Helicobacter pylori infection and gastroesophageal reflux disease (GERD) in OME pathogenesis remains uncertain and requires further investigation. NGS methods, including 16S rRNA sequencing, demonstrate higher sensitivity than conventional culture techniques, enabling the detection of fastidious and previously unrecognized microorganisms. Evidence also highlights the limited effectiveness of antibiotic therapy in OME, the persistent issue of antibiotic overuse, and the relative advantages of conservative management and microbiome-modulating approaches compared with antibiotics and surgical interventions. Conclusions: Current evidence suggests that OME is closely associated with microbiota dysbiosis and bacterial biofilm formation. Given the limited efficacy of antibiotics, microbiome-focused strategies—particularly probiotics—should be further explored. Molecular diagnostic methods, especially NGS, show clear advantages over traditional culture-based techniques. Future research should further evaluate microbiome modulation as a potential adjunctive or preventive strategy. Full article

Background: The molecular concordance between primary lung adenocarcinoma and metastatic lesions remains incompletely characterized despite its direct implications for precision oncology and biopsy-driven therapeutic decision-making. This prospective monocentric paired-sample study evaluated genomic concordance between primary lung adenocarcinoma and synchronous thoracic metastatic lesions using targeted next-generation sequencing (NGS). Methods: We identified 27 treatment-naïve patients with histologically confirmed lung adenocarcinoma who underwent paired molecular profiling of the primary tumor and a synchronous thoracic metastatic site (pleural or intrapulmonary). DNA and RNA were analyzed using validated institutional NGS platforms. Genomic alterations, including clinically actionable oncogenic drivers consistently covered by the sequencing panel used in each pair, were compared across matched samples. Concordance was assessed using exact binomial confidence intervals, Cohen’s κ statistics, McNemar tests, and paired Wilcoxon signed-rank tests. Results: Actionable driver alterations were identified in 17 of 27 patients (63.0%; 95% CI 42.4–80.6), including EGFR mutations (40.7%), KRAS alterations (18.5%), and one ALK gene rearrangement (3.7%). TP53 concurrent mutations were detected in 14 cases (51.9%). Across all 27 paired samples, driver-level concordance was 100% (95% CI 87.2–100), with perfect agreement for EGFR, KRAS, and ALK alterations (κ = 1.00). TP53 mutations showed high concordance (92.9%; κ = 0.85), while CNVs were concordant in 88.0% of evaluable pairs. Variant allele frequency (VAF) comparisons, adjusted for tumor cellularity, further supported the apparent clonal stability of driver alterations across paired samples. Conclusions: This study demonstrates very high molecular concordance between primary lung adenocarcinomas and their synchronous pleural or intrapulmonary metastases. The observed 100% concordance of actionable driver alterations across paired specimens supports the clinical reliability of thoracic metastatic biopsies for baseline molecular profiling in treatment-naïve disease. Although limited by sample size, these findings support the biological stability of actionable driver alterations during early thoracic metastatic dissemination. Full article

Lung cancer is a leading cause of global cancer mortality, with EGFR mutations serving as a primary therapeutic target. Although EGFR tyrosine kinase inhibitors (TKIs) are initially effective, acquired resistance inevitably develops. While ROS1 rearrangements are well-known baseline drivers, they are exceptionally rare as acquired resistance mechanisms. We utilized next-generation sequencing (NGS) to identify a rare ROS1 intragenic rearrangement (exons 35–37) in three never-smoking women with EGFR-mutant lung adenocarcinoma following progression on EGFR TKIs. Clinical courses were heterogeneous: one patient achieved a durable partial response using combined osimertinib and crizotinib. A second patient, intolerant to dual TKI therapy due to QTc prolongation and grade 3 edemas, achieved a sustained partial response with platinum-pemetrexed chemotherapy. The third patient exhibited polyclonal resistance, including EGFR C797S and TP53 mutations, with fatal central nervous system progression. In this three-patient case series, ROS1 exon 35–37 RNA-level intragenic rearrangements were repeatedly detected after EGFR-TKI progression, suggesting a rare transcript-level alteration within heterogeneous resistance evolution. However, its biological significance, driver versus passenger role, and therapeutic relevance remain uncertain. Combined EGFR and ROS1 inhibition may be considered in selected cases, but further validation is required. Full article

Hypertrophic scars are characterized by excessive collagen deposition, fibrotic remodeling, and functional impairment. However, the ability of current models is limited in recapitulating human pathology. This study presents a novel approach using induced pluripotent stem cell-derived scar organoids to model hypertrophic scar characteristics in vitro. Following established protocols, human pluripotent stem cells were differentiated into skin organoids and induced fibrotic transformation by treatment with TGF-β1 (10 ng/mL) and hypoxia (5% O₂) from day 45 onward. Scar organoids exhibited significant contraction and increased collagen I deposition compared with skin organoids. Immunofluorescence analysis showed reduced LHX2 expression, indicating loss of hair follicle development, while collagen I expression was significantly elevated. Dark-field imaging revealed marked morphological divergence between skin and scar organoids. RNA sequencing revealed distinct transcriptomic profiles. Expression of hair follicle-associated gene families (KRT and KRTAP) was upregulated in scar organoids, whereas epidermal structure-related genes (KRT4, KRT7, CLDN7, and WNT7) were downregulated. These findings demonstrate that iPSC-derived scar organoids successfully recapitulate key features of human hypertrophic scars, including excessive collagen production, loss of skin appendage development, and contractile behavior. This platform offers potential for future applications in drug screening, precision medicine, and understanding the molecular mechanisms underlying scar formation. Full article

Purpose: Prostate cancer is one of the most common malignancies in men, yet current prognostic methods remain suboptimal. Emerging evidence indicates that microRNAs (miRNAs) play critical roles in prostate cancer progression. This study aimed to identify miRNAs associated with adverse clinical outcomes by comparing miRNA expression profiles between prostate tumors with unfavorable versus favorable prognostic features. Materials and Methods: High-throughput next-generation sequencing (NGS) was used to analyze miRNA expression in formalin-fixed, paraffin-embedded prostate cancer tissue samples. Patients were classified into favorable or unfavorable prognosis groups based on risk stratification scores, Gleason grade group, and biochemical recurrence. Differentially expressed miRNAs were identified using a fold-change threshold ≥2 and a false discovery rate (FDR) <0.05. Predicted target genes and pathway analyses were conducted to generate candidate regulatory hypotheses rather than confirm mechanistic relationships. Results: Several miRNAs were differentially expressed according to prognostic category. miR-206 was significantly downregulated in high-risk tumors compared with low-risk tumors. High-Gleason-grade tumors showed reduced expression of miR-7704 and miR-4454, while miR-25-3p and let-7f-5p were upregulated. In patients with early biochemical recurrence, miR-7704 and miR-10400-5p were downregulated relative to those with prolonged recurrence-free survival. Target prediction analysis identified CPEB3, HAND1, PTAR1, and SPRYD4 as shared candidate targets, with CPEB3 emerging as a prioritized candidate supported by consistency in external datasets rather than a confirmed molecular target. Conclusions: Distinct miRNA expression patterns correlate with prostate cancer aggressiveness and clinical outcomes. miR-206, miR-7704, miR-4454, miR-25-3p, and let-7f-5p represent candidate prognostic biomarkers. Their shared target CPEB3 should be interpreted as a prioritized candidate for future investigation. Given the very small sample size and the lack of qRT-PCR and functional validation, these findings should be considered preliminary and hypothesis-generating, requiring validation in larger independent cohorts and experimental studies. Full article

Tumour necrosis factor-alpha (TNF-α) disrupts bioenergetic homeostasis in skeletal muscle cells through the suppression of ion-dependent ATPase activities, mitochondrial depolarisation, and impairment of antioxidant defences. Carvacrol, a phenolic monoterpenoid constituent of thyme and oregano essential oil, has been shown to exert cytoprotective effects in TNF-α-challenged L6 rat myoblasts. The mechanistic basis of these effects, specifically the relationship between membrane-associated ATPase function, mitochondrial polarisation status, and transcriptional regulation of metabolic stress-response genes, has not been formally characterised. L6 rat myoblasts were exposed to TNF-α (10 ng/mL, 1 h), then treated with carvacrol (6.25 µg/mL, 24 h) in a post-inflammatory rescue paradigm. Cell viability (MTT), membrane integrity (LDH), ion-dependent ATPase activities (Na⁺/K⁺, Ca²⁺, Mg²⁺), antioxidant enzyme activities (catalase, SOD), mitochondrial membrane potential (Muse™ MitoPotential flow cytometry), and SIRT1/AMPK mRNA expression were quantified. TNF-α significantly suppressed Na⁺/K⁺, Ca²⁺, and Mg²⁺-dependent ATPase activities (all p < 0.001), consistent with impaired membrane-associated bioenergetic function. Post-TNF-α carvacrol treatment partially restored all three ATPase activities (p < 0.05) and reduced the proportion of mitochondrially depolarised cells from 31.65 ± 4.25% to 19.0 ± 2.6% (p < 0.05). LDH release, catalase activity, and SOD activity were also significantly modulated. At the transcriptional level, carvacrol increased SIRT1 mRNA by 1.6-fold and AMPK mRNA by 2.0-fold relative to TNF-α-treated cells. An integrative bioenergetic model is proposed in which carvacrol’s membrane-intercalating properties restore the phospholipid environment required for ATPase conformational cycling, attenuating the Ca²⁺ overload that drives mitochondrial permeability transition, and thereby partially preserving Δψm. Transcriptional upregulation of SIRT1 and AMPKα may represent an adaptive response to residual energetic stress. The mechanistic relationships among these endpoints and the causal contribution of SIRT1 and AMPK to observed bioenergetic changes require protein-level and pathway-specific experimental validation. Full article

Circadian desynchronization is increasingly linked to metabolic, immune, neurocognitive, and oncological disease, but integrated clinical phenotyping across endocrine, microbiome, metabolic, and neuroimaging domains remains limited. We conducted a prospective, single-centre, observational study in 179 symptomatic patients referred for chronic multisystem features consistent with circadian dysregulation and 107 practically healthy controls. Circadian melatonin status was assessed using fractionated 24 h urinary 6-sulfatoxymelatonin (aMT6s) and standardized dim-light plasma sampling at daytime (14:00–16:00) and nocturnal (02:00–04:00) windows. Microbiome composition was assessed by 16S rRNA sequencing, urolithin A by targeted metabolomics, and putative glymphatic/perivascular clearance by MRI-derived DTI-ALPS index, perivascular space scoring, and white-matter-hyperintensity (WMH) volumetry. Patients showed markedly reduced nocturnal melatonin output and loss of day–night contrast (night aMT6s 10.2 vs. 40.6 ng/mL; urinary aMT6s day/night ratio 0.81 vs. 0.14; plasma nocturnal melatonin 12.7 vs. 54.4 pg/mL; all p < 0.0001), accompanied by altered cortisol day–night patterning. Patients also showed reduced microbiome diversity, depletion of Gordonibacter and Ellagibacter, lower plasma urolithin A, higher WMH volume and perivascular space scores, and a lower DTI-ALPS index. Age distributions broadly overlapped in the individual-level dataset, and key biomarkers were not significantly correlated with chronological age within the patient cohort; however, this finding is interpreted as an exploratory absence of detectable age gradient within the symptomatic cohort, not as proof of biological age-independence. Overall, the data support a coherent cross-sectional association among blunted nocturnal melatonin rhythmicity, dysbiosis/urolithin depletion, and MRI markers compatible with impaired perivascular clearance. The MGM axis framework should be regarded as hypothesis-generating; causal direction, melatonin receptor involvement, and AQP4-related mechanisms require longitudinal and mechanistic validation. Full article

Background/Objectives: Genomic alterations play a central role in diagnosis, prognostication, and therapeutic planning for hematolymphoid malignancies. At our tertiary care center, frontline genomic testing relies on optical genome mapping, karyotyping, and fluorescence in situ hybridization, with targeted next-generation sequencing (NGS) performed externally. To provide more comprehensive genomic profiling, we evaluated two large-panel NGS platforms and subsequently performed a clinical validation of the selected assay. Methods: The Illumina PanHeme DNA panel and the SOPHiA Genetics Community Myeloid Solution were compared using 24 bone marrow aspirate specimens with previously characterized alterations, including single nucleotide variants (SNVs), insertions/deletions (indels), and copy number variants (CNVs). The selected panel underwent full analytical validation using 60 specimens. Results: Both panels demonstrated excellent concordance for SNVs and indels, with comparable analytical performance and workflow. CNV calling with SOPHiA was notably strong. Platform selection was influenced by practical considerations, including panel content and cost, leading to a preference for further evaluation of the Illumina assay. Clinical validation of the Illumina PanHeme DNA panel, along with a complementary RNA Exome panel, was subsequently performed. Sequence variant detection showed 100% concordance with orthogonal testing, while CNV detection was variable, reflecting known limitations of targeted NGS. The RNA panel detected all expected fusion transcripts. Conclusions: These findings demonstrate robust analytical performance of both evaluated DNA panels. Clinical validation of the Illumina PanHeme DNA and RNA Exome assays supports their use for comprehensive molecular profiling of hematologic malignancies. Full article

Severe traumatic brain injury (TBI) is a leading cause of mortality and long-term disability in polytrauma (PT) patients, and its clinical outcome remains difficult to predict due to clinical heterogeneity and secondary injury mechanisms. Current diagnostic and prognostic approaches based on clinical assessment and imaging are limited, particularly in PT where neurological evaluation is often impaired. This study aimed to compare plasma- and extracellular vesicle (EV)-associated microRNA (miRNA) signatures in patients with severe TBI and healthy controls to identify their potential as minimally invasive biomarkers and to improve understanding of molecular responses. For profiling circulating miRNAs, blood samples were collected at ≤3 h and at 48 h after admission. In the screening phase, plasma samples of n = 15 patients with severe isolated TBI (Abbreviated Injury Scale [AIS]Head ≥ 4, all other AIS ≤ 1) and n = 15 age- and sex-matched healthy controls were pooled (n = 5/pool) and subjected to next-generation sequencing (NGS). In the following validation phase, n = 25 severely injured trauma patients (Injury Severity Score [ISS] ≥ 16) were enrolled and stratified into PT without TBI (PT; AISHead = 0; n = 13) and isolated TBI (n = 12). Differentially expressed candidate miRNAs identified in the screening phase were validated in individual plasma and EV samples using reverse transcription droplet digital polymerase chain reaction (RT-ddPCR). Functional enrichment and pathway analyses were performed using miRNet. NGS identified more differentially expressed miRNAs in plasma (ER: 103; 48 h: 65) than in EVs (Emergency Room [ER]: 14; 48 h: 32). Functional enrichment analysis indicated associations with pathways related to cellular stress, senescence, growth factor signaling, transcriptional regulation, and apoptosis. In validation, 12 of 16 plasma and 10 of 15 EV-miRNAs were confirmed as differentially expressed in TBI patients; among these, three plasma and four EV miRNAs differed between TBI and PT. After adjustment, most plasma miRNAs were associated with injury severity rather than group status. EV miRNA profiles showed heterogeneous patterns, with miR-1469 associated with TBI group status in adjusted analysis, while miR-1237-5p was linked to injury severity and other EV miRNAs showed no consistent group-specific effects. Plasma miRNAs mainly correlated with systemic injury markers, whereas EV miR-1469 showed a moderate association with the Glasgow Coma Scale (GCS). Overall, circulating miRNA profiles after injury appear to be predominantly influenced by systemic trauma severity rather than TBI-specific effects. Plasma miRNAs mainly reflected general injury burden, whereas EV-associated miRNAs showed more heterogeneous patterns, with miR-1469 emerging as a candidate associated with TBI after adjustment for clinical covariates. These findings suggest that EV-derived miRNAs, particularly miR-1469, may provide more targeted signals related to brain injury and warrant further investigation. Full article

Odontogenic sinusitis (ODS) is a common cause of unilateral maxillary sinusitis arising from periapical lesions (PALs) or other dental sources. The infection is typically polymicrobial and dominated by anaerobic bacteria, which are often under detected by routine culture. Molecular approaches such as quantitative polymerase chain reaction (QPCR) and next-generation sequencing (NGS) may provide improved characterization of the microbial burden and community structure. This study aimed to compare culture-based methods, targeted quantitative PCR, and 16S rRNA sequencing in paired samples to characterize microbial composition of ODS and evaluate diagnostic performance. Paired sinus mucosal biopsy (SIN) and periapical lesion (PAL) samples were collected from 28 patients with clinically confirmed ODS. Bacterial detection was performed using conventional culture and targeted QPCR assays for ten clinically relevant taxa. In three randomly selected patients, paired samples were additionally analyzed by 16S rRNA gene amplicon sequencing. Microbial load, taxa richness, and similarity between the two anatomically connected sites were assessed using Wilcoxon signed-rank, McNemar, Jaccard distance, and Bray–Curtis dissimilarity analyses. Results: Culture showed low sensitivity, identifying a limited number of pathogens, primarily Staphylococcus aureus, Streptococcus anginosus, and Fusobacterium nucleatum, in a minority of samples. In contrast, QPCR demonstrated substantially higher detection rates, particularly in PAL samples. Porphyromonas gingivalis (96.8%), Fusobacterium spp. (100.0%), and the S. anginosus group (90.3%) were highly prevalent in PAL specimens, with overlapping but lower detection in SIN samples. PAL samples exhibited significantly higher bacterial loads and taxa richness than paired SIN samples (Wilcoxon p = 0.0004). 16S rRNA sequencing confirmed polymicrobial communities at both sites and identified additional taxa not included in the QPCR panel. Similarity analyses revealed pronounced interindividual variability, ranging from near-identical to highly divergent paired microbiota. Periapical lesions act as reservoirs of predominantly anaerobic bacteria that may seed the maxillary sinus in ODS. Although microbial overlap exists, sinus communities display lower burden and site-specific compositional shifts. Culture-based diagnostics underestimate ODS microbial complexity, whereas combined molecular approaches provide a more comprehensive and clinically informative assessment. Full article

Background/Objectives: The gut microbiota has been increasingly studied in the context of gastrointestinal, neuropsychiatric, and immune-mediated disorders. However, comparative data across these conditions within the same population remain limited. This observational study aimed to perform a comparative analysis of gut microbiota composition across individuals with irritable bowel syndrome (IBS), anxiety disorders (AG), and autoimmune diseases (AI) and healthy controls (HC) from a Romanian cohort, aiming to identify population-specific microbial patterns. Methods: This study included 59 Romanian adults categorized into IBS (n = 27), AG (n = 13), AI (n = 11), and HC (n = 8) groups. Gut microbiota profiles were analyzed using 16S rRNA gene sequencing (NGS), and fecal short-chain fatty acids were quantified by headspace gas chromatography–mass spectrometry. Results: At the phylum level, Actinobacteria showed significantly higher relative abundance in HC compared to both the AI (adjusted p = 0.028) and the AG (adjusted p = 0.023) groups, with the AG–HC difference confirmed by LinDA analysis (FDR-adjusted p = 0.049). At the genus level, Bifidobacterium spp. was significantly lower in the AG compared to HC (adjusted p = 0.036), while Bacteroides spp. showed higher abundance in the AG compared to AI (adjusted p = 0.013), a finding supported by Kruskal–Wallis analysis. Lactate production was significantly lower in both the AI (p = 0.014) and AG (p = 0.038) compared to HC. The Simpson index revealed significantly lower microbial evenness in the AG compared to IBS and AI (FDR-adjusted p = 0.042). Beta diversity analysis identified significant differences in community composition between groups (PERMANOVA: Pseudo-F = 2.395, R² = 0.116, p = 0.039), with the most distinct separation between AI and AG (FDR-adjusted p = 0.048). Spearman correlation analysis revealed strong positive associations between Bifidobacterium and lactate, Bacteroides and the acetate/propionate ratio, and Actinobacteria and lactate (all FDR-adjusted p < 0.001). Conclusions: The present study provides exploratory evidence of disease-associated microbial and functional patterns in a Romanian cohort, with reduced Actinobacteria in the anxiety group as the most robust observation, suggesting a potential role of the gut microbiota in gut–brain–immune interactions and contributing preliminary population-specific data from an underrepresented Eastern European population. Full article

Background/Objectives: Colorectal cancer (CRC) is a leading cause of cancer-related mortality worldwide. Emerging evidence highlights the role of the gut microbiota in the development and progression of CRC. Microbial dysbiosis is hypothesized to contribute to chronic inflammation through a variety of mechanisms, such as the production of free radicals, which induce mutagenesis and immune dysregulation in the host, ultimately leading to diseases such as cancer. Methods: Tumor tissue samples or healthy mucosa tissue were collected for bacterial DNA extraction. The V3–V4 region of the 16S rRNA gene was amplified and sequenced using the Illumina MiSeq platform. Bioinformatics analysis was performed with QIIME2, including quality control, DADA2 denoising, alpha and beta diversity calculation, and taxonomic classification using the SILVA database. Results: Differences in microbial composition were observed between groups. The healthy controls exhibited high relative abundances of beneficial genera such as Faecalibacterium, Bacteroides, and Asteroleplasma, whereas the patients with CRC showed enrichment of atypical genera including Novosphingobium, Bradyrhizobium, and Undibacterium. Alpha diversity was lower in the CRC group, and clear clustering by group was observed in the beta diversity analysis. LEfSe analysis identified potential bacterial biomarkers associated with CRC at both the species and genus levels. Conclusions: The findings of this study support the hypothesis that colorectal cancer is associated with distinct alterations in gut microbiota composition, such as an increase in the Novosphingobium genus and a decrease in the Bacteroides genus. An exploratory description of these microbial profiles may aid in the development of microbiome-based diagnostic and therapeutic strategies and contribute to current knowledge of the role of the gut microbiota in CRC in southern Peru. Full article