Milk is a complete food that plays a fundamental role in supporting bone health with high content of calcium and protein. Recent research highlights the significance of milk-derived extracellular vesicles (mEVs) with emerging roles in modulating physiological processes, including bone remodeling. This study reports a comparative analysis of mEVs isolated from raw milk of three ruminant species: bovine, goat, and sheep. Extracellular vesicles (EVs) are characterized for size distribution, particle concentration, and protein composition. Functional relevance of each type of mEVs is evaluated on cell proliferation and osteogenic differentiation of murine fibroblasts model. Results reveal significant interspecies differences in both the structural and biological characteristics of mEVs. EVs derived from each milk type exhibit distinct effects on MC3T3-E1 viability, supporting the idea that mEVs have a beneficial role in cell growth and proliferation and induce osteogenic differentiation. Additionally, the microbial DNA content carried within mEVs from each ruminant species harbors distinct and structured microbial DNA profiles. Goat mEVs are enriched in beneficial commensals, whereas sheep samples reflect consistent yet mixed communities, and bovine mEVs exhibit environmental signatures and greater variability. Overall, these findings highlight the species-specific potential of mEVs, their compositional diversity, and potential functions of the microbial cargo. Furthermore, this study highlights mEVs as functional food components and opens new avenues for developing targeted dietary strategies leveraging mEVs to support personalised medicine in bone health and prevent or manage conditions such as osteoporosis.

Over the past decade, much attention has been directed towards the gut microbiota (GM) as a contributing factor to a wide range of diseases, especially in the aging population. In this study, we analyzed the GM of elderly patients with fragility fractures (age range 71-90 years, n = 13), and patients with severe osteoarthritis (age range 65-87 years, n = 11) using 16S rRNA amplicon sequencing of fecal samples and compared these profiles with those of healthy individuals (age range 65-100 years, n = 20). The aim was to improve understanding of GM alterations in skeletal diseases and to inform the future development of personalized therapeutic strategies. Analysis of microbial composition revealed that patients with skeletal disorders showed a dysbiotic GM profile, characterized by enrichment of Peptostreptococcaceae and Butyricicoccaceae, and other potentially pathogenic taxa, alongside depletion of healthy-associated, short-chain fatty acids (SCFA)-producing bacteria. Moreover, analysis of gut metabolic modules identified multiple functional differences between patients' groups and healthy controls, particularly in pathways related to SCFA production, amino acid metabolism, and energy metabolism. Overall, these findings demonstrate both shared and disease-specific dysbiotic signatures in the GM of elderly patients with fragility fractures or severe osteoarthritis compared with healthy individuals. Once validated in larger cohorts and supported by mechanistic studies, these results may pave the way for the development of novel GM-based intervention strategies for the management of skeletal diseases.

Gut microbiota may contribute to the adiposity-associated disease risk, but human studies reported inconsistent associations of adiposity with gut microbiota composition. We examined associations of body mass index (BMI) with alpha diversity and relative microbial abundance at the phylum and genus taxonomic levels (based on 16S rRNA amplicon sequencing or metagenomics) among 7415 adults from eight European observational studies in a joint federated analysis of harmonized data using DataSHIELD. Higher BMI (per 5 kg/m2) was associated with lower alpha diversity (β: -0.05; 95% CI: -0.07, -0.03) and, on the phylum level, positively associated with Proteobacteria, but neither with Firmicutes nor Bacteroidetes nor their ratio, where high between-study heterogeneity was observed. On the genus level, BMI was inversely associated with the relative abundance of Faecalibacterium of the Firmicutes phylum (β: -0.11; 95% CI: -0.14, -0.07) but positively with the odds of detection of Dorea, Streptococcus, and Clostridium (all three Firmicutes) as well as Collinsella (Actinobacteria). This federated analysis of multiple studies found lower alpha diversity, alongside depleted Faecalibacterium, as well as higher odds of detection of Dorea, Streptococcus, Clostridium, and Collinsella with higher adiposity. By combining data from diverse study populations using harmonized data and statistical methods, our analysis partly overcomes sources of heterogeneity that may explain previously observed inconsistencies.

The relationship between host microbiota and infective endocarditis (IE) has been investigated by few studies. We compared the oral, gut, and skin microbiota profiles of patients awaiting heart valve replacement, both with and without IE, and correlated the profiles with the patients' clinical data, including the etiological agents and antibiotics received. Thirty-six patients were enrolled in a prospective, observational pilot study: 25 with IE (cases) and 11 with non-infective valve disease requiring surgery (controls). Clinical data and oral, fecal, and skin samples were collected before surgical treatment. Microbiota was profiled using 16S rRNA amplicon sequencing. Cases and controls were comparable, except for inflammatory parameters and prevalence of prosthetic valves, which were higher in cases. Causative IE pathogens were Staphylococcus spp., Streptococcus spp., and Enterococcus spp. Compared to controls, cases showed dysbiotic features in their oral, gut, and skin microbiota, including lower diversity in oral and skin communities and overall community depletion. Only a few bacteria were enriched, such as Oxalobacteraceae in the oral cavity and Bacteroides in the gut. There was low concordance between bacterial changes and blood isolates for most IE-causing pathogens, suggesting that dysbiosis may have effects beyond merely acting as a reservoir for specific pathogens. Dysbiosis may be a relevant yet underexplored factor associated with IE. Further studies are needed to clarify the underlying pathogenetic mechanisms.IMPORTANCEThe relationship between microbiota alterations and infective endocarditis has been explored in the literature, but the majority of available studies focus only on the oral microbiota. In this prospective observational pilot study, we analyzed three microbial ecosystems (oral, skin, and gut) in patients awaiting heart valve replacement with and without infective endocarditis. We observed dysbiotic features that are potentially associated with infective endocarditis. While these findings are preliminary, they suggest that future studies should investigate whether microbiota-targeted interventions, such as a probiotic regimen, could play a preventive role in patients with established IE risk factors, including valve defects, presence of prosthesis, or a history of prior IE.

We recently showed that the individual gut microbiome (GM) configuration in children and adolescents, together with long-term dietary habits, can predict the development of obesity. Here, we expanded our previous cohort to include 218 individuals and used 16S rRNA amplicon sequencing, shotgun metatranscriptomics, and a network approach to analyze fecal samples collected at a baseline survey and after a 4-year follow-up, investigating associations with health status, dietary intake, and other health-related behaviors. Our results showed that an unbalanced GM profile in children/adolescents, with few represented species, poor connectivity, and low transcriptional activity (especially in relation to molecular effectors that positively influence gut and immune health), combined with unhealthy behaviors (i.e., low-fiber diet and reduced physical activity), may favor the onset of obesity. This knowledge may pave the way for the development of adjunct GM-based precision intervention strategies aimed at rewiring microbial networks to promote long-term health.

Evidence suggests that a gluten-free diet may increase the risk of metabolic abnormalities associated with cardiovascular disease in adults with Coeliac Disease (CeD). This 9-week, double-blind, placebo-controlled, randomised pilot study investigated the effects of a combined supplement containing probiotic Lactiplantibacillus plantarum ECGC 13110402 and plant sterols and stanols, on cardiometabolic biomarkers and gut microbiota diversity and composition in adults with CeD and hypercholesterolaemia. Blood lipid profiles and vitamin D concentrations were analysed, and gut microbiota was profiled via 16S rRNA amplicon sequencing. In the active group, significant reductions in total cholesterol, LDL-cholesterol, non-HDL cholesterol, and apolipoprotein B were observed at multiple time points during the treatment phase, with changes generally greater in magnitude compared with the placebo group. Vitamin D levels also increased in the active group during supplementation. Microbiota analysis revealed potentially beneficial changes in participants receiving the active formulation, including higher alpha diversity and higher proportions of Bifidobacterium spp., Christensenellaceae R-7 group, and Lachnospiraceae ND3007 group. Overall, this feasibility study provides exploratory findings that a combined Lactiplantibacillus plantarum ECGC 13110402-phytosterol formulation may support lipid management and beneficially modulate gut microbiota in adults with CeD, particularly for those seeking non-pharmacological approaches to improving cardiometabolic health biomarkers.

PurposeInteraction of gut microbiota (GM) with dietary sugars (glucose, sorbitol) and choline has been transversely implicated in the pathogenesis of multiple chronic diseases. Our aim was to develop functional PET imaging of GM, using a multi-tracer approach to capture bacteria classes involved in sugar fermentation and choline catabolism at their gastrointestinal (GI) location.MethodsAdult and young sex-balanced groups of mice underwent oral administration of [18F]FDG, [18F]FDS or [11C]choline ([11C]cho) and repeated PET imaging over 4-5 h. Antibiotics, probiotic or faecal microbiota transplantation (FMT) served to quantify the specific role and site of bacteria action. GM was sequenced ex-vivo; gut histology and metabolic profiles were assessed in subsets.Results[18F]FDG and [18F]FDS reflected caecum abundance of Clostridia and Bacteroidia fermenters, with [18F]FDG exhibiting strongest and broadest relations. Clearance of [11C]cho from small gut reflected Bacilli and Lactobacilli abundance. In vitro cultures supported these relationships. Urinary 11C-excretion was nearly abolished by antibiotics. PET imaging was able to differentiate and predict gut bacteria classes in mice receiving FMT from two age-extreme human donors. Urinary [18F]FDS excretion reflected small-gut goblet cell activation; high caecum [18F]FDG retention and small gut [11C]cho clearance predicted body glucose use and low systemic inflammation.ConclusionImaging of ingested probes is simple and effective to map GM characteristics in situ and the functional crosstalk with host processes in mice in real-time. Our data confirm that the GI ecosystem is highly diversified, pointing to small intestine and caecum GM as dominant players in gut-body handling of our target nutrients.

The Gut Microbiome-Liver-Brain Axis is a relatively novel construct with promising potential to enhance our understanding of Alcohol Use Disorder (AUD), and its therapeutic approaches. Significant alterations in the gut microbiome occur in AUD even before any other systemic signs or symptoms manifest. Prolonged and inappropriate alcohol consumption, by affecting the gut microbiota and gut mucosa permeability, is thought to contribute to the development of behavioral and cognitive impairments, leading to Alcohol-Related Liver Disorders and potentially progressing into alcoholic cirrhosis, which is often associated with severe cognitive impairment related to neurodegeneration, such as hepatic encephalopathy and alcoholic dementia.The critical role of the gut microbiota is further supported by the efficacy of FDA-approved treatments for hepatic encephalopathy in alcoholic cirrhosis (i.e., lactulose and rifaximin). To stimulate new research, we hypothesize that interactions between a maladaptive stress response and a constitutional predisposition to neurodegeneration underlie the progression of AUD to conditions of Alcohol-Related Clinical Concerns with severe cognitive impairment, which represent a significant and costly burden to society. Early identification of AUD individuals at risk for developing these conditions could help to prioritize integrated therapeutic interventions targeting different substrates of the Gut Microbiome-Liver-Brain axis. Specifically, addiction medications, microbiome modulators, stress-reducing interventions, and, possibly soon, novel agents that reduce hepatic steatosis/fibrosis will be discussed in the context of digitally supported integrated therapeutic approaches. The explicit goal of this AUD treatment performed on the early stage of the disorder would be to reduce the transition from AUD to those conditions of Alcohol-Related Common Clinical Concerns associated with severe cognitive impairment, a strategy recommended for most neurological neurodegenerative disorders.

Chronic kidney disease (CKD) may be associated with dysbiosis which may increase the risk of gastrointestinal infections. Patients with kidney failure have a predominance of bacteria responsible for the exacerbation of chronic inflammation through the production of ureases, uricase, and uremic toxins and a reduction of bacteria-producing protective molecules as short-chain fatty acids. Patients with CKD have an increased risk of Clostridioides difficile infection. Currently, besides antibiotic therapy, fecal microbiota transplantation (FMT) is the only effective gut microbiota-targeted therapy for treating this infection. Scant evidence is available on FMT in those receiving peritoneal dialysis (PD). In this case, we report a successful FMT performed by colonoscopy in a patient receiving PD for polycystic kidney disease suffering from recurrent Clostridioides difficile infections. The FMT was repeated to enhance microbiota engraftment. The role of FMT in treating Clostridioides difficile in individuals receiving PD may be an important and promising therapeutic strategy but requires further prospective study.

Acute lymphoblastic leukaemia (ALL) represents the most common childhood malignancy, and emerging evidence underscores the impact of the gut microbiome (GM) on its pathogenesis. In this study, we used shotgun metagenomics to investigate the GM of 30 ALL patients at diagnosis-19 with B-ALL and 11 with T-ALL-and compared them to 176 healthy controls (HCs). When considered as a single ALL group versus HCs, clear compositional differences emerged: ALL patients exhibited higher relative abundances of Enterococcus faecium, oral commensals such as Rothia dentocariosa, and multiple opportunistic species, whereas HCs were enriched in short-chain fatty acid producers like Anaerostipes hadrus and Intestinibacter bartlettii. Functionally, the ALL GM relied more on protein and amino acid catabolism, while HCs possessed enhanced pathways for carbohydrate and folate metabolism. These findings broadly align with 16S rRNA-based analyses from previous publications, though some discrepancies highlight differences in technique-driven resolution. In contrast, comparing the two major molecular phenotypes-B-ALL and T-ALL-revealed only minimal taxonomic and functional differences, primarily confined to BAs metabolism pathways. Overall, our results indicate that children with ALL at the time of diagnosis already display a dysbiotic signature, bolstering the notion that a disturbance in GM development during childhood may be linked to the multistep pathogenesis model of ALL.

BACKGROUND:The global prevalence of obesity and type 2 diabetes, particularly among children, is rising, yet the long-term impacts of early-life fecal microbiota transplantation (FMT) on metabolic health remain poorly understood. OBJECTIVES:To investigate how early-life FMT from children to young, sex-matched mice influences metabolic outcomes and adipose tissue function in later, adult life. METHODS:Germ-free mice were colonized with fecal microbiota from either lean children or children with obesity. The impacts on brown adipose tissue (BAT), white adipose tissue (WAT), glucose metabolism, and gut health were analyzed in male and female mice. Microbial communities and metabolite profiles were characterized using sequencing and metabolomics. RESULTS:Male mice receiving FMT from obese donors exhibited marked BAT whitening and impaired amino acid and glucose metabolism. In contrast, female recipients developed hyperglycemia, accompanied by gut barrier dysfunction and WAT impairment. Distinct microbial and metabolite profiles were associated with these phenotypes: Collinsella and trimethylamine in females; and Paraprevotella, Collinsella, Lachnospiraceae NK4A136, Bacteroides, Coprobacillus, and multiple metabolites in males. These phenotypic effects persisted despite changes in host environment and diet. CONCLUSIONS:Early-life FMT induced long-lasting effects on the metabolic landscape, profoundly affecting adipose tissue function and systemic glucose homeostasis in adulthood. Donor dietary habits correlated with the fecal microbial profiles observed in recipient mice. These findings highlight the critical need for identifying and leveraging beneficial exposures during early development to combat obesity and diabetes.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) profoundly reshapes the gut microbiome (GM), yet age-related differences in this process remain incompletely understood. In this study, we analyzed stool samples from a cohort of pediatric and adult patients collected before allo-HSCT and at neutrophil engraftment to assess GM dynamics. We observed a significant reduction in alpha diversity post-HSCT across both age groups, with young patients displaying notably lower biodiversity. While GM composition varies among age ranges before transplantation, these distinctions largely disappeared at neutrophil engraftment, indicating a convergence toward a more uniform “transplant-specific” signature. This shift entailed a loss of health-associated, butyrate-producing taxa and a rise in potentially opportunistic bacteria (e.g., Enterococcaceae, Staphylococcaceae). Low pre-allo-HSCT GM diversity correlated with an increased risk of clinically significant acute graft-versus-host disease, overwhelming the age differences, underscoring the clinical relevance of maintaining a balanced microbiome before transplantation. Overall, our findings highlight the importance of age in shaping the pre-transplant GM and reveal how allo-HSCT-driven factors homogenize microbial communities among age ranges, offering insights for future microbiome-targeted interventions to improve transplant outcomes.

Abstract Background Wnt/β-catenin signalling impairment accounts for 85% of colorectal cancers (CRCs), including sporadic and familial adenomatous polyposis (FAP) settings. An altered PI3K/mTOR pathway and gut microbiota also contribute to CRC carcinogenesis. We studied the interplay between the two pathways and the microbiota composition within each step of CRC carcinogenesis. Methods Proteins and target genes of both pathways were analysed by RT-qPCR and IHC in tissues from healthy faecal immunochemical test positive (FIT+, n = 17), FAP (n = 17) and CRC (n = 15) subjects. CRC-related mutations were analysed through NGS and Sanger. Oral, faecal and mucosal microbiota was profiled by 16 S rRNA-sequencing. Results We found simultaneous hyperactivation of Wnt/β-catenin and PI3K/mTOR pathways in FAP-lesions compared to CRCs. Wnt/β-catenin molecular markers positively correlated with Clostridium_sensu_stricto_1 and negatively with Bacteroides in FAP faecal microbiota. Alistipes, Lachnospiraceae, and Ruminococcaceae were enriched in FAP stools and adenomas, the latter also showing an overabundance of Lachnoclostridium, which positively correlated with cMYC. In impaired-mTOR-mutated CRC tissues, p-S6R correlated with Fusobacterium and Dialister, the latter also confirmed in the faecal-ecosystem. Conclusions Our study reveals an interplay between Wnt/β-catenin and PI3K/mTOR, whose derangement correlates with specific microbiota signatures in FAP and CRC patients, and identifies new potential biomarkers and targets to improve CRC prevention, early adenoma detection and treatment.