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Introduction: Calcific aortic valve disease (CAVD) is the most common heart valve disorder, defined by a remodeling multistep process: namely, valve fibrosis with its area narrowing, impaired blood flow, and final calcification phase. Nowadays, the only treatment is the surgical valve replacement. As for other cardiovascular diseases, growing evidence suggest an active role of the immune system in the calcification process that could be modulated by the microbiota. To address this point, we aimed to investigate and characterize, for the first time, the presence of a valve microbiota and associated immune response in human CAVD. Method: Calcified aortic valve (CAV) samples from twenty patients (11 from Germany and 9 from Italy) with diagnosis of severe symptomatic CAVD were used to assess the presence of infiltrating T cells, by cloning approach, and to characterize the valve microbiota, by 16S rRNA gene sequencing (NGS). Results: We documented the presence of infiltrating T lymphocytes, especially the T helper subset, in CAV samples. Moreover, we found a tissue-associated microbiota in freshly collected CAV samples, which was significantly different in Italian and German patients, suggesting potential correlation with other cardiovascular risk factors. Conclusion: The presence of microbiota in inflamed CAV samples represents the right trigger point to explain the valve calcification process, encouraging further studies to explore the potential link between bacteria and adaptive immune response and to define the critical role of local microbiota-immunity axis on CAVD development.

Microbiome research has enormous potential in cancer research and the use of formalin-fixed paraffin-embedded (FFPE) tissues could offer many advantages. The tumor microenvironment represents a suitable niche for specific microbes and evidence proves the presence of an endogenous tumor microbiota, here referred to as oncobiota. Awareness of the oncobiota role in tumorigenesis could have a large influence on cancer care, in terms of diagnosis, prevention, and treatment. Moreover, understanding the microbial-related tumor microenvironment, and its influence on tumor immune response and cancer cells will help define important pathogenetic mechanisms in cancer starting or progression. Routine collection of histopathological FFPE samples provides a large availability of specimens essential for affordable and impactful retrospective analyses and for getting robust statistical results. The FFPE tissues are common in the analysis of tumor biopsies including the tumor microbiota characterization which has an important role in the modulation of our immune system and consequently of tumor cells. However, the microbiota analysis starting from FFPE tissues presents methodological pitfalls and limits that may negatively affect the oncobiota research. After examining the methodological and analytical difficulties of this approach, this work seeks to offer workable solutions to promote that research area.

Objectives We characterized the microbiota in systemic sclerosis (SSc), focusing on the skin-oral-gut axis and serum and faecal free fatty acid (FFA) profile. Methods Twenty-five SSc patients with ACA or anti-Scl70 autoantibodies were enrolled. The microbiota of faecal, saliva and superficial epidermal samples was assessed through next generation-sequencing analysis. Gas chromatography-mass spectroscopy was used to quantity faecal and serum FFAs. Gastrointestinal symptoms were investigated with UCLA GIT-2.0-questionnaire. Results The ACA+ and anti-Scl70+ groups displayed different cutaneous and faecal microbiota profiles. The classes of cutaneous Sphingobacteria and Alphaproteobacteria, the faecal phylum of Lentisphaerae, the classes of Lentisphaeria and Opitutae, and the genus of NA-Acidaminococcaceae were significantly higher in faecal samples of the ACA+, compared with anti-Scl70+ patients. The cutaneous Sphingobacteria and the faecal Lentisphaerae were significantly correlated (rho = 0.42; p= 0.03). A significant increase in faecal propionic acid was observed in ACA+ patients. Moreover, all faecal medium-chain FFAs and hexanoic acids were significantly higher in ACA+ compared with anti-Scl70+ group (p< 0.05 and p< 0.001, respectively). In the ACA+ group, the analysis of serum FFA levels showed an increasing trend in valeric acid. Conclusions Different microbiota signatures and FFA profiles were found in the two groups of patients. Despite being in different body districts, the cutaneous Sphingobacteria and faecal Lentisphaerae appear interdependent.

Background and aimIn recent decades, obesity prevalence has reached epidemic proportions and considering the pivotal role of gut microbiota (GM) in the regulation of energy balance, alternative non-pharmacological approaches involving probiotics’ administration have been proposed. The aim of the present study was to evaluate the effect of Lactiplantibacillus plantarum IMC 510® supplementation on anthropometric and biochemical parameters, GM composition and functionality, and gastrointestinal and general symptoms of overweight/obese subjects.MethodsForty overweight/obese subjects were randomly assigned to daily consume the probiotic Lactiplantibacillus plantarum IMC 510® or placebo for 3 months. Before and after the administration period, anthropometric and biochemical parameters, self-administered questionnaires, and plasma and stool samples were obtained from each participant. The GM characterization was performed with 16S rRNA sequencing, while fecal short (SCFAs) and medium (MCFAs) chain fatty acids were analyzed with a gas chromatography–mass spectrometry protocol.ResultsCompared to placebo, probiotic supplementation determined a significant decrease in body weight, BMI, waist circumference, waist-to-height ratio, and blood glucose. Moreover, probiotic administration produced a significant decrease of the genera Hafnia-Obesumbacterium and Romboutsia and an increase of Succiniclasticum spp.; conversely, placebo administration resulted in the decrease of Actinomycetaceae and an increase of both Alloprevotella spp. and of the levels of pro-inflammatory hexanoic and heptanoic acids.ConclusionThanks to its effect in increasing some beneficial gut bacteria and lowering effects on waist circumference, fasting glucose levels and gastrointestinal symptoms of obese subjects, Lactiplantibacillus plantarum IMC 510® supplementation could represent a future and encouraging strategy for the prevention or treatment of obesity.

Background and aimsCrohn’s disease (CD) pathogenesis is still unclear. Remodeling in mucosal microbiota and systemic immunoregulation may represent an important component in tissue injury. Here, we aim to characterize the ileal microbiota in both pathological and healthy settings and to evaluate the correlated systemic microbial-associated inflammatory markers comparing first-time surgery and relapse clinical conditions.MethodsWe enrolled 28 CD patients at surgery; we collected inflamed and non-inflamed mucosa tissues and blood samples from each patient. Bacterial wall adherence was observed histologically, while its composition was assessed through amplicon sequencing of the 16S rRNA gene. In addition, we evaluated the systemic microRNA (miRNA) using quantitative real-time PCR amplification and free fatty acids (FFAs) using gas chromatography–mass spectroscopy.ResultsThe total number of mucosal adherent microbiota was enriched in healthy compared to inflamed mucosa. In contrast, the phylum Tenericutes, the family Ruminococcaceae, and the genera Mesoplasma and Mycoplasma were significantly enriched in the pathological setting. Significant microbiota differences were observed between the relapse and first surgery patients regarding the families Bacillaceae 2 and Brucellaceae and the genera Escherichia/Shigella, Finegoldia, Antrobacter, Gemmatimonas, Moraxella, Anoxibacillus, and Proteus. At the systemic level, we observed a significant downregulation of circulating miR-155 and miR-223, as well as 2-methyl butyric, isobutyric, and hexanoic (caproic) acids in recurrence compared to the first surgery patients. In addition, the level of hexanoic acid seems to act as a predictor of recurrence risk in CD patients (OR 18; 95% confidence interval 1.24–261.81; p = 0.006).ConclusionsWe describe a dissimilarity of ileal microbiota composition comparing CD and healthy settings, as well as systemic microbial-associated inflammatory factors between first surgery and surgical relapse. We suggest that patterns of microbiota, associated with healthy ileal tissue, could be involved in triggering CD recurrence. Our findings may provide insight into the dynamics of the gut microbiota–immunity axis in CD surgical recurrence, paving the way for new diagnostics and therapeutics aimed not only at reducing inflammation but also at maintaining a general state of eubiosis in healthy tissue.

Background and aimIn recent years, many studies have suggested that ancient wheat products might have beneficial effects on cardiometabolic risk profile, but little is known about their effect on gut microbiota (GM). The aim of the present study was to evaluate whether a replacement diet with pasta made from ancient wheat (AD) could influence the GM composition and its metabolites’ production compared to a replacement diet with pasta made from modern wheat (CD).MethodsA randomized, double-blinded crossover trial with two intervention phases was conducted on 20 clinically healthy adults (9 females; 11 males; mean age 43.1 ± 12.5 years). Study participants were assigned to consume pasta made using semi-whole flour from organic wheat that was either from ancient or modern control wheat for 8 weeks in a random order. An 8-week washout period was implemented between the interventions. Stool samples were collected from all subjects at the beginning and at the end of each intervention period. GM composition, and short- (SCFAs) and medium- chain fatty acids (MCFAs) production was evaluated.ResultsDietary interventions did not produce significant diversity in the GM composition at higher ranks (phylum, class, order and family), but only at genus level. In detail, the AD significantly (adj. p < 0.05) changed the abundance of Erysipelatoclostridium spp., Bacteroides_pectinophilus_group spp., CAG-873 spp., and Holdemanella spp. The CD significantly affected the abundance of Akkermansia spp., CAG-873 spp., Hungatella spp., Lachnospiraceae_UCG-008 spp., NK4A214_group spp., Frisingicoccus spp., Megasphaera spp., Synergistes spp., and Tyzzerella spp. Regarding the production of SCFAs and MCFAs, AD resulted in a significant increase of fecal acetic (+0.7%), isobutyric (+30.1%), 2-methylbutyric (+64.2%), and isovaleric (+22.5%) acids. On the other hand, CD resulted in increased levels of isobutyric (+71.4%), 2-methylbutyric (+116.2%), isovaleric (+99%), and valeric (+21.4%) acids, and a reduction of butyric (-31.6%) and hexanoic (-66.4%) acids.ConclusionA short-term replacement diet with both ancient and modern wheat pasta determined significant changes in GM composition at the genus level but notably the AD resulted in a greater beneficial impact on anti-inflammatory SCFAs.

Eosinophilic granulomatosis with polyangiitis (EGPA) is an anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis. A genome-wide association study showed a correlation between ANCA-negative EGPA and variants of genes encoding proteins with intestinal barrier functions, suggesting that modifications of the mucosal layer and consequent gut dysbiosis might be involved in EGPA pathogenesis. Here, we characterized the gut microbiota (GM) composition and the intestinal immune response in a cohort of EGPA patients. Faeces from 29 patients and 9 unrelated healthy cohabitants were collected, and GM and derived metabolites’ composition were compared. Seven intestinal biopsies from EGPA patients with gastrointestinal manifestations were analysed to assess the T-cell distribution and its correlation with GM and EGPA clinical and laboratory features. No significant differences in GM composition, nor in the total amount of faecal metabolites, emerged between patients and controls. Nevertheless, differences in bacterial taxa abundances and compositional GM-derived metabolites profile were observed. Notably, an enrichment of potential pathobionts (Enterobacteriacee and Streptococcaceae) was found in EGPA, particularly in patients with active disease, while lower levels were found in patients on immunosuppression, compared with non-immunosuppressed ones. Significantly lower amounts of hexanoic acid were found in patients, compared to controls. The analysis of the immune response in the gut mucosa revealed a high frequency of IFN-γ/IL-17-producing T lymphocytes, and a positive correlation between EGPA disease activity and intestinal T-cell levels. Our data suggest that an enrichment in potential intestinal pathobionts might drive an imbalanced inflammatory response in EGPA.

Crohn's disease (CD) pathogenesis is still unclear. Remodeling in mucosal microbiota and systemic immunoregulation may represent an important component in tissue injury. Here, we aim to characterize the ileal microbiota in both pathological and healthy settings and to evaluate the correlated systemic microbial-associated inflammatory markers comparing first-time surgery and relapse clinical conditions.We enrolled 28 CD patients at surgery; we collected inflamed and non-inflamed mucosa tissues and blood samples from each patient. Bacterial wall adherence was observed histologically, while its composition was assessed through amplicon sequencing of the 16S rRNA gene. In addition, we evaluated the systemic microRNA (miRNA) using quantitative real-time PCR amplification and free fatty acids (FFAs) using gas chromatography-mass spectroscopy.The total number of mucosal adherent microbiota was enriched in healthy compared to inflamed mucosa. In contrast, the phylumWe describe a dissimilarity of ileal microbiota composition comparing CD and healthy settings, as well as systemic microbial-associated inflammatory factors between first surgery and surgical relapse. We suggest that patterns of microbiota, associated with healthy ileal tissue, could be involved in triggering CD recurrence. Our findings may provide insight into the dynamics of the gut microbiota-immunity axis in CD surgical recurrence, paving the way for new diagnostics and therapeutics aimed not only at reducing inflammation but also at maintaining a general state of eubiosis in healthy tissue.

In this exploratory study, we investigate the variation in the facial skin microbiome architecture through aging and their functional association with host genetic factors in a cohort of healthy women, living in the same area and without cutaneous diseases. Notably, facial skin microbiota (SM) samples were collected from a cohort of 15 healthy Caucasian females, firstly divided into three age groups (younger women aged 20–35 years old; middle aged women of 36–52 years old; and older women aged 53–68 years old). Then, the recruited cohort was divided into two groups based on their facial hydration level (dry and normal skin). The facial SM revealed a different composition in the three analyzed aging groups and between normal and dry skins. The middle-aged women also revealed functional variations associated with collagen biosynthesis and oxidative stress damage repair. Otherwise, the association between selected host SNPs (single nucleotide polymorphisms) and the facial SM profile showed significant associations, suggesting a negative correlation with collagen metabolism and ROS damage protection. Finally, the composition and functionality of the facial SM seemed to affect the aging process through the two aging-correlated pathways of host ROS damage repair and collagen metabolism. Our exploratory data could be useful for future studies characterizing the structure, function, and dynamics of the SM in the aging process to design personalized therapeutic agents focusing on potential genomic targets, microbes, and their metabolites.

Fibromyalgia (FM) syndrome is mainly characterized by widespread pain, sleeping disorders, fatigue, and cognitive dysfunction. In many cases, gastrointestinal distress is also reported, suggesting the potential pathogenic role of the gut microbiota (GM). The GM is deeply influenced by several environmental factors, especially the diet, and recent findings highlighted significant symptom improvement in FM patients following various nutritional interventions such as vegetarian diet, low-fermentable oligosaccharides, disaccharides, monosaccharides, and polyols based diets, gluten-free diet, and especially an ancient grain supplementation. In particular, a recent study reported that a replacement diet with ancient Khorasan wheat led to an overall improvement in symptom severity of FM patients.To examine the effects of ancient Khorasan wheat on the GM, inflammation, and short-chain fatty acid production in FM patients.After a 2-wk run-in period, 20 FM patients were enrolled in this randomized, double-blind crossover trial. In detail, they were assigned to consume either Khorasan or control wheat products for 8 wk and then, following an 8-wk washout period, crossed. Before and after treatments, GM characterization was performed by 16S rRNA sequencing while the fecal molecular inflammatory response and the short-chain fatty acids (SCFAs) were respectively determined with the Luminex MAGPIX detection system and a mass chromatography-mass spectrometry method.The Khorasan wheat replacement diet, in comparison with the control wheat diet, had more positive effects on intestinal microbiota composition and on both the fecal immune and SCFAs profiles such as the significant increase of butyric acid levels (The replacement diet based on ancient Khorasan wheat results in beneficial GM compositional and functional modifications that positively correlate with an improvement of FM symptomatology.