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Ensuring the viability and efficacy of probiotic microorganisms during manufacturing and gastrointestinal transit remains challenging, particularly for sensitive strains such as certain lactic acid bacteria and bifidobacteria. This has led to increased interest in spore-forming bacteria, such as Heyndrickxia coagulans (formerly Bacillus coagulans), which can endure environmental stresses through their endospore forms. This study presents a comprehensive analysis of the probiotic potential of strain LMG S-24828, originally isolated from healthy human feces. The genomic analysis confirmed the strain's taxonomic placement within the species H. coagulans and revealed no extrachromosomal plasmid DNA, suggesting genetic stability. Safety assessments demonstrated that LMG S-24828 does not produce D-lactate, deconjugate bile salts, or exhibit hemolytic activity, and it lacks transmissible antibiotic resistances. Phenotypic tests showed the strain's metabolic versatility, including its ability to hydrolyze complex carbohydrates and adhere to intestinal epithelial cells. Moreover, LMG S-24828 exhibited robust survival and germination during in vitro and in vivo gastrointestinal simulations, with evidence of significant spore germination in the human gut. These findings suggest that H. coagulans LMG S-24828 possesses several advantageous traits for probiotic applications, warranting further clinical evaluation to confirm its health benefits., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Malnutrition is linked to 45% of global childhood mortality, however, the impact of maternal malnutrition on the child's health remains elusive. Previous studies suggested that maternal malnutrition does not affect breast milk composition. Yet, malnourished children often develop a so-called environmental enteropathy, assumed to be triggered by frequent pathogen uptake and unfavorable gut colonization. Here, we show in a murine model that maternal malnutrition induces a persistent inflammatory gut dysfunction in the offspring that establishes during nursing and does not recover after weaning onto standard diet. Early intestinal influx of neutrophils, impaired postnatal development of gut-regulatory functions, and expansion of Enterobacteriaceae were hallmarks of this enteropathy. This gut phenotype resembled those developing under deficient S100a8/a9-supply via breast milk, which is a known key factor for the postnatal development of gut homeostasis. We could confirm that S100a8/a9 is lacking in the breast milk of malnourished mothers and the offspring's intestine. Nutritional supply of S100a8 to neonates of malnourished mothers abrogated the aberrant development of gut mucosal immunity and microbiota colonization and protected them lifelong against severe enteric infections and non-infectious bowel diseases. S100a8 supplementation after birth might be a promising measure to counteract deleterious imprinting of gut immunity by maternal malnutrition., (© 2024. The Author(s).)

Background: Older subjects are at risk of elevated intestinal permeability (IP) which can lead to immune system activation and low-grade systemic inflammation. Dietary changes are a potential strategy to reduce IP. The MaPLE project evaluated the hypothesis that increasing (poly)phenol intake would beneficially impact on several important markers and pathways related to IP. The objective of the present study was to assess the effects of the MaPLE (poly)phenol-rich diet (PR-diet) on additional IP-related biomarkers and any relationships between biomarker responses., Methods: A randomised, controlled, crossover study was performed involving 51 participants (≥ 60 y) with increased IP, as determined by serum zonulin levels. Participants were randomly assigned to one of two intervention groups: a control diet (C-diet) or a PR-diet. Each intervention lasted 8 weeks and was separated by an 8-week washout period. For the present study, serum and faecal samples were used to measure zonula occludens-1 (ZO-1), occludin, adiponectin, calprotectin, faecal calprotectin, soluble cluster of differentiation 14 (sCD14), interleukin-6 receptor (IL-6R), and vascular endothelial-cadherin (VEC) levels using quantitative ELISA assays. Data were analysed using ANOVA, and Spearman and network correlation analysis were performed to identify the relationship among biomarkers at baseline., Results: Among the different markers analysed, a significant reduction was observed for faecal and serum calprotectin (p = 0.0378 and p = 0.0186, respectively) following the PR-diet, while a significant increase in ZO-1 was found (p = 0.001) after both the intervention periods (PR-diet and C-diet). In addition, a time effect was observed for VEC levels showing a reduction (p = 0.038) following the PR-diet. Based on network correlation analysis, two clusters of correlations were identified: one cluster with high levels of serum calprotectin, faecal calprotectin, sCD14, interleukin (IL)-6, tumor necrosis factor (TNF)-α, C-reactive protein (CRP) and bacterial DNAemia (16 S rRNA gene copies), with potential inflammatory-induced intestinal permeability. Differently, the other cluster had high levels of serum occludin, IL-6R, soluble intercellular adhesion molecule-1 (sICAM-1) and VEC, with potential inflammatory-induced endothelial dysfunction., Conclusions: Overall, this study provides further support to the hypothesis that a (poly)phenol-rich diet may help to ameliorate intestinal permeability-associated conditions. In this regard, calprotectin might represent a promising biomarker since it is a protein that typically increases with age and it is considered indicative of intestinal and systemic inflammation. Further research is needed to develop targeted (poly)phenol-rich diets against age-related gut dysfunction and inflammation., Trial Registration: 28/04/2017; ISRCTN10214981; https://doi.org/10.1186/ISRCTN10214981 ., (© 2024. The Author(s).)

Background & Aims: The pathophysiology of irritable bowel syndrome (IBS) is multifactorial and includes epithelial barrier dysfunction, a key element at the interface between the gut lumen and the deeper intestinal layers. Beneath the epithelial barrier there is the vascular one representing the last barrier to avoid luminal antigen dissemination The aims of this study were to correlate morpho-functional aspects of epithelial and vascular barriers with symptom perception in IBS., Methods: Seventy-eight healthy subjects (controls) and 223 patients with IBS were enrolled in the study and phenotyped according to validated questionnaires. Sugar test was used to evaluate in vivo permeability. Immunohistochemistry, western blot, and electron microscopy were used to characterize the vascular barrier. Vascular permeability was evaluated by assessing the mucosal expression of plasmalemma vesicle-associated protein-1 and vascular endothelial cadherin. Caco-2 or human umbilical vein endothelial cell monolayers were incubated with soluble mediators released by mucosal biopsies to highlight the mechanisms involved in permeability alteration. Correlation analyses have been performed among experimental and clinical data., Results: The intestinal epithelial barrier was compromised in patients with IBS throughout the gastrointestinal tract. IBS-soluble mediators increased Caco-2 permeability via a downregulation of tight junction gene expression. Blood vessel density and vascular permeability were increased in the IBS colonic mucosa. IBS mucosal mediators increased permeability in human umbilical vein endothelial cell monolayers through the activation of protease-activated receptor-2 and histone deacetylase 11, resulting in vascular endothelial cadherin downregulation. Permeability changes correlated with intestinal and behavioral symptoms and health-related quality of life of patients with IBS., Conclusions: Epithelial and vascular barriers are compromised in patients with IBS and contribute to clinical manifestations., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Environmental enteric dysfunction (EED) is a condition associated with malnutrition that can progress to malabsorption and villous atrophy. Severe EED results in linear growth stunting, slowed neurocognitive development, and unresponsiveness to oral vaccines. Prenatal exposure to malnutrition and breast feeding by malnourished mothers replicates EED. Pups are characterized by deprivation of secretory IgA (SIgA) and altered development of the gut immune system and microbiota. Extracellular ATP (eATP) released by microbiota limits T follicular helper (Tfh) cell activity and SIgA generation in Peyer's patches (PPs). Administration of a live biotherapeutic releasing the ATP-degrading enzyme apyrase to malnourished pups restores SIgA levels and ameliorates stunted growth. SIgA is instrumental in improving the growth and intestinal immune competence of mice while they are continuously fed a malnourished diet. The analysis of microbiota composition suggests that amplification of endogenous SIgA may exert a dominant function in correcting malnourishment dysbiosis and its consequences on host organisms, irrespective of the actual microbial ecology., Competing Interests: Declaration of interests F.G. and L.P. are inventors in a patent application covering the effect of apyrase in correcting dysbiosis. F.G. is the founder of MV BioTherapeutics, a company developing mucosal vaccines and biotherapeutics., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Confectionary products hold promise as unconventional food carriers for probiotic microorganisms. This study explored the delivery of Heyndrickxia coagulans SNZ1969, a spore-forming probiotic, using gummy candies. In this study, we prepared gummy candies containing bacterial spores with a viable count that remained stable during a 24-month shelf-life period, meeting the label claim of at least one billion CFUs per serving (24 g). Then, we carried out an intervention trial involving 24 healthy adults who consumed one serving per day for two weeks followed by an additional two weeks of follow-up. Fecal samples were collected and analyzed with a protocol that allowed the viable counts of SNZ1969, both in spore and vegetative forms. The obtained results revealed that bacterial spores germinated in all volunteers. SNZ1969 persistence in the gut was monitored for two weeks after the end of gummy candy consumption, indicating its potential for prolonged colonization. These findings highlight the potential of unconventional food carriers for probiotic delivery and suggest that spore-forming probiotics can be metabolically active in the human intestine. These findings provide information for the development of food products containing spore-forming probiotics and their potential benefits in promoting gastrointestinal health., Competing Interests: The following authors have affiliations with organizations with direct or indirect financial interest in the subject matter discussed in the manuscript:-Natalja Kirika and Andrea Sarrica are employed by Perfetti Van Melle S.p.A., the company that provided financial support for this study.-Simone Guglielmetti served as a consultant for Perfetti Van Melle S.p.A. during the period when this study was conducted., (© 2024 The Authors.)

It was theorized that modernization and the decline in harmless microbial populations associated with food have altered the gut microbiota, impacting host metabolism and immunity. Western dietary patterns, characterized by processed foods and preservation methods, may significantly reduce the microbial population associated with food. To mitigate the consequences of bacterial deprivation, the integration of these diets with fermented foods is commonly proposed. Nonetheless, non-fermented food consumed raw may also be an important source of viable microbial cells for the human microbiome. This study investigates whether salad-associated LAB can survive the gastrointestinal transit (GIT) and contribute to the gut microbiota. LAB strains were quantified and isolated from rocket salad (Eruca vesicaria subsp. sativa), and their survival through GIT was assessed via intervention trials in healthy adults and in vitro. Moreover, bacterial communities in fecal samples were analyzed after three days of rocket salad consumption. Washing with a sodium hypochlorite solution drastically reduced total bacterial load and eliminated viable LAB. The quantity of LAB introduced through salads did not significantly alter the gut microbiota composition. Rocket salads harbored Weissella and Leuconostoc species. A significant increase in Weissella spp. but not in Leuconostoc spp. was observed after the consumption of rocket salad. Simulated GIT experiments suggested that the food matrix and the initial number of ingested viable bacteria may have been important in determining survival. These findings propose that plant products could serve as sources of live LAB for the human gut. Further research with diverse vegetables and longer interventions is needed, encouraging studies on raw, non-fermented foods and their impact on the human intestinal microbiome., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Advanced glycation end products (AGEs) exert a key pathogenic role in the development of obesity and insulin resistance. Thanks to its abundance in bioactive compounds, the microalga Arthrospira platensis (spirulina, SP) is proposed as a nutritional supplement. Here, we investigated the potential anti-glycating properties of SP enriched with zinc (Zn-SP) and the following impact on diet-induced metabolic derangements. Thirty male C57Bl6 mice were fed a standard diet (SD) or a high-fat high-sugar diet (HFHS) for 12 weeks, and a subgroup of HFHS mice received 350 mg/kg Zn-SP three times a week. A HFHS diet induced obesity and glucose intolerance and increased plasma levels of pro-inflammatory cytokines and transaminases. Zn-SP administration restored glucose homeostasis and reduced hepatic dysfunction and systemic inflammation. In the liver of HFHS mice, a robust accumulation of AGEs was detected, paralleled by increased expression of the main AGE receptor (RAGE) and depletion of glyoxalase-1, whereas Zn-SP administration efficiently prevented these alterations reducing local pro-inflammatory responses. 16S rRNA gene profiling of feces and ileum content revealed altered bacterial community structure in HFHS mice compared to both SD and HFHS + Zn-SP groups. Overall, our study demonstrates relevant anti-glycation properties of Zn-SP which contribute to preventing AGE production and/or stimulate AGE detoxification, leading to the improvement of diet-related dysbiosis and metabolic derangements.

Purpose: To investigate the responsiveness and minimal clinically important difference (MCID) of the Functional Independence Measure (FIM) from a clinician's perspective in hip-fractured elderly undergoing inpatient rehabilitation., Materials and Methods: Data of 701 hip-fractured elderly admitted to our rehabilitation institute were retrospectively collected. Ten a priori hypotheses were formulated., Results: The effect size of the FIM scores after treatment were large (total = 1.38, motor = 1.78). Moderate correlations were found between the change total score of FIM and Barthel Index (BI, r s = 0.51), Functional Ambulation Categories (FAC, r s = 0.52) and a checklist regarding participants' independence in transfers (r s = 0.59). The area under the receiving operating characteristics were 0.82, 0.81, and 0.85 when BI, FAC and checklist were used as anchors, respectively. Based on these findings, the FIM showed high responsiveness (9/10 hypotheses met). Triangulation of findings showed that MCIDs of 22 and 21 points were the most appropriate for the FIM total and motor score, respectively., Conclusions: The FIM scale is appropriate for assessing independence and functional recovery in hip-fractured elderly. Improvements of 22 and 21 points at the total and motor FIM were identified as indicators of minimal clinical change in those admitted to a rehabilitation facility with an FIM total score between 40 and 80.

Probiotics are exploited for adjuvant treatment in IBS, but reliable guidance for selecting the appropriate probiotic to adopt for different forms of IBS is lacking. We aimed to identify markers for recognizing non-constipated (NC) IBS patients that may show significant clinical improvements upon treatment with the probiotic strain Lacticaseibacillus paracasei DG (LDG). To this purpose, we performed a post-hoc analysis of samples collected during a multicenter, double-blind, parallel-group, placebo-controlled trial in which NC-IBS patients were randomized to receive at least 24 billion CFU LDG or placebo capsules b.i.d . for 12 weeks. The primary clinical endpoint was the composite response based on improved abdominal pain and fecal type. The fecal microbiome and serum markers of intestinal (PV1 and zonulin), liver, and kidney functions were investigated. We found that responders (R) in the probiotic arm (25%) differed from non-responders (NR) based on the abundance of 18 bacterial taxa, including the families Coriobacteriaceae , Dorea spp. and Collinsella aerofaciens , which were overrepresented in R patients. These taxa also distinguished R (but not NR) patients from healthy controls. Probiotic intervention significantly reduced the abundance of these bacteria in R, but not in NR. Analogous results emerged for C. aerofaciens from the analysis of data from a previous trial on IBS with the same probiotic. Finally, C. aerofaciens was positively correlated with the plasmalemmal vesicle associated protein-1 (PV-1) and the markers of liver function. In conclusion, LDG is effective on NC-IBS patients with NC-IBS with a greater abundance of potential pathobionts. Among these, C. aerofaciens has emerged as a potential predictor of probiotic efficacy.

The gut microbiota is believed to be a critical factor in the pathogenesis of IBS, and its metabolic byproducts, such as short-chain fatty acids (SCFAs), are known to influence gut function and host health. Despite this, the precise role of SCFAs in IBS remains a topic of debate. In this study, we examined the bacterial community structure by 16S rRNA gene profiling and SCFA levels by UPLC-MS/MS in fecal samples from healthy controls (HC; n  = 100) and non-constipated patients (IBS-D and IBS-M; NC-IBS; n  = 240) enrolled in 19 hospitals in Italy. Our findings suggest a significant difference between the fecal microbiomes of NC-IBS patients and HC subjects, with HC exhibiting higher intra-sample biodiversity. Furthermore, we were able to classify non-constipated patients into two distinct subgroups based on their fecal SCFA levels (fecal catabotype "high" and "low"), each characterized by unique taxonomic bacterial signatures. Our results suggest that the fecal catabotype with higher SCFA levels may represent a distinct clinical phenotype of IBS that could have implications for its diagnosis and treatment. This study provides a new perspective on the intricate relationship between the gut microbiome and bowel symptoms in IBS, underscoring the importance of personalized strategies for its management.

The interplay between the intestinal microbiota and host is critical to intestinal ontogeny and homeostasis. MicroRNAs (miRNAs) may be an underlying link. Intestinal miRNAs are microbiota-dependent and, when shed in the lumen, affect resident microorganisms. Yet, longitudinal relationships between intestinal tissue miRNAs, luminal miRNAs, and luminal microorganisms have not been elucidated, especially in early life. Here, we investigated the postnatal cecal miRNA and microbiota populations, their relationship, and their impact on intestinal maturation in specific pathogen-free mice; we also assessed if they can be modified by intervention with allochthonous probiotic lactobacilli. We report that cecal and cecal content miRNA and microbiota signatures are temporally regulated, correlated, and modifiable by probiotics with implications for intestinal maturation. These findings help understand causal relationships within the gut ecosystem and provide a basis for preventing and managing their alterations in diseases throughout life. IMPORTANCE The gut microbiota affects intestinal microRNA (miRNA) signatures and is modified by host-derived luminal miRNA. This suggests the existence of close miRNA-microbiota relationships that are critical to intestinal homeostasis. However, an integrative analysis of these relationships and their evolution during intestinal postnatal maturation is lacking. We provide a system-level longitudinal analysis of miRNA-microbiota networks in the intestine of mice at the weaning transition, including tissue and luminal miRNA and luminal microbiota. To address causality and move toward translational applications, we used allochthonous probiotic lactobacilli to modify these longitudinal relationships and showed that they are critical for intestinal maturation in early life. These findings contribute to understand mechanisms that underlie the maturation of the intestinal ecosystem and suggest that interventions aiming at maintaining, or restoring, homeostasis cannot prescind from considering relationships among its components., Competing Interests: E.M.C. has received research support from the Lallemand Health Solutions and Ocean Spray and has received consultant fees from Danone. All other authors declare no conflict of interests. The probiotic strains utilized in this study were provided by the Lallemand Health Solutions (LHS), Inc.

Purpose: Garlic consumption has been inversely associated to intestinal adenoma (IA) and colorectal cancer (CRC) risk, although evidence is not consistent. Gut microbiota has been implied in CRC pathogenesis and is also influenced by garlic consumption. We analyzed whether dietary garlic influence CRC risk and bacterial DNA in blood., Methods: We conducted a case-control study in Italy involving 100 incident CRC cases, 100 IA and 100 healthy controls matched by center, sex and age. We used a validated food frequency questionnaire to assess dietary habits and garlic consumption. Blood bacterial DNA profile was estimated using qPCR and16S rRNA gene profiling. We derived odds ratios (ORs) and the corresponding 95% confidence intervals (CIs) of IA and CRC according to garlic consumption from multiple conditional logistic regression. We used Mann-Whitney and chi-square tests to evaluate taxa differences in abundance and prevalence., Results: The OR of CRC for medium/high versus low/null garlic consumption was 0.27 (95% CI = 0.11-0.66). Differences in garlic consumption were found for selected blood bacterial taxa. Medium/high garlic consumption was associated to an increase of Corynebacteriales order, Nocardiaceae family and Rhodococcus genus, and to a decrease of Family XI and Finegoldia genus., Conclusions: The study adds data on the protective effect of dietary garlic on CRC risk. Moreover, it supports evidence of a translocation of bacterial material to bloodstream and corroborates the hypothesis of a diet-microbiota axis as a mechanism behind the role of garlic in CRC prevention., (© 2023. The Author(s).)

The mammary gland hosts a microbiota, which differs between malignant versus normal tissue. We found that aerosolized antibiotics decrease murine mammary tumor growth and strongly limit lung metastasis. Oral absorbable antibiotics also reduced mammary tumors. In ampicillin-treated nodules, the immune microenvironment consisted of an M1 profile and improved T cell/macrophage infiltration. In these tumors, we noted an under-representation of microbial recognition and complement pathways, supported by TLR2/TLR7 protein and C3-fragment deposition reduction. By 16S rRNA gene profiling, we observed increased Staphylococcus levels in untreated tumors, among which we isolated Staphylococcus epidermidis, which had potent inflammatory activity and increased Tregs. Conversely, oral ampicillin lowered Staphylococcus epidermidis in mammary tumors and expanded bacteria promoting an M1 phenotype and reducing MDSCs and tumor growth. Ampicillin/paclitaxel combination improved the chemotherapeutic efficacy. Notably, an Amp-like signature, based on genes differentially expressed in murine tumors, identified breast cancer patients with better prognosis and high immune infiltration that correlated with a bacteria response signature. This study highlights the significant influence of mammary tumor microbiota on local immune status and the relevance of its treatment with antibiotics, in combination with breast cancer therapies., Competing Interests: Declaration of competing interest The authors declare no potential conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Reportedly, Western-type diets may induce the loss of key microbial taxa within the gastrointestinal microbiota, promoting the onset of noncommunicable diseases. It was hypothesized that the consumption of raw vegetables could contribute to the maintenance of the intestinal microbial community structure. In this context, we explored bacteria associated with commercial rocket salads produced through different farming practices: traditional (conventional, organic, and integrated) and vertical farming. Viable counts of mesophilic bacteria and lactic acid bacteria (LAB) were performed on plate count agar (PCA) and de Man-Rogosa-Sharpe (MRS) agar at pH 5.7, whereas metataxonomics through 16S rRNA gene sequencing was used to profile total bacteria associated with rocket salads. We found that rocket salads from vertical farming had much fewer viable bacteria and had a bacterial community structure markedly different from that of rocket salads from traditional farming. Furthermore, although α- and β-diversity analyses did not differentiate rocket samples according to farming techniques, several bacterial taxa distinguished organic and integrated from conventional farming salads, suggesting that farming practices could affect the taxonomic composition of rocket bacterial communities. LAB were isolated from only traditional farming samples and belonged to different species, which were variably distributed among samples and could be partly associated with farming practices. Finally, the INFOGEST protocol for in vitro simulation of gastrointestinal digestion revealed that several taxonomically different rocket-associated bacteria (particularly LAB) could survive gastrointestinal transit. This study suggests that commercial ready-to-eat rocket salads harbor live bacteria that possess the ability to survive gastrointestinal transit, potentially contributing to the taxonomic structure of the human gut microbiota. IMPORTANCE Western-type diets are composed of foods with a reduced amount of naturally occurring microorganisms. It was hypothesized that a microbe-depleted diet can favor the alteration of the human intestinal microbial ecosystem, therefore contributing to the onset of chronic metabolic and immune diseases currently recognized as the most significant causes of death in the developed world. Here, we studied the microorganisms that are associated with commercial ready-to-eat rocket salads produced through different farming practices. We showed that rocket salad (a widely consumed vegetal food frequently eaten raw) may be a source of lactic acid bacteria and other microbes that can survive gastrointestinal transit, potentially increasing the biodiversity of the intestinal microbiota. This deduction may be valid for virtually all vegetal foods that are consumed raw.

Fermented foods are receiving growing attention for their health promoting properties. In particular, there is a growing demand for plant-based fermented foods as dairy alternatives. Considering that soy is a vegetal food rich in nutrients and a source of the phytoestrogen isoflavones, the aim of this study was to select safe food microorganisms with the ability to ferment a soy drink resulting in a final product with an increased estrogenic activity and improved functional properties. We used milk kefir grains, a dairy source of microorganisms with proven health-promoting properties, as a starting inoculum for a soymilk. After 14 passages of daily inoculum in fresh soy drink, we isolated four lactic acid bacterial strains: Lactotoccus lactis subsp. lactis K03, Leuconostc pseudomesenteroides K05, Leuconostc mesenteroides K09 and Lentilactobacillus kefiri K10. Isolated strains were proven to be safe for human consumption according to the assessment of their antibiotic resistance profile and comparative genomics. Furthermore, functional characterization of the bacterial strains demonstrated their ability to ferment sugars naturally present in soybeans and produce a creamy texture. In addition, we demonstrated, by means of a yeast-based bioluminescence reporter system, that the two strains belonging to the genus Leuconostoc increased the estrogenic activity of the soybean drink. In conclusion, the proposed application of the bacterial strains characterized in this study meets the growing demand of consumers for health-promoting vegetal food alternatives to dairy products., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Background and Aim: An association between reflux and burning mouth syndrome (BMS) has been proposed. Aims of this study were: 1) to investigate the frequency of BMS in a sample of GERD patients; 2) to measure G17, in a sample of BMS patients; 3) to assess the efficacy of different therapeutical schedules for GERD in BMS patients., Methods: We divided the study in 3 main steps. In step one, we analyzed 500 consecutive GERD patients' type and frequency of extraesophageal manifestations including BMS. In step two, we collected 124 consecutive BMS patients' symptoms and  G17.  In step three, we evaluate the efficacy of 3 different drugs on BMS., Results: In step one, 204 patients complained heartburn; 31 globus pharyngeus; 52 chronic cough; 54 pharyngitis; 31 postnasal drip; 56 burning mouth symptoms; 34 noncardiac chest pain; 17 asthma and 21 sleep apnea. In step two, 29 patients had G17 ≤ 1 pg/L; 64 patients between 1 and 3; and 31 patients ≥ 3. In step three, 49 patients reported slight benefit with PPI, 75 no benefit. 61 patients reported slight benefit with sodium alginate and sodium bicarbonate, 63 no benefit. 23 reported an almost complete remission with HYCHSA, 26 slight benefit, 33 no benefit., Conclusions: Prevalence of BMS in GERD patients was similar to that reported for chronic chough and pharyngitis. Low levels of G17 were found in the majority of BMS patients. Finally, we observed a greater benefit from barrier drugs therapy than from PPI therapy in BMS patients. (www.actabiomedica.it).

Scope: Dietary polyphenols can alter the gut microbiota (GM) and promote the production of bioactive metabolites. Several indoles result of GM metabolism of dietary tryptophan have been associated with intestinal barrier integrity. Our aim is to study the changes in GM-derived indoles during a polyphenol-rich (PR) diet intervention in older adults., Methods and Results: Randomized, controlled, crossover trial in adults ≥ 60 years living in a residential care facility during an 8-week PR versus control diet (n = 51). Seven GM-tryptophan metabolites are measured in serum, and metataxonomic analysis of GM is performed on fecal samples. Exploratory subgroup analyses are performed based on renal function (RF). The PR-diet significantly increases serum indole 3-propionic acid (IPA) in subjects with normal RF, but not in subjects with impaired RF. Other GM-tryptophan metabolites are not affected. Comparison of baseline GM composition shows shifts in Bacteroidales order members as well as higher abundance of Clostridiales in participants with normal RF. During the trial, variations of IPA are associated with changes in C-reactive protein (β = 0.32, p = 0.010) and GM, particularly with the Clostridiales (r = 0.35, p < 0.001) and Enterobacteriales (r = -0.15, p < 0.05) orders., Conclusion: A PR diet increases the serum concentration of IPA in older adults with normal RF. Our findings may be important when defining appropriate dietary interventions for older adults., Trial Registration Number: ISRCTN10214981 (https://doi.org/10.1186/ISRCTN10214981)., (© 2022 The Authors. Molecular Nutrition & Food Research published by Wiley-VCH GmbH.)

Flavonoids have been inversely associated to colorectal cancer (CRC) and are plausible intermediaries for the relation among gut microbiome, intestinal permeability and CRC. We analyzed the relation of flavonoid intake with CRC and blood bacterial DNA. We conducted a case-control study in Italy involving 100 incident CRC cases and 200 controls. A valid and reproducible food-frequency questionnaire was used to assess dietary habits and to estimate six flavonoid subclass intakes. We applied qPCR and 16S rRNA gene profiling to assess blood bacterial DNA. We used multiple logistic regression to derive odds ratios (ORs) of CRC and Mann-Whitney and chi--square tests to evaluate abundance and prevalence of operational taxonomic units (OTUs) according to flavonoid intakes. Inverse associations with CRC were found for anthocyanidins (OR for the highest versus the lowest tertile = 0.24, 95% confidence interval, CI = 0.11-0.52) and flavanones (OR = 0.18, 95% CI = 0.08-0.42). We found different abundance and prevalence according to anthocyanidin and flavanone intake for OTUs referring to Oligoflexales order, Diplorickettsiaceae family, Staphylococcus , Brevundimonas , Pelomonas and Escherischia - Shigella genera, and Flavobacterium and Legionella species. The study provides evidence to a protective effect of dietary anthocyanidins and flavanones on CRC and suggests an influence of flavonoids on blood bacterial DNA, possibly through intestinal permeability changes.

An immunosuppressive microenvironment in lung concurs to pre-malignant lesions progression to cancer. Here, we explore if perturbing lung microbiota, which contribute to immunosuppression, by antibiotics or probiotic aerosol interferes with lung cancer development in a mouse carcinogen-induced tumor model. Urethane-injected mice were vancomycin/neomycin (V/N)-aerosolized or live or dead L. rhamnosus GG ( L.RGG )-aerosolized, and tumor development was evaluated. Transcriptional profiling of lungs and IHC were performed. Tumor nodules number, diameter and area were reduced by live or heat-killed L.RGG , while only a decrease in nodule diameter was observed in V/N-treated lungs. Both L.RGG and V/N reduced Tregs in the lung. In L.RGG -treated groups, the gene encoding the joining chain (J chain) of immunoglobulins was increased, and higher J chain protein and IgA levels were observed. An increased infiltration of B, NK and myeloid-derived cells was predicted by TIMER 2.0. The Kaplan-Meier plotter revealed an association between high levels of J chain mRNA and good prognosis in lung adenocarcinoma patients that correlated with increased B and CD4 T cells and reduced Tregs and M2 macrophages. This study highlights L.RGG aerosol efficacy in impairing lung cancer growth by promoting local immunity and points to this non-invasive strategy to treat individuals at risk of lung cancer.

A polyphenol-rich diet reduced intestinal permeability (IP) in older adults. Our aim was to evaluate if participants categorized according to urolithin metabotypes (UMs) exhibited different responses in the MaPLE trial. Fifty-one older adults (mean age: 78 years) completed an 8-week randomized-controlled-crossover trial comparing the effects of a polyphenol-rich vs. a control diet on IP, assessed through zonulin levels. Plasma and urinary metabolomics were evaluated with a semi-targeted UHPLC-MS/MS method. Gut microbiota was characterized by 16S rRNA gene profiling. UMs were determined according to urolithin excretion in 24 h urine samples. Multivariate statistics were used to characterize the differences in metabolomic and metataxonomic responses across UMs. Thirty-three participants were classified as urolithin metabotype A (UMA), 13 as urolithin metabotype B (UMB), and 5 as urolithin metabotype 0 (UM0) according to their urinary excretion of urolithins. Clinical, dietary, and biochemical characteristics at baseline were similar between UMs (all p > 0.05). After the polyphenol-rich diet, UMB vs. UMA participants showed a 2-fold higher improvement of zonulin levels (p for interaction = 0.033). Moreover, UMB vs. UMA participants were characterized for alterations in fatty acid metabolism, kynurenine pathway of tryptophan catabolism, and microbial metabolization of phenolic acids. These changes were correlated with the reduction of zonulin levels and modifications of gut microbes (increased Clostridiales, including, R. lactaris, and G. formicilis). In conclusion, urolithin-based metabotyping identified older adults with a higher improvement of IP after a polyphenol-rich diet. Our results reinforce the concept that UMs may contribute to tailor personalized nutrition interventions., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Recent studies suggest that also the non-critical form of COVID-19 infection may be associated with executive function impairments. However, it is not clear if they result from cognitive impairments or by COVID-19 infection per se. We aimed to investigate if patients in the post-acute stage of severe COVID-19 (PwCOVID), without manifest cognitive deficits, reveal impairments in performing dual-task (DT) activities compared to healthy controls (HS). We assessed balance in 31 PwCOVID vs. 30 age-matched HS by stabilometry and the Timed Up and Go (TUG) test with/without a cognitive DT. The DT cost (DTC), TUG test time and sway oscillations were recorded; correct cognitive responses (CCR) were calculated to evaluate cognitive performance. Results show a significant difference in overall DT performance between PwCOVID and HS in both stabilometry ( p < 0.01) and the TUG test ( p < 0.0005), although with similar DTCs. The main difference in the DTs between groups emerged in the CCR (effect size > 0.8). Substantially, PwCOVID gave priority to the motor task, leaving out the cognitive one, while HS performed both tasks simultaneously. Our findings suggest that PwCOVID, even without a manifest cognitive impairment, may present a deficit in executive function during DTs. These results encourage the use of DTs and CCR in PwCOVID.

Intense physical activity is often associated with undesirable physiological changes, including increased inflammation, transient immunodepression, increased susceptibility to infections, altered intestinal barrier integrity, and increased oxidative stress. Several trials suggested that probiotics supplementation may have beneficial effects on sport-associated gastro-intestinal and immune disorders. Recently, in a placebo-controlled human trial, the AminoAlta™ probiotic formulation (AApf) was demonstrated to increase the absorption of amino acids from pea protein, suggesting that the administration of AApf could overcome the compositional limitations of plant proteins. In this study, human cell line models were used to assess in vitro the potential capacity of AApf to protect from the physiological damages that an intense physical activity may cause. The obtained results revealed that the bacteria in the AApf have the ability to adhere to differentiated Caco-2 epithelial cell layer. In addition, the AApf was shown to reduce the activation of NF-κB in Caco-2 cells under inflammatory stimulation. Notably, this anti-inflammatory activity was enhanced in the presence of partially hydrolyzed plant proteins. The AApf also triggered the expression of cytokines by the THP-1 macrophage model in a dose-dependent manner. In particular, the expression of cytokines IL-1β, IL-6, and TNF-α was higher than that of the regulatory cytokine IL-10, resembling a cytokine profile characteristic of M1 phenotype, which typically intervene in counteracting bacterial and viral infections. Finally, AApf was shown to reduce transepithelial permeability and increase superoxide dismutase activity in the Caco-2 cell model. In conclusion, this study suggests that the AApf may potentially provide a spectrum of benefits useful to dampen the gastro-intestinal and immune detrimental consequences of an intense physical activity., Competing Interests: Authors LB, VV, RF, MM, and FW were employed by Sofar S.p.A., the company that commercialized the probiotic supplement investigated. RJ was employed by Increnovo LCC and a consultant to Sofar S.p.A. SG is a consultant to Sofar S.p.A. Sofar S.p.A. had a role in study design, data collection, and decision to publish. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Brunelli, De Vitis, Ferrari, Minuzzo, Fiore, Jäger, Taverniti and Guglielmetti.)

Secretory immunoglobulin A (SIgA) interaction with commensal bacteria conditions microbiota composition and function. However, mechanisms regulating reciprocal control of microbiota and SIgA are not defined. Bacteria-derived adenosine triphosphate (ATP) limits T follicular helper (Tfh) cells in the Peyer's patches (PPs) via P2X7 receptor (P2X7R) and thereby SIgA generation. Here we show that hydrolysis of extracellular ATP (eATP) by apyrase results in amplification of the SIgA repertoire. The enhanced breadth of SIgA in mice colonized with apyrase-releasing Escherichia coli influences topographical distribution of bacteria and expression of genes involved in metabolic versus immune functions in the intestinal epithelium. SIgA-mediated conditioning of bacteria and enterocyte function is reflected by differences in nutrient absorption in mice colonized with apyrase-expressing bacteria. Apyrase-induced SIgA improves intestinal homeostasis and attenuates barrier impairment and susceptibility to infection by enteric pathogens in antibiotic-induced dysbiosis. Therefore, amplification of SIgA by apyrase can be leveraged to restore intestinal fitness in dysbiotic conditions., Competing Interests: Declaration of interests F.G. is the founder of MV BioTherapeutics. F.G. and L.P. are inventors in a patent application covering the dysbiosis-correcting effect of apyrase., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

There is an increasing interest in investigating dietary strategies able to modulate the gut microbial ecosystem which, in turn, may play a key role in human health. Dietary fibers (DFs) are widely recognized as molecules with prebiotic effects. The main objective of this systematic review was to: (i) analyze the results available on the impact of DF intervention on short chain fatty acids (SCFAs) production; (ii) evaluate the interplay between the type of DF intervention, the gut microbiota composition and its metabolic activities, and any other health associated outcome evaluated in the host. To this aim, initially, a comprehensive database of literature on human intervention studies assessing the effect of confirmed and candidate prebiotics on the microbial ecosystem was developed. Subsequently, studies performed on DFs and analyzing at least the impact on SCFA levels were extracted from the database. A total of 44 studies from 42 manuscripts were selected for the analysis. Among the different types of fiber, inulin was the DF investigated the most ( n = 11). Regarding the results obtained on the ability of fiber to modulate total SCFAs, seven studies reported a significant increase, while no significant changes were reported in five studies, depending on the analytical methodology used. A total of 26 studies did not show significant differences in individual SCFAs, while the others reported significant differences for one or more SCFAs. The effect of DF interventions on the SCFA profile seemed to be strictly dependent on the dose and the type and structure of DFs. Overall, these results underline that, although affecting microbiota composition and derived metabolites, DFs do not produce univocal significant increase in SCFA levels in apparently healthy adults. In this regard, several factors (i.e., related to the study protocols and analytical methods) have been identified that could have affected the results obtained in the studies evaluated. Future studies are needed to better elucidate the relationship between DFs and gut microbiota in terms of SCFA production and impact on health-related markers.

Burning mouth syndrome (BMS) is defined as "idiopathic orofacial pain with intraoral burning or dysesthesia recurring daily for more than 2 hours per day and more than 3 months, without any identifiable causative lesions, with or without somatosensory changes" in International Classification of Orofacial Pain, 2020. Worldwide prevalence of BMS was estimated to be 1.73% in population-based studies, while female and elderly are at higher risk of BMS. The aim of this narrative review is to clarify the main etiopathogenetic factors of BMS investigated so far in the scientific literature. There is growing evidence of an important role of peripheral neuropathology in BMS, supported by immunohistochemical studies which have demonstrated a significant loss of epithelial and subepithelial nerve fibers. Other possible etiopathogenetic factors emerging from literature are laryngopharyngeal reflux and hormonal and salivary changes related to aging and menopause. Finally, the role of the oral microbiota in BMS has not yet been thoroughly investigated. Further studies are necessary to investigate the probably multifactorial etiopathogenesis of primary BMS, a pathology which has a serious impact on the quality of life of our patients, a disease we find ourselves treating without the adequate therapy and the necessary knowledge.

Purpose: Aging can be characterized by increased systemic low-grade inflammation, altered gut microbiota composition, and increased intestinal permeability (IP). The intake of polyphenol-rich foods is proposed as a promising strategy to positively affect the gut microbiota-immune system-intestinal barrier (IB) axis. In this context, we tested the hypothesis that a PR-dietary intervention would affect the presence of bacterial factors in the bloodstream of older adults., Methods: We collected blood samples within a randomized, controlled, crossover intervention trial in which older volunteers (n = 51) received a polyphenol-enriched and a control diet. We quantified the presence of bacterial DNA in blood by qPCR targeting the 16S rRNA gene (16S; bacterial DNAemia). Blood DNA was taxonomically profiled via 16S sequencing., Results: Higher blood 16S levels were associated with higher BMI and markers of IP, inflammation, and dyslipidemia. PR-intervention did not significantly change bacterial DNAemia in the older population (P = 0.103). Nonetheless, the beneficial changes caused by the polyphenol-enriched diet were greatest in participants with higher bacterial DNAemia, specifically in markers related to IP, inflammation and dyslipidemia, and in fecal bacterial taxa. Finally, we found that the bacterial DNA detected in blood mostly belonged to γ-Proteobacteria, whose abundance significantly decreased after the polyphenol-rich diet in subjects with higher bacterial DNAemia at baseline., Conclusions: This study shows that older subjects with higher bacterial DNAemia experienced a beneficial effect from a polyphenol-rich diet. Bacterial DNAemia may be a further relevant marker for the identification of target populations that could benefit more from a protective dietary treatment., Registration: This trial was retrospectively registered at www.isrctn.org (ISRCTN10214981) on April 28, 2017., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

COVID-19 leaves important sequelae in patients, not only in those who had the experience of a critical illness but also in patients with severe form. Understanding the impairments allows us to target rehabilitation to patients' real needs; balance impairments are an assumed sequela of COVID-19, but no study has specifically evaluated balance performance in these patients. Their performance was compared to that of patients with a pulmonary disease that leads to systemic diseases, such as patients with an acute exacerbation of chronic obstructive pulmonary disease (PwAECOPD), and of healthy subjects. A total of 75 subjects were assessed: 25 patients with COVID-19 (PwCOVID) with a severe form in the acute phase, 25 PwAECOPD and 25 healthy subjects sex- and age-matched. A stabilometric platform was used to evaluate static balance, both with eyes open and closed, while the dynamic balance was assessed with the Mini-BESTest and the Timed Up and Go test. When compared to healthy subjects, results showed that PwCOVID had worse performance in both static (P < 0.005) and dynamic (P < 0.0001) balance, with a large effect size in all measures (>0.8). Moreover, PwCOVID showed similar results to those of PwAECOPD. In conclusion, PwCOVID showed a balance deficit in both dynamic and static conditions. Therefore, as for PwAECOPD, they should require not only respiratory rehabilitation but also balance and mobility physiotherapy to prevent today's PwCOVID from becoming tomorrow's fallers., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

GPR120 (encoded by FFAR4 gene) is a receptor for long chain fatty acids, activated by ω-3 Polyunsaturated Fatty Acids (PUFAs), and expressed in many cell types. Its role in the context of colorectal cancer (CRC) is still puzzling with many controversial evidences. Here, we explored the involvement of epithelial GPR120 in the CRC development. Both in vitro and in vivo experiments were conducted to mimic the conditional deletion of the receptor from gut epithelium. Intestinal permeability and integrity of mucus layer were assessed by using Evans blue dye and immunofluorescence for MUC-2 protein, respectively. Microbiota composition, presence of lipid mediators and short chain fatty acids were analyzed in the stools of conditional GPR120 and wild type (WT) mice. Incidence and grade of tumors were evaluated in all groups of mice before and after colitis-associated cancer. Finally, GPR120 expression was analyzed in 9 human normal tissues, 9 adenomas, and 17 primary adenocarcinomas. Our work for the first time highlights the role of the receptor in the progression of colorectal cancer. We observed that the loss of epithelial GPR120 in the gut results into increased intestinal permeability, microbiota translocation and dysbiosis, which turns into hyperproliferation of epithelial cells, likely through the activation of β -catenin signaling. Therefore, the loss of GPR120 represents an early event of CRC, but avoid its progression as invasive cancer. these results demonstrate that the epithelial GPR120 receptor is essential to maintain the mucosal barrier integrity and to prevent CRC developing. Therefore, our data pave the way to GPR120 as an useful marker for the phenotypic characterization of CRC lesions and as new potential target for CRC prevention., (© 2022. The Author(s).)

Inflammation and immunity are linked to intestinal adenoma (IA) and colorectal cancer (CRC) development. The gut microbiota is associated with CRC risk. Epithelial barrier dysfunction can occur, possibly leading to increased intestinal permeability in CRC patients. We conducted a case-control study including 100 incident histologically confirmed CRC cases, and 100 IA and 100 healthy subjects, matched to cases by center, sex and age. We performed 16S rRNA gene analysis of blood and applied conditional logistic regression. Further analyses were based on negative binomial distribution normalization and Random Forest algorithm. We found an overrepresentation of blood 16S rRNA gene copies in colon cancer as compared to tumor-free controls. For high levels of gene copies, community diversity was higher in colon cancer cases than controls. Bacterial taxa and operational taxonomic unit abundances were different between groups and were able to predict CRC with an accuracy of 0.70. Our data support the hypothesis of a higher passage of bacteria from gastrointestinal tract to bloodstream in colon cancer. This result can be applied on non-invasive diagnostic tests for colon cancer control.

Gut microbial processing of dietary flaxseed (FS) contributes to its health benefits, but the relative effects of its bioactive components (lignans, omega-3 fatty acids, fiber) on the microbiota are unclear. We investigated the gut microbial compositional and functional responses to whole FS and its isolated components, FS oil (FSO) and secoisolariciresinol diglucoside (SDG) (precursor to microbial-derived enterolignans) to help understand their contribution to whole FS benefits. Cecum content and fecal samples were collected from C57BL/6 female mice fed a basal diet (AIN93G) or isocaloric diets containing 10% FS or 10% FS-equivalent amounts of FSO or SDG for 21 days. Cecal and fecal microbiota composition and predicted genomic functions, and their relationship with serum enterolignans were evaluated. Only FS modified the community structure. Shared- and diet-specific enriched taxa and functions were identified. Carbohydrate and protein processing functions were enriched in FS mice, and there was a positive correlation between select enriched taxa, encompassing fiber degraders and SDG metabolizers, and serum enterolignans. This was not observed in mice receiving isolated FSO and SDG, suggesting that FS fiber supports SDG microbial metabolism. In conclusion, the cooperative activities of a diverse microbiota are necessary to process FS components and, when administered at the amount present in FS, these components may act together to affect SDG-derived enterolignans production. This has implications for the use of FS, FSO and SDG in clinical practice., (Copyright © 2021. Published by Elsevier Inc.)

Background &aim: The MaPLE study was a randomized, controlled, crossover trial involving adults ≥60 y.o. (n = 51) living in a residential care facility during an 8-week polyphenol-rich (PR)-diet. Results from the MaPLE trial showed that the PR-diet reduced the intestinal permeability (IP) in older adults by inducing changes to gut microbiota (GM). The present work aimed at studying the changes in serum metabolome in the MaPLE trial, as a further necessary step to depict the complex crosstalk between dietary polyphenols, GM, and intestinal barrier., Methods: Serum metabolome was monitored using a semi-targeted UHPLC-MS/MS analysis. Metataxonomic analysis (16S rRNA gene profiling) of GM was performed on faecal samples. Clinical characteristics and serum levels of the IP marker zonulin were linked to GM and metabolomics data in a multi-omics network., Results: Compared to the control diet, the PR-diet increased serum metabolites related to polyphenols and methylxanthine intake. Theobromine and methylxanthines, derived from cocoa and/or green tea, were positively correlated with butyrate-producing bacteria (the order Clostridiales and the genera Roseburia, Butyricicoccus and Faecalibacterium) and inversely with zonulin. A direct correlation between polyphenol metabolites hydroxyphenylpropionic acid-sulfate, 2-methylpyrogallol-sulfate and catechol-sulfate with Butyricicoccus was also observed, while hydroxyphenylpropionic acid-sulfate and 2-methylpyrogallol-sulfate negatively correlated with Methanobrevibacter. The multi-omics network indicated that participant's age, baseline zonulin levels, and changes in Porphyromonadaceae abundance were the main factors driving the effects of a PR-diet on zonulin., Conclusion: Overall, these results reveal the complex relationships among polyphenols consumption, intestinal permeability, and GM composition in older adults, and they may be important when setting personalized dietary interventions for older adults., Trial Registration Number: ISRCTN10214981., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Dietary flaxseed (FS) and its components including FS oil (FSO), secoisolariciresinol diglucoside (SDG) and fiber, are processed by the gut microbiota. These data are in support of the article entitled "Discriminatory and cooperative effects within the mouse gut microbiota in response to flaxseed and its oil and lignan components", Journal of Nutritional Biochemistry [1]. Here we describe data generated by 16S rRNA sequencing of DNA obtained from cecum contents and feces of C57BL/6 female mice fed either a basal diet (BD, AIN93G), or isocaloric diets containing 10% FS, or 10% FS-equivalent amounts of FSO or SDG for 21 days. These include bacterial community composition and inferred KEGG pathways; the raw data are publicly available at the NCBI SRA database (BioProject ID PRJNA683934). Furthermore, this work includes detailed experimentation procedures, total bacterial counts (qPCR) in the cecum content and feces, and correlation analysis between a selected bacterial genus, Bacteroides and a predicted metabolic pathway. FS is utilized worldwide, especially for the prevention and/or treatment of diseases including cardiovascular diseases, diabetes and cancer. These data will be valuable as a reference to study different FS cultivars and SDG- or FSO- enriched products on the gut microbiota, to study gut microbial responses to FS and its components in different mouse strains and mammalian hosts to elucidate individualized effects, and to understand the importance of the gut microbiota for FS benefits., Competing Interests: All funding agencies have been listed in the Acknowledgments section above. EMC has received research support from Lallemand Health Solutions and Ocean Spray and has received consultant fees or speaker or travel support from Danone, Nestlé, and Lallemand Health Solutions (All are outside of this work). The authors declare that they have no known competing financial interests or personal relationships which have or could be perceived to have influenced the work reported in this article., (© 2021 The Authors. Published by Elsevier Inc.)

Probiotic microorganisms may benefit the host by influencing diverse physiological processes, whose nature and underlying mechanisms are still largely unexplored. Animal models are a unique tool to understand the complexity of the interactions between probiotic microorganisms, the intestinal microbiota, and the host. In this regard, in this pilot study, we compared the effects of 5-day administration of three different probiotic bacterial strains ( Bifidobacterium bifidum MIMBb23sg, Lactobacillus helveticus MIMLh5, and Lacticaseibacillus paracasei DG) on three distinct murine intestinal sites (ileum, cecum, and colon). All probiotics preferentially colonized the cecum and colon. In addition, probiotics reduced in the ileum and increased in the cecum and colon the relative abundance of numerous bacterial taxonomic units. MIMBb23sg and DG increased the inducible nitric oxide synthase (iNOS) in the ileum, which is involved in epithelial homeostasis. In addition, MIMBb23sg upregulated cytokine IL-10 in the ileum and downregulated the cyclooxygenase COX-2 in the colon, suggesting an anti-inflammatory/regulatory activity. MIMBb23sg significantly affected the expression of the main gene involved in serotonin synthesis (TPH1) and the gene coding for the serotonin reuptake protein (SERT) in the ileum and colon, suggesting a potential propulsive effect toward the distal part of the gut, whereas the impact of MIMLh5 and DG on serotonergic genes suggested an effect toward motility control. The three probiotics decreased the expression of the permeability marker zonulin in gut distal sites. This preliminary in vivo study demonstrated the safety of the tested probiotic strains and their common ability to modulate the intestinal microbiota. The probiotics affected host gene expression in a strain-specific manner. Notably, the observed effects in the gut were site dependent. This study provides a rationale for investigating the effects of probiotics on the serotonergic system, which is a topic still widely unexplored., Competing Interests: SG was consultant of Sofar S.p.A., the private company that commercializes strains L. paracasei DG and B. bifidum MIMBb23g. WF was an employee of Sofar S.p.A. SG and VT received royalties from the sale of the strain B. bifidum MIMBb23sg. SG and VT received royalties from the sale of the strain L. helveticus MIMLh5. The remanining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Taverniti, Cesari, Gargari, Rossi, Biddau, Lecchi, Fiore, Arioli, Toschi and Guglielmetti.)

Background: Hip fractures should not be considered as a single, homogeneous condition. Various determinants of functional recovery of patients with hip fracture have been proposed, such as age or type of fracture. The aim of this study was to determine if patients with an intertrochanteric fracture (ITF) had lower functional recovery characteristics with respect to those with a femoral neck fracture (FNF)., Methods: A retrospective study was carried out on 531 elderly patients with hip fracture, surgically treated, admitted to a rehabilitation institute between December 1, 2014, and December 31, 2017. Patients underwent an individualized rehabilitation program for improving their physical function. The outcome measures of this study were the length of stay (LOS) in the rehabilitation institute, the Functional Independence Measure (FIM) gain, i.e. the difference in FIM score between discharge and admission, and the FIM efficiency, which represents the daily gain. Patients were also stratified by age categories and by different levels of functional independence, as evaluated with the FIM total score at baseline., Results: Age (p<0.05), LOS (p<0.005) and rehabilitation outcomes (FIM score and efficiency; p<0.05) differed significantly between patients with FNF and ITF. In particular, patients with ITF were older and more dependent at baseline. Moreover, they showed a lower FIM efficiency with respect to patients with FNF (p<0.05), which obtained a similar improvement in FIM total score of about 26 points but were discharged on average 3 days before. With aging, FIM efficiency decreased, while LOS increased. Patients highly dependent at baseline (FIM total score <40) had the lowest FIM gain and efficiency and the longest LOS respect to the more independent patients. Moreover, in older (85+ years) and in higher dependent patients the differences in the outcome measures between ITF and FNF were reduced., Conclusions: The type of hip fracture could affect the LOS and FIM efficiency of younger and more independent patients with hip fracture during inpatient rehabilitation. In particular, patients with ITF require a longer rehabilitation period to achieve a similar functional gain as those with FNF., Competing Interests: Declaration of Competing Interest None., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The increased presence of bacteria in blood is a plausible contributing factor in the development and progression of aging-associated diseases. In this context, we performed the quantification and the taxonomic profiling of the bacterial DNA in blood samples collected from forty-three older subjects enrolled in a nursing home. Quantitative PCR targeting the 16S rRNA gene revealed that all samples contained detectable amounts of bacterial DNA with a concentration that varied considerably between subjects. Correlation analyses revealed that the bacterial DNAemia (expressed as concentration of 16S rRNA gene copies in blood) significantly associated with the serum levels of zonulin, a marker of intestinal permeability. This result was confirmed by the analysis of a second set of blood samples collected from the same subjects. 16S rRNA gene profiling revealed that most of the bacterial DNA detected in blood was ascribable to the phylum Proteobacteria with a predominance of the genus Pseudomonas. Several control samples were also analyzed to assess the influence of contaminant bacterial DNA potentially originating from reagents and materials. The data reported here suggest that para-cellular permeability of epithelial (and, potentially, endothelial) cell layers may play an important role in bacterial migration into the bloodstream. Bacterial DNAemia is likely to impact on several aspects of host physiology and could underpin the development and prognosis of various diseases in older subjects.

The use of DNA damage as marker of oxidative stress, metabolic dysfunction and age-related diseases is debated. The present study aimed at assessing the level of DNA damage (evaluated as DNA strand-breaks, endogenous and oxidatively-induced DNA damage) in a group of older subjects with intestinal permeability enrolled within the MaPLE (Gut and Blood Microbiomics for Studying the Effect of a Polyphenol-Rich Dietary Pattern on Intestinal Permeability in the Elderly) intervention trial, to evaluate its association with clinical, metabolic and dietary markers. DNA damage in peripheral blood mononuclear cells was assessed by the comet assay in 49 older subjects participating in the study. Clinical and metabolic markers, markers of inflammation, vascular function and intestinal permeability were determined in serum. Food intake was estimated by weighted food diaries. On the whole, a trend towards higher levels of DNA damage was observed in men compared to women ( p = 0.071). A positive association between DNA damage and clinical/metabolic markers (e.g., uric acid, lipid profile) and an inverse association with dietary markers (e.g., vitamin C, E, B 6 , folates) were found and differed based on sex. By considering the importance of DNA stability during aging, the results obtained on sex differences and the potential role of dietary and metabolic factors on DNA damage underline the need for further investigations in a larger group of older adults to confirm the associations found and to promote preventive strategies.

Background & Aim: Increased intestinal permeability (IP) can occur in older people and contribute to the activation of the immune system and inflammation. Dietary interventions may represent a potential strategy to reduce IP. In this regard, specific food bioactives such as polyphenols have been proposed as potential IP modulator due to their ability to affect several critical targets and pathways that control IP. The trial aimed to test the hypothesis that a polyphenol-rich dietary pattern can decrease serum zonulin levels, an IP surrogate marker involved in tight junction modulation, and can beneficially alter the intestinal microbiota, and IP-associated biochemical and clinical markers in older subjects., Methods: A randomised, controlled, cross-over intervention trial was performed. Sixty-six subjects (aged ≥ 60 y) with increased IP based on serum zonulin levels, were randomly allocated to one of the two arms of the intervention consisting of a control diet (C-diet) vs. a polyphenol-rich diet (PR-diet). Each intervention was 8-week long and separated by an 8-week wash out period. At the beginning and at the end of each intervention period, serum samples were collected for the quantification of zonulin and other biological markers. Faecal samples were also collected to investigate the intestinal microbial ecosystem. In addition, anthropometrical/physical/biochemical parameters and food intake were evaluated., Results: Fifty-one subjects successfully completed the intervention and a high compliance to the dietary protocols was demonstrated. Overall, polyphenol intake significantly increased from a mean of 812 mg/day in the C diet to 1391 mg/day in the PR-diet. Two-way analysis of variance showed a significant effect of treatment (p = 0.008) and treatment × time interaction (p = 0.025) on serum zonulin levels, which decreased after the 8-week PR-diet. In addition, a treatment × time interaction was observed showing a reduction of diastolic blood pressure (p = 0.028) following the PR-diet, which was strongest in those not using antihypertensive drugs. A decrease in both diastolic (p = 0.043) and systolic blood pressure (p = 0.042) was observed in women. Interestingly, a significant increase in fibre-fermenting and butyrate-producing bacteria such as the family Ruminococcaceae and members of the genus Faecalibacterium was observed following the PR intervention. The efficacy of this dietary intervention was greater in subjects with higher serum zonulin at baseline, who showed more pronounced alterations in the markers under study. Furthermore, zonulin reduction was also stronger among subjects with higher body mass index and with insulin resistance at baseline, thus demonstrating the close interplay between IP and metabolic features., Conclusions: These data show, for the first time, that a PR-diet can reduce serum zonulin levels, an indirect marker of IP. In addition, PR-diet reduced blood pressure and increased fibre-fermenting and butyrate-producing bacteria. These findings may represent an initial breakthrough for further intervention studies evaluating possible dietary treatments for the management of IP, inflammation and gut function in different target populations. THIS STUDY WAS REGISTERED AT WWW.ISRCTN., Org as: ISRCTN10214981., Competing Interests: Conflict of interest The authors declare no conflicts of interest., (Copyright © 2020 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)

Emerging evidence indicates that gut microbiota affect the response to anticancer therapies by modulating the host immune system. In this study, we investigated the impact of gut microbiota on immune-mediated trastuzumab antitumor efficacy in preclinical models of HER2-positive breast cancer and in 24 patients with primary HER2-positive breast cancer undergoing trastuzumab-containing neoadjuvant treatment. In mice, the antitumor activity of trastuzumab was impaired by antibiotic administration or fecal microbiota transplantation from antibiotic-treated donors. Modulation of the intestinal microbiota was reflected in tumors by impaired recruitment of CD4 + T cells and granzyme B-positive cells after trastuzumab treatment. Antibiotics caused reductions in dendritic cell (DC) activation and the release of IL12p70 upon trastuzumab treatment, a mechanism that was necessary for trastuzumab effectiveness in our model. In patients, lower α-diversity and lower abundance of Lachnospiraceae, Turicibacteraceae, Bifidobacteriaceae , and Prevotellaceae characterized nonresponsive patients (NR) compared with those who achieved pathologic complete response (R), similar to antibiotic-treated mice. The transfer of fecal microbiota from R and NR into mice bearing HER2-positive breast cancer recapitulated the response to trastuzumab observed in patients. Fecal microbiota β-diversity segregated patients according to response and positively correlated with immune signature related to interferon (IFN) and NO2-IL12 as well as activated CD4 + T cells and activated DCs in tumors. Overall, our data reveal the direct involvement of the gut microbiota in trastuzumab efficacy, suggesting that manipulation of the gut microbiota is an optimal future strategy to achieve a therapeutic effect or to exploit its potential as a biomarker for treatment response. SIGNIFICANCE: Evidence of gut microbiota involvement in trastuzumab efficacy represents the foundation for new therapeutic strategies aimed at manipulating commensal bacteria to improve response in trastuzumab-resistant patients. See related commentary by Sharma, p. 1937 GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/8/2195/F1.large.jpg., (©2021 American Association for Cancer Research.)

SARS-CoV-2 is a newly emerging virus that currently lacks curative treatments. Lactoferrin (LF) is a naturally occurring non-toxic glycoprotein with broad-spectrum antiviral, immunomodulatory and anti-inflammatory effects. In this study, we assessed the potential of LF in the prevention of SARS-CoV-2 infection in vitro. Antiviral immune response gene expression was analyzed by qRT-PCR in uninfected Caco-2 intestinal epithelial cells treated with LF. An infection assay for SARS-CoV-2 was performed in Caco-2 cells treated or not with LF. SARS-CoV-2 titer was determined by qRT-PCR, plaque assay and immunostaining. Inflammatory and anti-inflammatory cytokine production was determined by qRT-PCR. LF significantly induced the expression of IFNA1 , IFNB1 , TLR3 , TLR7 , IRF3 , IRF7 and MAVS genes. Furthermore, LF partially inhibited SARS-CoV-2 infection and replication in Caco-2 intestinal epithelial cells. Our in vitro data support LF as an immune modulator of the antiviral immune response with moderate effects against SARS-CoV-2 infection.

Purpose: Subclinical vitamin D (vitD) deficiency enhances the predisposition to a myriad of acute and chronic pathologies in many people worldwide. Due to the scarcity of vitD-rich foods, the consumption of supplements or fortified foods can be required to maintain healthy serum levels of 25-hydroxyvitamin D [25(OH)D], and the major circulating form of vitD that is commonly measured in serum to determine the vitD status. Since the vitD absorption seems to resemble that of lipids, improved emulsification in the gut could favor vitD permeation through the enterocyte membrane. Contextually, we hypothesized that a microorganism with cholecalciferol (vitD3)-solubilization properties may potentially result in enhanced serum vitD levels., Methods and Results: Six probiotic strains were screened for their ability to create a stable suspension of vitD3 in water: Lacticaseibacillus paracasei DG, L. paracasei LPC-S01, L. paracasei Shirota, L. rhamnosus GG, Limosilactobacillus reuteri DSM 17938, and Lactobacillus acidophilus LA5. The DG strain displayed the strongest vitD3 solubilization ability and, consequently, were used in an in vivo trial where a commercial preparation of vitD3 in refined olive oil was administered by gavage to CD-1 mice with or without the concurrent administration of L. paracasei DG. ELISA measurements showed that the DG strain significantly increased the serum levels of 25(OH) D when administered once a day for 1 week in association with the vitD3 supplement., Conclusion: This preliminary pre-clinical study suggests that the combined administration of L. paracasei DG with an oil-based cholecalciferol supplement could contribute to the maintenance of the adequate 25(OH) D serum levels in people at risk of vitD deficiency., Competing Interests: Competing interestsSofar S.p.A. is a private company that commercializes probiotic supplements containing L. paracasei DG. Walter Fiore and Valerio De Vitis are employees of Sofar S.p.A. Simone Guglielmetti is consultant of Sofar S.p.A. Other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© The Author(s) 2021.)

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causing the ongoing global pandemic of coronavirus disease 2019 (COVID-19), which primarily manifests with respiratory distress and may also lead to symptoms associated with the gastrointestinal tract. Probiotics are living microorganisms that have been shown to confer immune benefits. In this study, we investigated the immunomodulatory effects and anti-SARS-CoV-2 activity of three different Lacticaseibacillus probiotic strains, either alone or in combination with lactoferrin, using the intestinal epithelial Caco-2 cell line. Our results revealed that the Lacticaseibacillus paracasei DG strain significantly induced the expression of genes involved in protective antiviral immunity and prevented the expression of proinflammatory genes triggered by SARS-CoV-2 infection. Moreover, L. paracasei DG significantly inhibited SARS-CoV-2 infection in vitro. L. paracasei DG also positively affected the antiviral immune activity of lactoferrin and significantly augmented its anti-SARS-CoV-2 activity in Caco-2 intestinal epithelial cells. Overall, our work shows that the probiotic strain L. paracasei DG is a promising candidate that exhibits prophylactic potential against SARS-CoV-2 infection.

Polyphenols have great potential in regulating intestinal health and ameliorating pathological conditions related to increased intestinal permeability (IP). However, the efficacy of dietary interventions with these phytochemicals may significantly be influenced by interindividual variability factors affecting their bioavailability and consequent biological activity. In the present study, urine samples collected from older subjects undergoing a crossover intervention trial with polyphenol-rich foods were subjected to metabolomics analysis for investigating the impact of increased IP on the bioavailability of polyphenols. Interestingly, urinary levels of phase II and microbiota-derived metabolites were significantly different between subjects with healthier intestinal barrier integrity and those with increased IP disruption. Our results support that this IP-dependent impaired bioavailability of polyphenols could be attributed to disturbances in the gut microbial metabolism and phase II methylation processes. Furthermore, we also observed that microbiota-derived metabolites could be largely responsible for the biological activity elicited by dietary polyphenols against age-related disrupted IP.

Background: Severe early-onset erythroderma and gut inflammation, with massive tissue infiltration of oligoclonal activated T cells are the hallmark of Omenn syndrome (OS)., Objective: The impact of altered gut homeostasis in the cutaneous manifestations of OS remains to be clarified., Methods: We analyzed a cohort of 15 patients with OS and the 129Sv/C57BL/6 knock-in Rag2 R229Q/R229Q (Rag2 R229Q ) mouse model. Homing phenotypes of circulating lymphocytes were analyzed by flow cytometry. Inflammatory cytokines and chemokines were examined in the sera by ELISA and in skin biopsies by immunohistochemistry and in situ RNA hybridization. Experimental colitis was induced in mice by dextran sulfate sodium salt., Results: We show that memory/activated T cells from patients with OS and from the Rag2 R229Q mouse model of OS abundantly express the skin homing receptors cutaneous lymphocyte associated antigen and CCR4 (Ccr4), associated with high levels of chemokine C-C motif ligands 17 and 22. Serum levels of LPS are also elevated. A broad T h 1/T h 2/T h 17 inflammatory signature is detected in the periphery and in the skin. Increased Tlr4 expression in the skin of Rag2 R229Q mice is associated with enhanced cutaneous inflammation on local and systemic administration of LPS. Likewise, boosting colitis in Rag2 R229Q mice results in increased frequency of Ccr4 + splenic T cells and worsening of skin inflammation, as indicated by epidermal thickening, enhanced epithelial cell activation, and dermal infiltration by T h 1 effector T cells., Conclusions: These results support the existence of an interplay between gut and skin that can sustain skin inflammation in OS., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

The evaluation of food intake in older subjects is crucial in order to be able to verify adherence to nutritional recommendations. In this context, estimation of the intake of specific dietary bioactives, such as polyphenols, although particularly challenging, is necessary to plan possible intervention strategies to increase their intake. The aims of the present study were to: (i) evaluate the nutritional composition of dietary menus provided in a residential care setting; (ii) estimate the actual intake of nutrients and polyphenols in a group of older subjects participating in the MaPLE study; and (iii) investigate the impact of an eight-week polyphenol-rich dietary pattern, compared to an eight-week control diet, on overall nutrient and polyphenol intake in older participants. The menus served to the participants provided ~770 mg per day of total polyphenols on average with small variations between seasons. The analysis of real consumption, measured using weighed food diaries, demonstrated a lower nutrient (~20%) and polyphenol intake (~15%) compared to that provided by the menus. The feasibility of dietary patterns that enable an increase in polyphenol intake with putative health benefits for age-related conditions is discussed, with a perspective to developing dietary guidelines for this target population.

Producers of probiotic products are legally required to indicate on the label only the minimum numbers of viable microorganisms at the end of shelf life expressed as colony-forming units (CFUs). Label specifications, however, describe only a fraction of the actual microbiological content of a probiotic formulation. This paper describes the microbiological components of a probiotic product that are not mentioned on the label, such as the actual number of CFUs, the presence of viable cells that cannot generate colonies on agar plates, and the abundance of dead cells. These "hidden" microbial fractions in probiotic products, the abundance of which may change during the shelf life, can promote biological responses in the host. Therefore, they should not be ignored because they may influence the efficacy and can be relevant for immunocompromised or fragile consumers. In conclusion, we propose the minimum requirements for microbiological characterization of probiotic products to be adopted for label specifications and clinical studies.

Oral consumption of probiotics is practical and can be an effective solution to preserve vaginal eubiosis. Here, we studied the ability of orally administered Lactobacillus paracasei LPC-S01 (DSM 26760) to affect the composition of the vaginal microbiota and colonize the vaginal mucosa in nondiseased adult women. A total of 40 volunteers took oral probiotic (24 billion CFU) or placebo capsules daily for 4 weeks, and after a 4-week washout, they switched to placebo or probiotic capsules according to the crossover design. A total of 23 volunteers completed the study according to the protocol. Before and after capsule ingestion, vaginal swabs were collected for qPCR quantification to detect L. paracasei LPC-S01 and for 16S rRNA gene sequencing. Vaginal swabs were grouped according to their bacterial taxonomic structure into nine community state types (CSTs), four of which were dominated by lactobacilli. Lactobacillus paracasei LPC-S01 was detected in the vagina of two participants. Statistical modeling (including linear mixed-effects model analysis) demonstrated that daily intake of probiotic capsules reduced the relative abundance of Gardnerella spp. Quantitative PCR with Gardnerella vaginalis primers confirmed this result. Considering the pathogenic nature of G. vaginalis, these results suggest a potential positive effect of this probiotic capsule on the vaginal microbial ecosystem., (© FEMS 2020.)