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The human body represents the habitat of trillions of symbiotic microorganisms, collectively known as human microbiota, approximately half of which residing in the gut. The development of next-generation sequencing techniques has boosted the profiling of human microbiota in recent years. A growing body of evidence seems to support a strict relationship between the disruption of the mutualistic relationship between the microbiota and the host (i.e., dysbiosis) and the development of several diseases, including breast malignancies. Breast cancer still represents the most frequent cause of cancer-related death in women. Its complex relationship with gut microbiota is the object of a growing body of evidence. In fact, the interaction with the host immune system and a direct impact of gut microbiota on estrogen, lipid and polyphenols metabolism, seem to potentially affect breast tumor development, progression and response to treatments. In this review, in an attempt to help oncologists navigating this rapidly-evolving research field, we provide an essential overview on the taxonomy, main analytical techniques and terminology most commonly adopted. We discuss what is currently known regarding the interaction between gut microbiota and breast cancer and potential efforts to harness this complex interplay for therapeutic purposes, and revise main ongoing studies. We also briefly provide an overview on breast cancer intratumoral microbiota and its potential role beyond gut microbiota., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Francesco Schettini reports honoraria from Novartis, Gilead and Daiichy-Sankyo for educational events/materials and travel expenses from Novartis, Gilead and Daiichy-Sankyo. Daniele Generali declares personal fees for educational events by Novartis, Lilly, Pfizer, Daiichy-Sankyo, Roche; research funds from Astrazeneca, Novartis and LILT. The other authors have nothing to declare., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Chronic Kidney Disease (CKD) is characterized by a methionine-related metabolic disorder involving reduced plasma levels of hydrogen sulfide (H 2 S) and increased lanthionine. The gut microbiota influences methionine metabolism, potentially impacting sulfur metabolite dysfunctions in CKD. We evaluated whether gut microbiota dysbiosis contributes to H 2 S and lanthionine metabolic alterations in CKD., Methods: The gut microbiota of 88 CKD patients (non-dialysis, hemodialysis, and transplant patients) and 26 healthy controls were profiled using 16 S-amplicon sequencing. H 2 S and lanthionine concentrations were measured in serum and fecal samples using the methylene blue method and LC-MS/MS, respectively., Results: The CKD population exhibited a tenfold increase in serum lanthionine associated with kidney dysfunction. Despite lanthionine retention, hemodialysis and transplant patients had significantly lower serum H 2 S than healthy controls. Fecal H 2 S levels were not altered or related to bloodstream H 2 S concentrations. Conversely, fecal lanthionine was significantly increased in CKD compared to healthy controls and associated with kidney dysfunction. Microbiota composition varied among CKD groups and healthy controls, with the greatest dissimilarity observed between hemodialysis and transplant patients. Changes relative to the healthy group included uneven Ruminococcus gnavus distribution (higher in transplant patients and lower in non-dialysis CKD patients), reduced abundance of the short-chain fatty acid-producing bacteria Alistipes indistinctus and Coprococcus eutactus among transplant patients, and depleted Streptococcus salivarius in non-dialysis CKD patients. A higher abundance of Methanobrevibacter smithii, Christensenella minuta, and Negativibacillus massiliensis differentiated hemodialysis patients from controls. No correlation was found between differentially abundant species and the metabolic profile that could account for the H 2 S and lanthionine alterations observed., Conclusions: The metabolic deregulation of H 2 S and lanthionine observed in the study was not associated with alterations in the gut microbiota composition in CKD patients. Further research on microbial sulfur pathways may provide a better understanding of the role of gut microbiota in maintaining H 2 S and lanthionine homeostasis., (© 2024. The Author(s).)

Purpose: This study aimed to explore metabolic tumor volume (tMTV) as assessed 18F-fluorodeoxyglucose positron emission tomography-computed tomography (18F-FDG-PET/CT), and understand its biological meaning in patients with NSCLC exposed to immune checkpoint blockers(ICBs)., Experimental Design: In this study, patients with advanced NSCLC and a positive PET scan within 42 days of first line treatment were enrolled in 11 institutions across 4 countries. Total MTV (tMTV) was analyzed, with a 42% SUVmax threshold. Survival was analyzed according to high tMTV (≥ median). Plasma proteomic profile, whole exome, transcriptome and other analysis were performed on monocentric cohorts to explore its biological correlates., Results: Of the 518 patients included, 167 received ICBs, 257 had chemotherapy plus ICBs, and 94 had chemotherapy. Median tMTV was 99 cm3. Median overall survival (OS) for patients with high tMTV treated with ICBs was 11.4 months vs 29.6 months (P<0.0012) for those with low tMTV. In patients receiving chemotherapy-ICB tMTV did not correlate with OS (P=0.099). In patients with PD-L1≥1% and high tMTV, chemotherapy-ICB combination was associated with longer OS compared with ICBs alone (20 vs 11.4 months,p=0.026), while no survival differences observed in low tMTV group. High tMTV correlated (and its detrimental effect seems to be driven by) a specific proteomic profile and increase in genomic instability., Conclusion: Our analysis indicates high tTMV is linked to an increase in systemic inflammation, specific cytokines production and chromosomal instability. tTMV may serve as one of the biomarker to select the best upfront strategy in patients with PD-L1 positive advanced NSCLC.

Besides suppressing immune responses, regulatory T cells (Tregs) maintain tissue homeostasis and control systemic metabolism. Whether iron is involved in Treg-mediated tolerance is completely unknown. Here, we showed that the transferrin receptor CD71 was upregulated on activated Tregs infiltrating human liver cancer. Mice with a Treg-restricted CD71 deficiency spontaneously developed a scurfy-like disease, caused by impaired perinatal Treg expansion. CD71-null Tregs displayed decreased proliferation and tissue-Treg signature loss. In perinatal life, CD71 deficiency in Tregs triggered hepatic iron overload response, characterized by increased hepcidin transcription and iron accumulation in macrophages. Lower bacterial diversity, and reduction of beneficial species, were detected in the fecal microbiota of CD71 conditional knockout neonates. Our findings indicate that CD71-mediated iron absorption is required for Treg perinatal expansion and is related to systemic iron homeostasis and bacterial gut colonization. Therefore, we hypothesize that Tregs establish nutritional tolerance through competition for iron during bacterial colonization after birth.

The gut microbiota influences the clinical responses of cancer patients to immunecheckpoint inhibitors (ICIs). However, there is no consensus definition of detrimental dysbiosis. Based on metagenomics (MG) sequencing of 245 non-small cell lung cancer (NSCLC) patient feces, we constructed species-level co-abundance networks that were clustered into species-interacting groups (SIGs) correlating with overall survival. Thirty-seven and forty-five MG species (MGSs) were associated with resistance (SIG1) and response (SIG2) to ICIs, respectively. When combined with the quantification of Akkermansia species, this procedure allowed a person-based calculation of a topological score (TOPOSCORE) that was validated in an additional 254 NSCLC patients and in 216 genitourinary cancer patients. Finally, this TOPOSCORE was translated into a 21-bacterial probe set-based qPCR scoring that was validated in a prospective cohort of NSCLC patients as well as in colorectal and melanoma patients. This approach could represent a dynamic diagnosis tool for intestinal dysbiosis to guide personalized microbiota-centered interventions., Competing Interests: Declaration of interests L. Zitvogel is founder of everImmune and its SAB President. L.D. is an everImmune SAB member. R.D. is a scientific co-founder of everImmune. L. Zitvogel received a research contract from Kaleido, 9 meters/Innovate Pharma, and is currently sponsored by Pileje. G.Z. received a research grant from Fondation Roche; received fees from Roche, MSD, BMS, and Astra Zeneca; and is a consultant for Da Volterra & Inventiva. E.D. reports grants and personal fees from Roche Genentech, grants from Boehringer, grants from AstraZeneca, grants and personal fees from Merck Serono, grants from BMS, and grants from MSD. E.D. is founder of Graegis/Alys. M.L.I., L.B., and A.F. are employees and shareholders of Bio-Me. P.D. had consulting and advisory roles for AstraZeneca, Bristol-Myers Squibb, Boehringer Ingelheim, Celgene, Daiichi Sankyo, Eli Lilly, Merck, Novartis, Pfizer, prIME Oncology, Peer CME, Roche, and Samsung, as well as honoraria from AstraZeneca, Bristol-Myers Squibb, Boehringer Ingelheim, Celgene, Eli Lilly, Merck, Novartis, Pfizer, prIME Oncology, Peer CME, Roche, and Samsung. P.D. ran clinical trials as principal or co-investigator for AstraZeneca, Bristol-Myers Squibb, Boehringer Ingelheim, Eli Lilly, Merck, Novartis, Pfizer, Roche, MedImmune, Sanofi-Aventis, Taiho Pharma, Novocure, and Daiichi Sankyo and received travel, accommodation, and expenses from AstraZeneca, Roche, Novartis, prIME Oncology, and Pfizer. M.F. and R.K.W. were supported by the Seerave Foundation. R.K.W. received unrestricted research grants from Takeda, J&J, Ferring, and Tramedico; speaker’s fee from AbbVie, MSD., and Boston Scientific; and has acted as consultant for Takeda Pharmaceuticals. C.C. reported personal fees from Amgen, Bayer, Merck Serono, MSD, Nordic Pharma, Roche, Pierre Fabre, Servier, and Takeda and research grants from Bayer, Merck Serono, Pierre Fabre, Seagen, Servier, and Tempus. F.G. received honoraria from Amgen, Sanofi, Merk Serono, MSD, BMS, and Astra Zeneca; had a consultancy or advisory role for Roche and Enterome; and received direct research fundings from Roche, Enterome, Astra Zeneca, and Servier and traveling supports from Servier, Amgen, and Roche. L. Zaho is cofounder of Notitia Biotechnologies. F.B. institutional interest AbbVie, ACEA, Amgen, Astra Zeneca, Bayer, Bristol-Myers Squibb, Boehringer Ingelheim, EISAI, Eli Lilly Oncology, F. Hoffmann–La Roche Ltd, Genentech, Ipsen, Ignyta, Innate Pharma, Loxo, Novartis, MedImmune, Merck, Mirati, MSD, Pierre Fabre, Pfizer, Sanofi-Aventis, and Takeda.art, (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Cyclin-dependent kinase (CDK)4/6-inhibitors with endocrine therapy represent the standard of treatment of hormone receptor-positive(HR+)/human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer (MBC). Gut microbiota seems to predict treatment response in several tumour types, being directly implied in chemotherapy resistance and development of adverse effects. No evidence is available on gut microbiota impact on efficacy of HR+ breast cancer treatment., Patients and Methods: We assessed the potential association among faecal microbiota and therapeutic efficacy of CDK4/6-inhibitors on 14 MBC patients classified as responders (R) and non-responders (NR) according to progression-free survival. A stool sample was collected at baseline and V3-V4 16S targeted sequencing was employed to assess its bacterial composition. Statistical associations with R and NR were studied., Results: No significant differences were observed between R and NR in terms of α-/β-diversity at the phylum and species level. Machine-learning (ML) algorithms evidenced four bacterial species as a discriminant for R (Bifidobacterium longum, Ruminococcus callidus) and NR (Clostridium innocuum, Schaalia odontolytica), and an area under curve (AUC) of 0.946 after Random Forest modelling. Network analysis evidenced two major clusters of bacterial species, named Species Interacting Groups (SIG)1-2, with SIG1 harbouring 75% of NR-related bacterial species, and SIG2 regrouping 76% of R-related species (p < 0.001). Cross-correlations among several patients' circulating immune cells or biomarkers and bacterial species' relative abundances showed associations with potential prognostic implications., Conclusions: Our results provide initial insights into the gut microbiota involvement in sensitivity and/or resistance to CDK4/6-inhibitors + endocrine therapy in MBC. If confirmed in larger trials, several microbiota manipulation strategies might be hypothesised to improve response to CDK4/6-inhibitors., Competing Interests: Declaration of Competing Interest FS declares travel expenses by Novartis and Gilead and personal fees for educational events by Novartis, Daiichy-Sankyo and Gilead. DG declares personal fees for educational events by Novartis, Lilly, Pfizer, Daiichy-Sankyo, Roche; research funds from Astrazeneca, Novartis and LILT. GC declares reports advisory/consulting fees from Seagen, Roche, Novartis, Eli Lilly, Daiichi Sankyo, AstraZeneca, Pfizer, Sanofi, Pierre Fabre and Gilead, and fees for non-CME services (e.g. speakers' bureaus) from Eli Lilly, Pfizer Inc and Daiichi Sankyo. The other authors have nothing to declare., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Formyl peptide receptor-1 (FPR1) is a pattern recognition receptor that is mostly expressed by myeloid cells. In patients with colorectal cancer (CRC), a loss-of-function polymorphism (rs867228) in the gene coding for FPR1 has been associated with reduced responses to chemotherapy or chemoradiotherapy. Moreover, rs867228 is associated with accelerated esophageal and colorectal carcinogenesis. Here, we show that dendritic cells from Fpr1 -/- mice exhibit reduced migration in response to chemotherapy-treated CRC cells. Moreover, Fpr1 -/- mice are particularly susceptible to chronic ulcerative colitis and colorectal oncogenesis induced by the mutagen azoxymethane followed by oral dextran sodium sulfate, a detergent that induces colitis. These experiments were performed after initial co-housing of Fpr1 -/- mice and wild-type controls, precluding major Fpr1-driven differences in the microbiota. Pharmacological inhibition of Fpr1 by cyclosporin H also tended to increase intestinal oncogenesis in mice bearing the Apc Min mutation, and this effect was reversed by the anti-inflammatory drug sulindac. We conclude that defective FPR1 signaling favors intestinal tumorigenesis through the modulation of the innate inflammatory/immune response., Competing Interests: No potential conflict of interest was reported by the author(s)., (© 2023 The Author(s). Published with license by Taylor & Francis Group, LLC.)

Gut microorganisms and the products of their metabolism thoroughly affect host brain development, function and behavior. Since alterations of brain plasticity and cognition have been demonstrated upon motor, sensorial and social enrichment of the housing conditions, we hypothesized that gut microbiota and metabolome could be altered by environmental stimuli, providing part of the missing link among environmental signals and brain effects. In this preliminary study, metagenomic and metabolomic analyses of mice housed in different environmental conditions, standard and enriched, identify environment-specific microbial communities and metabolic profiles. We show that mice housed in an enriched environment have distinctive microbiota composition with a reduction in gut bacterial richness and biodiversity and are characterized by a metabolomic fingerprint with the increase of formate and acetate and the decrease of bile salts. We demonstrate that mice treated with a mixture of formate and acetate recapitulate some of the brain plasticity effects modulated by environmental enrichment, such as hippocampal neurogenesis, neurotrophin production, short-term plasticity and cognitive behaviors, that can be further exploited to decipher the mechanisms involved in experience-dependent brain plasticity., (© 2022. The Author(s).)

Gut dysbiosis has been associated with intestinal and extraintestinal malignancies, but whether and how carcinogenesis drives compositional shifts of the microbiome to its own benefit remains an open conundrum. Here, we show that malignant processes can cause ileal mucosa atrophy, with villous microvascular constriction associated with dominance of sympathetic over cholinergic signaling. The rapid onset of tumorigenesis induced a burst of REG3γ release by ileal cells, and transient epithelial barrier permeability that culminated in overt and long-lasting dysbiosis dominated by Gram-positive Clostridium species. Pharmacologic blockade of β-adrenergic receptors or genetic deficiency in Adrb2 gene, vancomycin, or cohousing of tumor bearers with tumor-free littermates prevented cancer-induced ileopathy, eventually slowing tumor growth kinetics. Patients with cancer harbor distinct hallmarks of this stress ileopathy dominated by Clostridium species. Hence, stress ileopathy is a corollary disease of extraintestinal malignancies requiring specific therapies., Significance: Whether gut dysbiosis promotes tumorigenesis and how it controls tumor progression remain open questions. We show that 50% of transplantable extraintestinal malignancies triggered a β-adrenergic receptor-dependent ileal mucosa atrophy, associated with increased gut permeability, sustained Clostridium spp.-related dysbiosis, and cancer growth. Vancomycin or propranolol prevented cancer-associated stress ileopathy. This article is highlighted in the In This Issue feature, p. 873., (©2021 American Association for Cancer Research.)

Aside from PD-L1 expression, biomarkers of response to immune checkpoint inhibitors (ICIs) in non-small-cell lung cancer (NSCLC) are needed. In a previous retrospective analysis, we documented that fecal Akkermansia muciniphila (Akk) was associated with clinical benefit of ICI in patients with NSCLC or kidney cancer. In the current study, we performed shotgun-metagenomics-based microbiome profiling in a large cohort of patients with advanced NSCLC (n = 338) treated with first- or second-line ICIs to prospectively validate the predictive value of fecal Akk. Baseline stool Akk was associated with increased objective response rates and overall survival in multivariate analyses, independent of PD-L1 expression, antibiotics, and performance status. Intestinal Akk was accompanied by a richer commensalism, including Eubacterium hallii and Bifidobacterium adolescentis, and a more inflamed tumor microenvironment in a subset of patients. However, antibiotic use (20% of cases) coincided with a relative dominance of Akk above 4.8% accompanied with the genus Clostridium, both associated with resistance to ICI. Our study shows significant differences in relative abundance of Akk that may represent potential biomarkers to refine patient stratification in future studies., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The prognosis of early breast cancer (BC) relies on cell autonomous and immune parameters. The impact of the intestinal microbiome on clinical outcome has not yet been evaluated. Shotgun metagenomics was used to determine the composition of the fecal microbiota in 121 specimens from 76 early BC patients, 45 of whom were paired before and after chemotherapy. These patients were enrolled in the CANTO prospective study designed to record the side effects associated with the clinical management of BC. We analyzed associations between baseline or post-chemotherapy fecal microbiota and plasma metabolomics with BC prognosis, as well as with therapy-induced side effects. We examined the clinical relevance of these findings in immunocompetent mice colonized with BC patient microbiota that were subsequently challenged with histo-compatible mouse BC and chemotherapy. We conclude that specific gut commensals that are overabundant in BC patients compared with healthy individuals negatively impact BC prognosis, are modulated by chemotherapy, and may influence weight gain and neurological side effects of BC therapies. These findings obtained in adjuvant and neoadjuvant settings warrant prospective validation., (© 2021. The Author(s).)

Treatment with combined immune checkpoint blockade (CICB) targeting CTLA-4 and PD-1 is associated with clinical benefit across tumor types, but also a high rate of immune-related adverse events. Insights into biomarkers and mechanisms of response and toxicity to CICB are needed. To address this, we profiled the blood, tumor and gut microbiome of 77 patients with advanced melanoma treated with CICB, with a high rate of any ≥grade 3 immune-related adverse events (49%) with parallel studies in pre-clinical models. Tumor-associated immune and genomic biomarkers of response to CICB were similar to those identified for ICB monotherapy, and toxicity from CICB was associated with a more diverse peripheral T-cell repertoire. Profiling of gut microbiota demonstrated a significantly higher abundance of Bacteroides intestinalis in patients with toxicity, with upregulation of mucosal IL-1β in patient samples of colitis and in pre-clinical models. Together, these data offer potential new therapeutic angles for targeting toxicity to CICB., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been recently demonstrated in the sputum or saliva, suggesting how the shedding of viral RNA outlasts the end of symptoms. Recent data from transcriptome analysis show that the oral cavity mucosa harbors high levels of angiotensin-converting enzyme 2 (ACE2) and transmembrane protease, serine 2 (TMPRSS2), highlighting its role as a double-edged sword for SARS-CoV-2 body entrance or interpersonal transmission. Here, we studied the oral microbiota structure and inflammatory profile of 26 naive severe coronavirus disease 2019 (COVID-19) patients and 15 controls by 16S rRNA V2 automated targeted sequencing and magnetic bead-based multiplex immunoassays, respectively. A significant diminution in species richness was observed in COVID-19 patients, along with a marked difference in beta-diversity. Species such as Prevotella salivae and Veillonella infantium were distinctive for COVID-19 patients, while Neisseria perflava and Rothia mucilaginosa were predominant in controls. Interestingly, these two groups of oral species oppositely clustered within the bacterial network, defining two distinct Species Interacting Groups (SIGs). COVID-19-related pro-inflammatory cytokines were found in both oral and serum samples, along with a specific bacterial consortium able to counteract them. We introduced a new parameter, named CytoCOV, able to predict COVID-19 susceptibility for an unknown subject at 71% of power with an Area Under Curve (AUC) equal to 0.995. This pilot study evidenced a distinctive oral microbiota composition in COVID-19 subjects, with a definite structural network in relation to secreted cytokines. Our results would be usable in clinics against COVID-19, using bacterial consortia as biomarkers or to reduce local inflammation., Competing Interests: The 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 Iebba, Zanotta, Campisciano, Zerbato, Di Bella, Cason, Luzzati, Confalonieri, Palamara and Comar.)

A deviated repertoire of the gut microbiome predicts resistance to cancer immunotherapy. Enterococcus hirae compensated cancer-associated dysbiosis in various tumor models. However, the mechanisms by which E. hirae restored the efficacy of cyclophosphamide administered with concomitant antibiotics remain ill defined. Here, we analyzed the multifaceted modes of action of this anticancer probiotic. Firstly, E. hirae elicited emigration of thymocytes and triggered systemic and intratumoral IFNγ-producing and CD137-expressing effector memory T cell responses. Secondly, E. hirae activated the autophagy machinery in enterocytes and mediated ATG4B-dependent anticancer effects, likely as a consequence of its ability to increase local delivery of polyamines. Thirdly, E. hirae shifted the host microbiome toward a Bifidobacteria-enriched ecosystem. In contrast to the live bacterium, its pasteurized cells or membrane vesicles were devoid of anticancer properties. These pleiotropic functions allow the design of optimal immunotherapies combining E. hirae with CD137 agonistic antibodies, spermidine, or Bifidobacterium animalis. We surmise that immunological, metabolic, epithelial, and microbial modes of action of the live E. hirae cooperate to circumvent primary resistance to therapy.

Ileal epithelial cell apoptosis and the local microbiota modulate the effects of oxaliplatin against proximal colon cancer by modulating tumor immunosurveillance. Here, we identified an ileal immune profile associated with the prognosis of colon cancer and responses to chemotherapy. The whole immune ileal transcriptome was upregulated in poor-prognosis patients with proximal colon cancer, while the colonic immunity of healthy and neoplastic areas was downregulated (except for the Th17 fingerprint) in such patients. Similar observations were made across experimental models of implanted and spontaneous murine colon cancer, showing a relationship between carcinogenesis and ileal inflammation. Conversely, oxaliplatin-based chemotherapy could restore a favorable, attenuated ileal immune fingerprint in responders. These results suggest that chemotherapy inversely shapes the immune profile of the ileum-tumor axis, influencing clinical outcome.

Limited experimental evidence bridges nutrition and cancer immunosurveillance. Here, we show that ketogenic diet (KD) - or its principal ketone body, 3-hydroxybutyrate (3HB), most specifically in intermittent scheduling - induced T cell-dependent tumor growth retardation of aggressive tumor models. In conditions in which anti-PD-1 alone or in combination with anti-CTLA-4 failed to reduce tumor growth in mice receiving a standard diet, KD, or oral supplementation of 3HB reestablished therapeutic responses. Supplementation of KD with sucrose (which breaks ketogenesis, abolishing 3HB production) or with a pharmacological antagonist of the 3HB receptor GPR109A abolished the antitumor effects. Mechanistically, 3HB prevented the immune checkpoint blockade-linked upregulation of PD-L1 on myeloid cells, while favoring the expansion of CXCR3+ T cells. KD induced compositional changes of the gut microbiota, with distinct species such as Eisenbergiella massiliensis commonly emerging in mice and humans subjected to carbohydrate-low diet interventions and highly correlating with serum concentrations of 3HB. Altogether, these results demonstrate that KD induces a 3HB-mediated antineoplastic effect that relies on T cell-mediated cancer immunosurveillance.

Infertile couples undergoing the use of assisted reproductive technology are a good study model to evaluate the microbiological signatures affecting reproductive health. We tested vaginal lavages, follicular fluids, embryo culture mediums, and seminal fluids from 47 couples for their microbiome composition and HPV infection. Twenty-five infertile couples were diagnosed with unexplained infertility, whereas 22 were diagnosed with explained infertility. Lactobacilli were dominant in the vaginal lavages of both patient groups, and the most abundant species was L. iners (CST III), which is linked to a decreased fertility rate. Besides this, L. gasseri -which is known to be associated with oocyte DNA fragmentation and decreased sperm mobility-was identified in the seminal fluids, follicular fluids, and embryo culture media of the unexplained infertility group. Prevotella was increased in the seminal fluids of the explained infertility group, along with HPV-positive seminal fluids: an infection commonly associated with infertility, especially male infertility. Prevotella has been described to negatively affect sperm motility. Taken together, these results suggest that the profiling of the reproductive tract microbiome can add new perspectives to human reproduction.

Intestinal microbiota have been proposed to induce commensal-specific memory T cells that cross-react with tumor-associated antigens. We identified major histocompatibility complex (MHC) class I-binding epitopes in the tail length tape measure protein (TMP) of a prophage found in the genome of the bacteriophage Enterococcus hirae Mice bearing E. hirae harboring this prophage mounted a TMP-specific H-2K b -restricted CD8 + T lymphocyte response upon immunotherapy with cyclophosphamide or anti-PD-1 antibodies. Administration of bacterial strains engineered to express the TMP epitope improved immunotherapy in mice. In renal and lung cancer patients, the presence of the enterococcal prophage in stools and expression of a TMP-cross-reactive antigen by tumors correlated with long-term benefit of PD-1 blockade therapy. In melanoma patients, T cell clones recognizing naturally processed cancer antigens that are cross-reactive with microbial peptides were detected., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Background: The development of immune checkpoint blockade (ICB) has revolutionized the clinical outcome of renal cell carcinoma (RCC). Nevertheless, improvement of durability and prediction of responses remain unmet medical needs. While it has been recognized that antibiotics (ATBs) decrease the clinical activity of ICB across various malignancies, little is known about the direct impact of distinct intestinal nonpathogenic bacteria (commensals) on therapeutic outcomes of ICB in RCC., Objective: To evaluate the predictive value of stool bacteria composition for ICB efficacy in a cohort of advanced RCC patients., Design, Setting, and Participants: We prospectively collected fecal samples from 69 advanced RCC patients treated with nivolumab and enrolled in the GETUG-AFU 26 NIVOREN microbiota translational substudy phase 2 trial (NCT03013335) at Gustave Roussy. We recorded patient characteristics including ATB use, prior systemic therapies, and response criteria. We analyzed 2994 samples of feces from healthy volunteers (HVs). In parallel, preclinical studies performed in RCC-bearing mice that received fecal transplant (FMT) from RCC patients resistant to ICB (NR-FMT) allowed us to draw a cause-effect relationship between gut bacteria composition and clinical outcomes for ICB. The influence of tyrosine kinase inhibitors (TKIs) taken before starting nivolumab on the microbiota composition has also been assessed., Outcome Measurements and Statistical Analysis: Metagenomic data (MG) from whole genome sequencing (WGS) were analyzed by multivariate and pairwise comparisons/fold ratio to identify bacterial fingerprints related to ATB or prior TKI exposure and patients' therapeutic response (overall response and progression-free survival), and compared with the data from cancer-free donors., Results and Limitations: Recent ATB use (n = 11; 16%) reduced objective response rates (from 28% to 9%, p < 0.03) and markedly affected the composition of the microbiota, facilitating the dominance of distinct species such as Clostridium hathewayi, which were also preferentially over-represented in stools from RCC patients compared with HVs. Importantly, TKIs taken prior to nivolumab had implications in shifting the microbiota composition. To establish a cause-effect relationship between gut bacteria composition and ICB efficacy, NR-FMT mice were successfully compensated with either FMT from responding RCC patients or beneficial commensals identified by WGS-MG (Akkermansia muciniphila and Bacteroides salyersiae)., Conclusions: The composition of the microbiota is influenced by TKIs and ATBs, and impacts the success of immunotherapy. Future studies will help sharpen the role of these specific bacteria and their potential as new biomarkers., Patient Summary: We used quantitative shotgun DNA sequencing of fecal microbes as well as preclinical models of fecal or bacterial transfer to study the association between stool composition and (pre)clinical outcome to immune checkpoint blockade. Novel insights into the pathophysiological relevance of intestinal dysbiosis in the prognosis of kidney cancer may lead to innovative therapeutic solutions, such as supplementation with probiotics to prevent primary resistance to therapy., (Copyright © 2020. Published by Elsevier B.V.)

Objective: HIV-1-associated dysbiosis is most commonly characterized by overall decreased diversity, with abundance of the genus Prevotella, recently related to inflammatory responses., Design: A pilot study including 10 antiretroviral therapy-treated HIV-1-infected men and 50 uninfected controls was performed to identify the main gut dysbiosis determinants (e.g. Prevotella enrichment), that may affect mucosal antiviral defenses and T cell immunity in HIV-1-infected individuals., Methods: 16rRNA gene sequencing was applied to the HIV-1-infected individuals' fecal microbiota and compared with controls. Measurements of CD4 and CD8 T cell activation [CD38, human leukocyte antigen (HLA)-DR, CD38 HLA-DR] and frequencies of Th17, obtained from lamina propria lymphocytes isolated from five different intestinal sites, were performed by flow cytometry. IFNβ, IFNAR1 and MxA gene expression level was evaluated by real-time PCR in lamina propria lymphocytes. Nonparametric t tests were used for statistical analysis., Results: HIV-1-infected men had a significant fecal microbial communities' imbalance, including different levels of genera Faecalibacterium, Prevotella, Alistipes and Bacteroides, compared with controls. Notably, Prevotella abundance positively correlated with frequencies of CD4 T cells expressing CD38 or HLA-DR and coexpressing CD38 and HLA-DR (P < 0.05 for all these measures). The same trend was observed for the activated CD8 T cells. Moreover, Prevotella levels were inversely correlated with IFN-I genes (P < 0.05 for IFNβ, IFNAR1 and MxA genes) and the frequencies of Th17 cells (P < 0.05). By contrast, no statistically significant correlations were observed for the remaining bacterial genera., Conclusion: Our findings suggest that Prevotella enrichment might affect gut mucosal IFN-I pathways and T cell response in HIV-1-infected patients, thus contributing to immune dysfunction.

Microbiota are microorganismal communities colonizing human tissues exposed to the external environment, including the urogenital tract. The bacterial composition of the vaginal microbiota has been established and is partially related to obstetric outcome, while the uterine microbiota, considered to be a sterile environment for years, is now the focus of more extensive studies and debates. The characterization of the microbiota contained in the reproductive tract (RT) of asymptomatic and infertile women, could define a specific RT microbiota associated with implantation failure. In this pilot study, 34 women undergoing personalized hormonal stimulation were recruited and the biological samples of each patient, vaginal fluid, and endometrial biopsy, were collected immediately prior to oocyte-pick up, and sequenced. Women were subsequently divided into groups according to fertilization outcome. Analysis of the 16s rRNA V4-V5 region revealed a significant difference between vaginal and endometrial microbiota. The vaginal microbiota of pregnant women corroborated previous data, exhibiting a lactobacilli-dominant habitat compared to non-pregnant cases, while the endometrial bacterial colonization was characterized by a polymicrobial ecosystem in which lactobacilli were exclusively detected in the group that displayed unsuccessful in vitro fertilization. Overall, these preliminary results revisit our knowledge of the genitourinary microbiota, and highlight a putative relationship between vaginal/endometrial microbiota and reproductive success., (Copyright © 2020 Riganelli, Iebba, Piccioni, Illuminati, Bonfiglio, Neroni, Calvo, Gagliardi, Levrero, Merlino, Mariani, Capri, Pietrangeli, Schippa and Guerrieri.)

The prognosis of colon cancer (CC) is dictated by tumor-infiltrating lymphocytes, including follicular helper T (T FH ) cells and the efficacy of chemotherapy-induced immune responses. It remains unclear whether gut microbes contribute to the elicitation of T FH cell-driven responses. Here, we show that the ileal microbiota dictates tolerogenic versus immunogenic cell death of ileal intestinal epithelial cells (IECs) and the accumulation of T FH cells in patients with CC and mice. Suppression of IEC apoptosis led to compromised chemotherapy-induced immunosurveillance against CC in mice. Protective immune responses against CC were associated with residence of Bacteroides fragilis and Erysipelotrichaceae in the ileum. In the presence of these commensals, apoptotic ileal IECs elicited PD-1 + T FH cells in an interleukin-1R1- and interleukin-12-dependent manner. The ileal microbiome governed the efficacy of chemotherapy and PD-1 blockade in CC independently of microsatellite instability. These findings demonstrate that immunogenic ileal apoptosis contributes to the prognosis of chemotherapy-treated CC.

This short communication reports the preliminary results of Fecal Microbial Transplantation (FMT) impact on microbiota, microbial translocation (MT), and immune activation in four recurrent Clostridium difficile infection (R-CDI) patients. After FMT a restore of gut microbiota composition with a significant increase of fecal acetyl-putrescine and spermidine and fecal acetate and butyrate, a decrease of immune activation of T cells CD4+ and CD8+levels, and of LPS binding protein (LBP) level, were observed. Preliminary results indicate that FMT seems to be helpful not only as a CDI radical cure, with an impact on fecal microbiota and metabolome profiles, but also on MT and immune activation.

A microbial ecosystem in which bacteria no longer live in a mutualistic association is called dysbiotic. Gut microbiota dysbiosis is a condition related with the pathogenesis of intestinal illnesses (irritable bowel syndrome, celiac disease, and inflammatory bowel disease) and extra-intestinal illnesses (obesity, metabolic disorder, cardiovascular syndrome, allergy, and asthma). Dysbiosis status has been related to various important pathologies, and many therapeutic strategies aimed at restoring the balance of the intestinal ecosystem have been implemented. These strategies include the administration of probiotics, prebiotics, and synbiotics; phage therapy; fecal transplantation; bacterial consortium transplantation; and a still poorly investigated approach based on predatory bacteria. This review discusses the various aspects of these strategies to counteract intestinal dysbiosis.

In liver cirrhosis (LC), impaired intestinal functions lead to dysbiosis and possible bacterial translocation (BT). Bacteria or their byproducts within the bloodstream can thus play a role in systemic inflammation and hepatic encephalopathy (HE). We combined 16S sequencing, NMR metabolomics and network analysis to describe the interrelationships of members of the microbiota in LC biopsies, faeces, peripheral/portal blood and faecal metabolites with clinical parameters. LC faeces and biopsies showed marked dysbiosis with a heightened proportion of Enterobacteriaceae. Our approach showed impaired faecal bacterial metabolism of short-chain fatty acids (SCFAs) and carbon/methane sources in LC, along with an enhanced stress-related response. Sixteen species, mainly belonging to the Proteobacteria phylum, were shared between LC peripheral and portal blood and were functionally linked to iron metabolism. Faecal Enterobacteriaceae and trimethylamine were positively correlated with blood proinflammatory cytokines, while Ruminococcaceae and SCFAs played a protective role. Within the peripheral blood and faeces, certain species (Stenotrophomonas pavanii, Methylobacterium extorquens) and metabolites (methanol, threonine) were positively related to HE. Cirrhotic patients thus harbour a 'functional dysbiosis' in the faeces and peripheral/portal blood, with specific keystone species and metabolites related to clinical markers of systemic inflammation and HE.

The present study aimed to characterize the behavior of Bdellovibrio bacteriovorus in the presence of Staphylococcus aureus. B. bacteriovorus was co-cultured with S. aureus or Pseudomonas aeruginosa or Streptococcus mutans, in planktonic and sessile conditions. Co-cultures were studied by Field-Emission Scanning Electron Microscopy (FESEM), Scanning Transmission Electron Microscopy (STEM), turbidimetry, quantitative PCR (qPCR), and sequencing of gene Bd0108 of B. bacteriovorus. Results indicated that B. bacteriovorus comparably inhibited planktonic growth of P. aeruginosa and S. aureus, but not of S. mutans. FESEM and STEM showed that B. bacteriovorus interacts with S. aureus affecting its cell wall and membrane. Sequencing of gene Bd0108 did not reveal any of the mutations that can arise from the host-interaction (hit) locus. Although some Gram-negative species are reported to be B. bacteriovorus prey, it seems that in case of nutrient deficiency this predatory bacterium can also take advantage of some Gram-positive species. B. bacteriovorus behaviour in the presence of S. aureus is relevant for its possible therapeutic use in several pathologies, like cystic fibrosis in which S. aureus and P. aeruginosa frequently coexist as infectious agents.

The dynamics of swimming microorganisms is strongly affected by solid-liquid and air-liquid interfaces. In this paper, we characterize the motion of both single bacteria and microcolonies at an air-liquid interface. Both of them follow circular trajectories. Single bacteria preferentially show a counter-clockwise motion, in agreement with previous experimental and theoretical findings. Instead, no preferential rotation direction is observed for microcolonies suggesting that their motion is due to a different physical mechanism. We propose a simple mechanical model where the microcolonies move like rafts constrained to the air-liquid interface. Finally, we observed that the microcolony growth is due to the aggregation of colliding single-swimmers, suggesting that the microcolony formation resembles a condensation process where the first nucleus originates by the collision between two single-swimmers. Implications of microcolony splitting and aggregation on biofilm growth and dispersion at air-liquid interface are discussed., (© 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

There is a relationship between hepatic encephalopathy (HE) protein malnutrition and muscle wasting. Muscle may play an alternative role in ammonia detoxification. Molecular mechanisms responsible for muscle depletion are under investigation. Specific nutrients may interact to reverse the molecular pathways involved in muscle wasting at an early stage. Training exercises have also been proposed to improve skeletal muscle mass. However, these data refer to small groups of patients. The amelioration of muscle mass may potentially help to prevent HE. The pathogenesis of HE is associated with modifications of the gut microbiota and diet is emerging to play a relevant role in the modulation of the gut milieu. Vegetarian and fibre-rich diets have been shown to induce beneficial changes on gut microbiota in healthy people, with reduction of Bacteroides spp., Enterobacteriaceae, and Clostridium cluster XIVa bacteria. By way of contrast, it has been suggested that a high-fat or protein diet may increase Firmicutes and reduce Bacteroidetes phylum. Milk-lysozyme and milk-oligosaccharides have also been proposed to induce a "healthy" microbiota. At present, no studies have been published describing the modification of the gut microbiota in cirrhotic patients with HE as a response to specific diets. New research is needed to evaluate the potentiality of foods in the modulation of gut microbiota in liver disease and HE.

Background: Some beneficial effects of probiotics may be due to secreted probiotic-derived factors, identified as "postbiotic" mediators. The aim of this study was to evaluate whether supernatants harvested from Lactobacillus rhamnosus GG (LGG) cultures (ATCC53103 strain) protect colonic human smooth muscle cells (HSMCs) from lipopolysaccharide (LPS)-induced myogenic damage., Materials and Methods: LGG was grown in de Man, Rogosa, Share medium at 37°C and samples were collected in middle and late exponential, stationary, and overnight phases. Supernatants were recovered by centrifugation, filtered, and stored at -20°C. The primary HSMCs culture was exposed for 24 hours to purified LPS of a pathogen strain of Escherichia coli (O111:B4) (1 μg/mL) with and without supernatants. Postbiotic effects were evaluated on the basis of HSMCs morphofunctional alterations and interleukin-6 (IL-6) production. Data are expressed as mean±SE (P<0.05 significant)., Results: LPS induced persistent, significant, 20.5%±0.7% cell shortening and 34.5%±2.2% decrease in acetylcholine-induced contraction of human HSMCs. These morphofunctional alterations were paralleled to a 365.65%±203.13% increase in IL-6 production. All these effects were dose-dependently reduced by LGG supernatants. Supernatants of the middle exponential phase already partially restored LPS-induced cell shortening by 57.34%±12.7% and IL-6 increase by 145.8%±4.3% but had no effect on LPS-induced inhibition of contraction. Maximal protective effects were obtained with supernatants of the late stationary phase with LPS-induced cell shortening restored by 84.1%±4.7%, inhibition of contraction by 85.5%±6.4%, and IL-6 basal production by 92.7%±1.2%., Conclusions: LGG-derived products are able to protect human SMCs from LPS-induced myogenic damage. Novel insights have been provided for the possibility that LGG-derived products could reduce the risk of progression to postinfective motor disorders.

Introduction: Literature data provide little information about protozoa infections and gut microbiota compositional shifts in humans. This preliminary study aimed to describe the fecal bacterial community composition of people from Côte d'Ivoire harboring Giardia duodenalis, Entamoeba spp., and Blastocystis hominis, in trying to discover possible alterations in their fecal microbiota structure related to the presence of such parasites., Methodology: Twenty fecal samples were collected from people inhabiting three different localities of Côte d'Ivoire for copromicroscopic analysis and molecular identification of G. duodenalis, Entamoeba spp., and B. hominis. Temporal temperature gradient gel electrophoresis (TTGE) was used to obtain a fingerprint of the overall bacterial community; quantitative polymerase chain reaction (qPCR) was used to define the relative abundances of selected bacterial species/group, and multivariate statistical analyses were employed to correlate all data., Results: Cluster analysis revealed a significant separation of TTGE profiles into four clusters (p < 0.0001), with a marked difference for G. duodenalis-positive samples in relation to the others (p = 5.4×10-6). Interestingly, qPCR data showed how G. duodenalis-positive samples were related to a dysbiotic condition that favors potentially harmful species (such as Escherichia coli), while Entamoeba spp./B. hominis-positive subjects were linked to a eubiotic condition, as shown by a significantly higher Faecalibacterium prausnitzii-Escherichia coli ratio., Conclusions: This preliminary investigation demonstrates a differential fecal microbiota structure in subjects infected with G. duodenalis or Entamoeba spp./B. hominis, paving the way for using further next-generation DNA technologies to better understand host-parasite-bacteria interactions, aimed at identifying potential indicators of microbiota changes.

Microbial epidemiology and population genomics have previously been carried out near-exclusively for organisms grown in vitro. Metagenomics helps to overcome this limitation, but it is still challenging to achieve strain-level characterization of microorganisms from culture-independent data with sufficient resolution for epidemiological modelling. Here, we have developed multiple complementary approaches that can be combined to profile and track individual microbial strains. To specifically profile highly recombinant neisseriae from oral metagenomes, we integrated four metagenomic analysis techniques: single nucleotide polymorphisms in the clade's core genome, DNA uptake sequence signatures, metagenomic multilocus sequence typing and strain-specific marker genes. We applied these tools to 520 oral metagenomes from the Human Microbiome Project, finding evidence of site tropism and temporal intra-subject strain retention. Although the opportunistic pathogen Neisseria meningitidis is enriched for colonization in the throat, N. flavescens and N. subflava populate the tongue dorsum, and N. sicca, N. mucosa and N. elongata the gingival plaque. The buccal mucosa appeared as an intermediate ecological niche between the plaque and the tongue. The resulting approaches to metagenomic strain profiling are generalizable and can be extended to other organisms and microbiomes across environments.

Objective: The pathogenesis of salivary calculi is not yet clear; however, 2 theories have been formulated: (1) "the classic theory," based on calcium microdeposits in serous and ductal acinous cells, successively discharged into the ducts; (2) "the retrograde theory," based on a retrograde migration of food, bacteria, and so on from the oral cavity to the salivary duct. The aim of the present study is to highlight the role of bacteria and biofilm in stone formation., Study Design: Case series without comparison., Setting: Laboratory of the Department of Anatomical Pathology., Subjects and Methods: Traditional optic microscopy and scanning electron microscopy were carried out on 15 salivary gland calculi that were collected from 12 patients. A qPCR (quantitative real-time polymerase chain reaction) assay was performed to highlight the presence of bacterial DNA on each stone., Results: Optic microscopy showed formations that-due to their size, shape, and Gram and Giemsa staining-seemed to be Gram-positive bacterial cells. PAS- (periodic acid-Schiff) and alcian-PAS-positive staining matrix was present around them. The ultrastructural observation of the material processed for scanning electron microscopy showed the presence of structures resembling bacterial cells in the middle of the stones, surrounded by soft, amorphous material. Results of qPCR showed the presence of bacterial DNA in the internal part of the tissue sample., Conclusions: The presence of bacteria and/or bacterial products resembling biofilm in salivary gland stones supports the "retrograde theory." This evidence may support the hypothesis that biofilm could be the causative effect of lithiasic formations., (© American Academy of Otolaryngology—Head and Neck Surgery Foundation 2016.)

DNA methylation damage can be induced by endogenous and exogenous chemical agents, which has led every living organism to develop suitable response strategies. We investigated protein expression profiles of Escherichia coli upon exposure to the alkylating agent methyl-methane sulfonate (MMS) by differential proteomics. Quantitative proteomic data showed a massive downregulation of enzymes belonging to the glycolytic pathway and fatty acids degradation, strongly suggesting a decrease of energy production. A strong reduction in the expression of the N-acetylneuraminate lyases (NanA) involved in the sialic acid metabolism was also observed. Using a null NanA mutant and DANA, a substrate analog acting as competitive inhibitor, we demonstrated that down regulation of NanA affects biofilm formation and adhesion properties of E. coli MV1161. Exposure to alkylating agents also decreased biofilm formation and bacterial adhesion to Caco-2 eukaryotic cell line by the adherent invasive E. coli (AIEC) strain LF82. Our data showed that methylation stress impairs E. coli adhesion properties and suggest a possible role of NanA in biofilm formation and bacteria host interactions.

The microbial ecosystem of the gastrointestinal tract is characterized by a great number of microbial species living in balance by adopting mutualistic strategies. The eubiosis/dysbiosis condition of the gut microbiota strongly influences our healthy and disease status. This review briefly describes microbiota composition and functions, to then focus on eubiosis and dysbiosis status: the two sides of the microbiota.

Liver disease is associated with qualitative and quantitative changes in the intestinal microbiota. In cirrhotic patients the alteration in gut microbiota is characterized by an overgrowth of potentially pathogenic bacteria (i.e., gram negative species) and a decrease in autochthonous familiae. Here we summarize the available literature on the risk of gut dysbiosis in liver cirrhosis and its clinical consequences. We therefore described the features of the complex interaction between gut microbiota and cirrhotic host, the so called "gut-liver axis", with a particular attention to the acquired risk of bacterial translocation, systemic inflammation and the relationship with systemic infections in the cirrhotic patient. Such knowledge might help to develop novel and innovative strategies for the prevention and therapy of gut dysbiosis and its complication in liver cirrhosis.

Bdellovibrio bacteriovorus is a predator bacterial species found in the environment and within the human gut, able to attack Gram-negative prey. Cystic fibrosis (CF) is a genetic disease which usually presents lung colonization by Pseudomonas aeruginosa or Staphylococcus aureus biofilms. Here, we investigated the predatory behavior of B. bacteriovorus against these two pathogenic species with: (1) broth culture; (2) "static" biofilms; (3) field emission scanning electron microscope (FESEM); (4) "flow" biofilms; (5) zymographic technique. We had the first evidence of B. bacteriovorus survival with a Gram-positive prey, revealing a direct cell-to-cell contact with S. aureus and a new "epibiotic" foraging strategy imaged with FESEM. Mean attaching time of HD100 to S. aureus cells was 185 s, while "static" and "flow" S. aureus biofilms were reduced by 74 (at 24 h) and 46% (at 20 h), respectively. Furthermore, zymograms showed a differential bacteriolytic activity exerted by the B. bacteriovorus lysates on P. aeruginosa and S. aureus. The dual foraging system against Gram-negative (periplasmic) and Gram-positive (epibiotic) prey could suggest the use of B. bacteriovorus as a "living antibiotic" in CF, even if further studies are required to simulate its in vivo predatory behavior.

Cystic fibrosis (CF) patients have chronic airway infection and frequent exposure to antibiotics, which often leads to the emergence of resistant organisms. Achromobacter xylosoxidans is a new emergent pathogen in CF spectrum. From 2005 to 2010 we had an outbreak in A. xylosoxidans prevalence in our CF center, thus, the present study was aimed at deeply investigating virulence traits of A. xylosoxidans strains isolated from infected CF patients. To this purpose, we assessed A. xylosoxidans genome variability by randomly amplified polymorphic DNA (RAPD), biofilm production, antibiotic resistances, and motility. All A. xylosoxidans strains resulted to be biofilm producers, and were resistant to antibiotics usually employed in CF treatment. Hodge Test showed the ability to produce carbapenemase in some strains. Strains who were resistant to β-lactamics antibiotics, showed the specific band related to metal β-lactamase (blaIMP-1), and some of them showed to possess the integron1. Around 81% of A. xylosoxidans strains were motile. Multivariate analysis showed that RAPD profiles were able to predict Forced Expiratory Volume (FEV1%) and biofilm classes. A significant prevalence of strong biofilm producers strains was found in CF patients with severely impaired lung functions (FEV1% class 1). The outbreak we had in our center (prevalence from 8.9 to 16%) could be explained by an enhanced adaptation of A. xylosoxidans in the nosocomial environment, despite of aggressive antibiotic regimens that CF patients usually undergo.

Introduction: In this study we investigated the effects of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene variants on the composition of faecal microbiota, in patients affected by Cystic Fibrosis (CF). CFTR mutations (F508del is the most common) lead to a decreased secretion of chloride/water, and to mucus sticky secretions, in pancreas, respiratory and gastrointestinal tracts. Intestinal manifestations are underestimated in CF, leading to ileum meconium at birth, or small bowel bacterial overgrowth in adult age., Methods: Thirty-six CF patients, fasting and under no-antibiotic treatment, were CFTR genotyped on both alleles. Faecal samples were subjected to molecular microbial profiling through Temporal Temperature Gradient Electrophoresis and species-specific PCR. Ecological parameters and multivariate algorithms were employed to find out if CFTR variants could be related to the microbiota structure., Results: Patients were classified by two different criteria: 1) presence/absence of F508del mutation; 2) disease severity in heterozygous and homozygous F508del patients. We found that homozygous-F508del and severe CF patients exhibited an enhanced dysbiotic faecal microbiota composition, even within the CF cohort itself, with higher biodiversity and evenness. We also found, by species-specific PCR, that potentially harmful species (Escherichia coli and Eubacterium biforme) were abundant in homozygous-F508del and severe CF patients, while beneficial species (Faecalibacterium prausnitzii, Bifidobacterium spp., and Eubacterium limosum) were reduced., Conclusions: This is the first report that establishes a link among CFTR variants and shifts in faecal microbiota, opening the way to studies that perceive CF as a 'systemic disease', linking the lung and the gut in a joined axis.

Introduction: Members of the human intestinal microbiota are key players in maintaining human health. Alterations in the composition of gut microbial community (dysbiosis) have been linked with important human diseases. Understanding the underlying processes that control community structure, including the bacterial interactions within the microbiota itself, is essential. Bdellovibrio bacteriovorus is a gram-negative bacterium that preys other gram-negative species for survival, acting as a population-balancer. It was found in terrestrial/aquatic ecosystems, and in animal intestines, postulating its presence also in the human gut., Methods: The present study was aimed to evaluate, by end-point PCR and qPCR, the presence of B. bacteriovorus in intestinal and faecal biopsy specimens from 92 paediatric healthy subjects and patients, suffering from Inflammatory Bowel Diseases (IBD), Celiac disease and Cystic fibrosis (CF)., Results: i) B. bacteriovorus was present and abundant only in healthy individuals, while it was heavily reduced in patients, as in the case of IBD and Celiac, while in CF patients and relative controls we observed comparable results; ii) B. bacteriovorus seemed to be mucosa-associated, because all IBD and Celiac biopsies (and related controls) were treated with mucus-removing agents, leaving only the mucosa-attached microflora; iii) B. bacteriovorus abundance was district-dependent, with a major preponderance in duodenum, and gradually decreasing up to rectum; iv) B. bacteriovorus levels significantly dropped in disease status, in duodenum and ileum., Conclusions: Results obtained in this study could represent the first step for new therapeutic strategies aimed to restore a balance in the intestinal ecosystem, utilizing Bdellovibrio as a probiotic.

Several studies reported increased numbers of mucosa-associated Escherichia coli strains in patients with inflammatory bowel disease (IBD), encompassing Crohn's disease (CD) and ulcerative colitis (UC). The majority of E. coli strains possess type 1 fimbriae, whose tip fibrillum protein, FimH, naturally undergoes amino acid replacements, an important process in the adaptation of commensal E. coli strains to environmental changes, like those observed in IBD and urinary tract infections. In this study, we analyzed mutational patterns in the fimH gene of 52 mucosa-associated E. coli strains isolated from IBD and non-IBD pediatric patients, in order to investigate microevolution of this genetic trait. FimH-positive strains were also phylogenetically typed and tested for their adhesive ability on Caco-2 cells. Specific FimH alleles for each grouping feature were found. Mutations G66S and V27A were related to CD, while mutations A242V, V163A, and T74I were attributed to UC. Otherwise, the G66S, N70S, and S78N mutations were specifically attributed to B2/D phylogroups. The N70S and A119V mutations were related to adhesive E. coli strains. Phylogroup B2, adhesive, and IBD E. coli strains showed a higher site substitution rate (SSR) in the fimH gene, together with a higher number of mutations. The degree of naïve mucosal inflammation was related to specific FimH alleles. Moreover, we could suggest that the V27A mutation is pathoadaptive for the CD intestinal habitat, while we could also suggest that both the N70S and S78N mutations are related to the more virulent E. coli B2 phylogroup. In conclusion, we found some FimH variants that seem to be more involved than others in the evolution of IBD pathogenesis.

Through genomic analysis of mucosa-associated Escherichia coli strains, we found a close genetic association among isolates from pediatric inflammatory bowel disease (IBD) patients. A specific E. coli pathovar, adherent-invasive E. coli (AIEC), was found in Crohn's disease (CD) adult patients - this pathovar has enhanced adhesive and invasive properties, mainly due to the mannose-bonding FimH protein. We aimed to characterize 52 mucosa-associated E. coli strains isolated from pediatric IBD and non-IBD patients. Eleven E. coli strains, showing a strong similarity in fimH gene sequence to that of E. coli AIEC LF82, were characterized for fimH gene sequence, genomic profiling, adhesive and invasive ability, and phylogrouping. The results were compared with E. coli strains AIEC LF82 and MG1655. The 11 E. coli isolates showed 82.4% ± 1.4% fimH sequence similarity and 80.6% ± 1.3% genomic similarity to strain AIEC LF82. All these strains harbored V27A and S78N FimH mutations, as found in LF82. Nine of them belonged to the more virulent B2 and D phylogroups. Neuraminidase treatment, mimicking inflamed mucosa, enhanced adhesion of all 11 strains by 3.5-fold, but none showed invasion ability. It could be argued that the 11 selected strains could be a branch of an E. coli subpopulation (pathobionts), that could take advantage in an inflamed context because of a suitable genomic and (or) genetic backdrop.

Background: Researchers have made every effort to assess the role of gut microbiota in pediatric diseases like inflammatory bowel disease (IBD), celiac disease, asthma, allergy, and autism. The leading hypothesis is that an altered microbial composition is present (other than the presence of a specific pathogen) and that it could be involved in the pathogenesis or progression of such disorders., Methods: Cultural, molecular, metabolomic, and metagenomic approaches are trying to define the pediatric gut microbiota imbalances in different diseases., Results and Conclusion: In pediatric IBD, a marked increase in aerobes and facultative anaerobes was found, along with an increase in Enterobacteriaceae members (Escherichia coli). In both pediatric IBD and celiac disease (Th1-mediated disorders), higher bacterial cell counts were observed, jointly with a general gain of biodiversity. A preponderance of Bacteroidetes and a parallel decrease of Firmicutes was also reported in IBD, celiac disease and autism. Contrarily, dietary changes due to Western lifestyles increase Firmicutes populations and lower short-chain fatty acids production, possibly exposing 'developed' children to the infectious challenge (Escherichia and Shigella spp.). Lactobacillus and Bifidobacterium species could be protective agents for atopic diseases, while Clostridia, Enterobacteriaceae, and staphylococci can be associated with an increased risk of such Th2-mediated disorders. In the brain-gut axis view, gut microbiota could also play a role in autism., (Copyright © 2011 S. Karger AG, Basel.)