Intestinal inflammation and permeability in patients recovered from SARS-CoV-2 infection.

INTRODUCTION: Different works suggest a close link between Long COVID Gastrointestinal (GI) manifestations and the post-infection disorders of gut-brain interaction (PI-DGBIs). However, the actual mechanisms underlying long-term GI sequelae are still not clear. Our study was aimed to assess both intestinal inflammation and permeability among subjects recovered from SARS-CoV-2 infection and their eventual correlation with long-term GI sequelae. METHODS: Eighty-six subjects attending the Post-COVID Service and recovered from SARS-CoV-2 infection for six months, were investigated for Long COVID manifestations. Those subjects complaining of long-term GI symptoms were further evaluated by Rome IV questionnaire to assess PI-DGBIs. Intestinal inflammation (by fecal calprotectin, FC), and permeability (by serum and fecal levels of zonulin) were evaluated in all subjects. The Hospital Anxiety and Depression Scale (HADS) and The Gastrointestinal Quality of Life Index (GIQLI) questionnaires were further provided to all participants. RESULTS: Thirty-seven subjects (43%) complained of long-term GI symptoms, while 49 subjects (57%) did not. Thirty-three subjects fulfilled Rome IV criteria for PI-DGBIs. FC values resulted higher in those subjects who did not complain GI symptoms (p=0.03), although remaining quite close to the normal range. No significant differences were shown regarding the assessment of intestinal permeability. By GIQLI, long-term gastrointestinal sequelae were inversely correlated with quality of life (p=0.009). CONCLUSION: Long COVID GI complaints unlikely recognize underlying local inflammatory mechanisms. Since the healthcare, economic, and social burden of post-COVID DGBIs, a deeper understanding of this emerging condition should be encouraged to improve management of the affected subjects.

Faecal Transplantation for Ulcerative Colitis From Diet Conditioned Donors Followed by Dietary Intervention Results in Favourable Gut Microbial Profile Compared to Faecal Transplantation Alone.

BACKGROUND AND AIMS: Several faecal microbial transplantation [FMT] approaches for ulcerative colitis [UC] have been investigated with conflicting results. We have recently published the clinical outcomes from the CRAFT UC Trial using FMT with the UC Exclusion Diet [UCED], compared with FMT alone. Here we aimed to compare the two FMT strategies in terms of microbial profile and function. METHODS: Subjects recruited to the CRAFT UC study with available pre- and post-intervention faecal samples were included. Donors received diet conditioning for 14 days based on the UCED principles. Group 1 received single FMT by colonoscopy [Day 1] and enemas [Days 2 and 14] without donors' dietary conditioning [N = 11]. Group 2 received FMT but with donors' dietary pre-conditioning and UCED for the patients [N = 10]. Faecal samples were assessed by DNA shotgun metagenomic sequencing. RESULTS: Following diet conditioning, donors showed depletion in metabolic pathways involved in biosynthesis of sulphur-containing amino acids. Only Group 2 showed significant shifts towards the donors' microbial composition [ADONIS: R2 = 0.15, p = 0.008] and significantly increased Eubacterium_sp_AF228LB post-intervention [ß-coefficient 2.66, 95% confidence interval 2.1-3.3, q < 0.05] which was inversely correlated with faecal calprotectin [rho = -0.52, p = 0.035]. Moreover, pathways involved in gut inflammation and barrier function including branched chain amino acids were enriched post-intervention in Group 2 and were significantly inversely correlated with faecal calprotectin. CONCLUSION: FMT from diet conditioned donors followed by the UCED led to microbial alterations associated with favourable microbial profiles which correlated with decreased faecal calprotectin. Our findings support further exploration of the additive benefit of dietary intervention for both donors and patients undergoing FMT as a potential treatment of UC.

Amyotrophic lateral sclerosis stratification: unveiling patterns with virome, inflammation, and metabolism molecules.

Amyotrophic lateral sclerosis (ALS) is an untreatable and clinically heterogeneous condition primarily affecting motor neurons. The ongoing quest for reliable biomarkers that mirror the disease status and progression has led to investigations that extend beyond motor neurons' pathology, encompassing broader systemic factors such as metabolism, immunity, and the microbiome. Our study contributes to this effort by examining the potential role of microbiome-related components, including viral elements, such as torque tenovirus (TTV), and various inflammatory factors, in ALS. In our analysis of serum samples from 100 ALS patients and 34 healthy controls (HC), we evaluated 14 cytokines, TTV DNA load, and 18 free fatty acids (FFA). We found that the evaluated variables are effective in differentiating ALS patients from healthy controls. In addition, our research identifies four unique patient clusters, each characterized by distinct biological profiles. Intriguingly, no correlations were found with site of onset, sex, progression rate, phenotype, or C9ORF72 expansion. A remarkable aspect of our findings is the discovery of a gender-specific relationship between levels of 2-ethylhexanoic acid and patient survival. In addition to contributing to the growing body of evidence suggesting altered peripheral immune responses in ALS, our exploratory research underscores metabolic diversity challenging conventional clinical classifications. If our exploratory findings are validated by further research, they could significantly impact disease understanding and patient care customization. Identifying groups based on biological profiles might aid in clustering patients with varying responses to treatments.

Donor screening for fecal microbiota transplantation with a direct stool testing-based strategy: a prospective cohort study.

Fecal microbiota transplantation (FMT) is effective against recurrent Clostridioides difficile infection (rCDI), but its safety is jeopardized by the potential transmission of pathogens, so international guidelines recommend either a quarantine or a direct stool testing. Whereas reports of the quarantine-based approach are emerging, data on the direct testing-based approach are not available. Our aim is to report outcomes of a donor screening framework for FMT including direct stool testing. In this prospective cohort study, all donor candidates recruited at our FMT centre underwent a four-step screening process to be enrolled as actual donors. Each collected stool donation was then evaluated with a direct stool testing including a molecular assay for gut pathogens and a culture assay for multi-drug resistant organisms (MDRO). From January 2019 to June 2023, 72 of 227 candidates (32%) were considered eligible and provided 277 stool donations. Ninety-nine donations (36%) were discarded for positivity to intestinal pathogens, most commonly enteropathogenic Escherichia coli (n = 37) and Blastocystis hominis (n = 20). Overall, 337 stool aliquots were obtained from 165 approved donations. All suspensions were used for patients with rCDI, and no serious adverse events or clinically evident infections were observed at 12 weeks after procedures. In our study, screening of donor faeces including direct stool testing led to the discard of a considerable rate of stool donations but was also extremely safe. This approach may represent a reliable strategy to guarantee the safety of FMT programs, especially in countries with high prevalence of MDRO.

Fecal Microbiota Transplantation in Amyotrophic Lateral Sclerosis: Clinical Protocol and Evaluation of Microbiota Immunity Axis.

The fecal microbial transplantation (FMT) is a therapeutic transplant of fecal microbiota from healthy donors to patients. This practice is aimed at restoring eubiosis and rebalancing the enteric and systemic immune responses, and then eliminating pathogenic triggers of multiple disease, including neurodegenerative diseases. Alterations of gut microbiota (GM) affect the central nervous system (CNS) health, impacting neuro-immune interactions, synaptic plasticity, myelination, and skeletal muscle function. T-regulatory lymphocytes (Treg) are among the most important players in the pathogenesis of amyotrophic lateral sclerosis (ALS), altering the disease course. Along with circulating neuropeptides, other immune cells, and the gut-brain axis, the GM influences immunological tolerance and controls Treg's number and suppressive functions. A double-blind, controlled, multicenter study on FMT in ALS patients has been designed to evaluate if FMT can modulate neuroinflammation, by restoring Treg number, thus modifying disease activity and progression.

Reporting a single case of cystic echinococcosis in retroperitoneal mass of uncertain origin.

Echinococcal disease (hydatid disease (HD) is an endemic parasitosis caused by Echinococcus granulosus in the larval stage, and it is typically due to the production of unilocular cystic lesions, usually involving the liver for the majority of patients and the lungs in 25%, but also any other organs can be potentially involved in developing echinococcal disease. We report a case of extrahepatic, retroperitoneal echinococcal disease, caused by Echinococcus granulosus. The patient underwent a surgical removal of the abdominal mass, revealed by abdominal ultrasound and computerized tomography scanning, and in the founded clinical and radiological suspicion of echinococcal disease, multiple bioptical samples were sent for microbiological analysis and albendazole therapy was started; Echinococcus granulosus protoscolices were found on the bioptical sample, and the diagnosis was successfully confirmed. According to the current parasitology literature on echinococcal disease, extrahepatic localization, although rare, can be found, and it should be considered in the differential diagnosis of an abdominal mass when epidemiological risk factors and anamnestic data are present, regardless of the usual site of the disease.

Serial Fecal Microbiota Infusions via Colonoscopy for Active Ulcerative Colitis: A Feasibility, Safety, and Translational Monocentric Italian Study.

The effectiveness of fecal microbiota transplantation (FMT) in ulcerative colitis (UC) remains unclear. This study aimed to investigate the feasibility and effectiveness of serial fecal infusions via colonoscopy in patients with active UC. Subjects with mild-to-moderate UC received three consecutive fecal infusions via colonoscopy. A control population with the same baseline features receiving Infliximab treatment was enrolled. Adverse events and clinical, endoscopic, and microbial outcomes were investigated. Nineteen patients with mildly-to-moderately active UC were enrolled. Clinical response was obtained in six patients at week 2, in eight at week 6, and in nine at week 12. Clinical response was maintained in eight patients at week 24. Endoscopic remission at week 12 was reached in six patients. In the control population, 13/19 patients achieved clinical response at week 6, and 10/19 patients maintained clinical response after 6 months. Microbiota richness was higher in responders compared with the non-responders. Peptostreptococcus, Lactobacillus, and Veillonella were higher in non-responders, while Parabacteroides, Bacteroides, Faecalibacterium, and Akkermansia were higher in responders at all timepoints. Serial FMT infusions appear to be feasible, safe, and effective in UC patients, with a potential role in inducing and maintaining clinical response. Specific bacteria predict the response to FMT.

The first international Rome consensus conference on gut microbiota and faecal microbiota transplantation in inflammatory bowel disease.

BACKGROUND: Several randomised clinical trials (RCTs) performing faecal microbiota transplantation (FMT) for the management of inflammatory bowel disease (IBD), particularly for ulcerative colitis, have recently been published, but with major variations in study design. These include differences in administered dose, route and frequency of delivery, type of placebo and evaluated endpoints. Although the overall outcomes appear to be promising, they are highly dependent on both donor and recipient factors. OBJECTIVE: To develop concensus-based statements and recommendations for the evaluation, management and potential treatment of IBD using FMT in order to move towards standardised practices. DESIGN: An international panel of experts convened several times to generate evidence-based guidelines by performing a deep evaluation of currently available and/or published data. Twenty-five experts in IBD, immunology and microbiology collaborated in different working groups to provide statements on the following key issues related to FMT in IBD: (A) pathogenesis and rationale, (B) donor selection and biobanking, (C) FMT practices and (D) consideration of future studies and perspectives. Statements were evaluated and voted on by all members using an electronic Delphi process, culminating in a plenary consensus conference and generation of proposed guidelines. RESULTS AND CONCLUSIONS: Our group has provided specific statements and recommendations, based on best available evidence, with the end goal of providing guidance and general criteria required to promote FMT as a recognised strategy for the treatment of IBD.

Fecal microbiota transplantation for recurrent Clostridioides difficile infection in patients with concurrent ulcerative colitis.

AIMS: Clostridioides difficile infection (CDI) is a major challenge for healthcare systems. Inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn's disease, is a risk factor for primary and recurrent CDI (rCDI). Moreover, CDI itself often worsens the clinical picture of IBD, increasing the risk of complications. Fecal microbiota transplantation (FMT) is a highly effective treatment for rCDI, but data from patients with IBD and CDI are limited and often referred to mixed cohorts. We aimed to report outcomes from a cohort of patients with UC treated with FMT for rCDI superinfection. METHODS AND RESULTS: In a retrospective, single-centre cohort study we evaluated characteristics and outcomes of patients with UC who received FMT for rCDI. The primary outcome was negative C. difficile toxin 8 weeks after FMT. Thirty-five patients were included in the analysis. Sixteen patients were cured after single FMT, while 19 patients received repeat FMT. Overall, FMT cured rCDI in 32 patients (91%), and repeat FMT was significantly associated with sustained cure of CDI compared with single FMT (84% vs 50%, p = 0.018). Twenty-four patients (69%) experienced remission or an amelioration of UC activity. Serious adverse events were not observed. CONCLUSIONS: In our cohort of patients with UC, FMT was highly effective in curing rCDI without severe adverse events and repeat FMT was significantly associated with CDI cure. Most patients also experienced remission or amelioration of UC activity after FMT. Our findings suggest that a sequential FMT protocol may be used routinely in patients with UC and rCDI.

Celiac Disease Predisposition and Genital Tract Microbiota in Women Affected by Recurrent Pregnancy Loss.

The incidence of Idiopathic Recurrent Pregnancy Loss (RPL) is doubled in patients suffering from Celiac Disease (CD) compared to healthy populations. CD genetic components are HLA class II genes known as HLA-DQ2 and DQ8. Genetically susceptible women can remain asymptomatic even though they are exposed to a doubled risk of RPL compared to the general population. Furthermore, CD has been associated with microbiota alterations. The aim of this study is to evaluate endometrial and vaginal microbiota in HLA-DQ2/DQ8 positive and negative RPL patients compared to healthy pregnant women. Endometrial and vaginal microbiota of 3 subgroups were evaluated: 15 HLA-DQ2/DQ8 positive RPL women, 25 HLA DQ2/DQ8 negative RPL women (for a total of 40 RPL women) and 7 healthy fertile controls with previous uncomplicated pregnancies (all HLA-DQ2/DQ8 negative). The 2 RPL subgroups (HLA-DQ2/DQ8 positive and negative) showed a different endometrial and vaginal composition in the Lactobacillacae family compared to controls: Lactobacillus acidophilus was absent both in the vaginal and endometrial samples of RPL women, while Lactobaciluus iners, which can favor a less stable vaginal microbiota, was found only in RPL women (26.4% in HLA DQ2/DQ8 positive and 22.1% HLA DQ2/DQ8 negative) in both the vaginal and endometrial districts. In conclusion, both HLA DQ2/DQ8 positive-RPL and HLA DQ2/DQ8 negative-RPL women showed different endometrial and vaginal microbiota composition compared to healthy controls.

Fecal Microbiota Transplantation and Other Gut Microbiota Manipulation Strategies.

The gut microbiota is composed of bacteria, archaea, phages, and protozoa. It is now well known that their mutual interactions and metabolism influence host organism pathophysiology. Over the years, there has been growing interest in the composition of the gut microbiota and intervention strategies in order to modulate it. Characterizing the gut microbial populations represents the first step to clarifying the impact on the health/illness equilibrium, and then developing potential tools suited for each clinical disorder. In this review, we discuss the current gut microbiota manipulation strategies available and their clinical applications in personalized medicine. Among them, FMT represents the most widely explored therapeutic tools as recent guidelines and standardization protocols, not only for intestinal disorders. On the other hand, the use of prebiotics and probiotics has evidence of encouraging findings on their safety, patient compliance, and inter-individual effectiveness. In recent years, avant-garde approaches have emerged, including engineered bacterial strains, phage therapy, and genome editing (CRISPR-Cas9), which require further investigation through clinical trials.

Variability of strain engraftment and predictability of microbiome composition after fecal microbiota transplantation across different diseases.

Fecal microbiota transplantation (FMT) is highly effective against recurrent Clostridioides difficile infection and is considered a promising treatment for other microbiome-related disorders, but a comprehensive understanding of microbial engraftment dynamics is lacking, which prevents informed applications of this therapeutic approach. Here, we performed an integrated shotgun metagenomic systematic meta-analysis of new and publicly available stool microbiomes collected from 226 triads of donors, pre-FMT recipients and post-FMT recipients across eight different disease types. By leveraging improved metagenomic strain-profiling to infer strain sharing, we found that recipients with higher donor strain engraftment were more likely to experience clinical success after FMT (P = 0.017) when evaluated across studies. Considering all cohorts, increased engraftment was noted in individuals receiving FMT from multiple routes (for example, both via capsules and colonoscopy during the same treatment) as well as in antibiotic-treated recipients with infectious diseases compared with antibiotic-naïve patients with noncommunicable diseases. Bacteroidetes and Actinobacteria species (including Bifidobacteria) displayed higher engraftment than Firmicutes except for six under-characterized Firmicutes species. Cross-dataset machine learning predicted the presence or absence of species in the post-FMT recipient at 0.77 average AUROC in leave-one-dataset-out evaluation, and highlighted the relevance of microbial abundance, prevalence and taxonomy to infer post-FMT species presence. By exploring the dynamics of microbiome engraftment after FMT and their association with clinical variables, our study uncovered species-specific engraftment patterns and presented machine learning models able to predict donors that might optimize post-FMT specific microbiome characteristics for disease-targeted FMT protocols.

"Bacterial Consortium": A Potential Evolution of Fecal Microbiota Transplantation for the Treatment of Clostridioides difficile Infection.

Fecal microbiota transplantation (FMT) consists of infusion of feces from a donor to a recipient patient in order to restore the resident microbial population. FMT has shown to be a valid clinical option for Clostridioides difficile infections (CDI). However, this approach shows several criticalities, such as the recruiting and screening of voluntary donors. Our aim was to evaluate the therapeutic efficacy of a synthetic bacterial suspension defined "Bacterial Consortium" (BC) infused in the colon of CDI patients. The suspension was composed by 13 microbial species isolated by culturomics protocols from healthy donors' feces. The efficacy of the treatment was assessed both clinically and by metagenomics typing. Fecal samples of the recipient patients were collected before and after infusion. DNA samples obtained from feces at different time points (preinfusion, 7, 15, 30, and 90 days after infusion) were analyzed by next-generation sequencing. Before infusion, patient 1 showed an intestinal microbiota dominated by the phylum Bacteroidetes. Seven days after the infusion, Bacteroidetes decreased, followed by an implementation of Firmicutes and Verrucomicrobia. Patient 2, before infusion, showed a strong abundance of Proteobacteria and a significant deficiency of Bacteroidetes and Verrucomicrobia. Seven days after infusion, Proteobacteria strongly decreased, while Bacteroidetes and Verrucomicrobia increased. Metagenomics data revealed an "awakening" by microbial species absent or low concentrated at time T0 and present after the infusion. In conclusion, the infusion of selected bacteria would act as a trigger factor for "bacterial repopulation" representing an innovative treatment in patients with Clostridioides difficile infections.

How the gut parasitome affects human health.

The human gut microbiome (GM) is a complex ecosystem that includes numerous prokaryotic and eukaryotic inhabitants. The composition of GM can influence an array of host physiological functions including immune development. Accumulating evidence suggest that several members of non-bacterial microbiota, including protozoa and helminths, that were earlier considered as pathogens, could have a commensal or beneficial relationship with the host. Here we examine the most recent data from omics studies on prokaryota-meiofauna-host interaction as well as the impact of gut parasitome on gut bacterial ecology and its role as 'immunological driver' in health and disease to glimpse new therapeutic perspectives.

Rummeliibacillus suwonensis: First Time Isolation from Human Feces by Culturomics.

Gut microbiota is a complex ecosystem composed by trillions of microorganisms that are crucial for human health or disease status. Currently, there are two methodological options to explore its complexity: metagenomics and culturomics. Culturomics is an approach that uses multiple culture conditions (days of incubation, enrichment factors and growth temperature) and MALDI-TOF mass spectrometry for the identification of bacterial species and sequencing when this method fails. In this paper, we describe how Colturomic's protocol has allowed the first isolation in human sample of Rummeliibacillus suwonensis, a Gram positive, facultative anaerobe bacterium. The bacterium was isolated from feces of a 69 years old male with amyotrophic lateral sclerosis (ALS) recruited for a clinical trial assessing safety and efficacy of fecal microbiota transplantation in ALS. The first isolation of the microorganism dates back to 2013 from the soil of a South Korean mountain area. In this report, morphological description, biochemical characterization and antibiotic susceptibility tests were performed to outline the bacterial properties.

Clostridioides difficile Infection in Children: A 5-Year Multicenter Retrospective Study.

While there are numerous studies regarding Clostridioides difficile infection (CDI) in adults, literature on the pediatric population is scarce. Therefore, we performed a 5-year retrospective study between January 2014 and December 2018 in two referral centers in Rome, Italy. There were 359 patients tested for CDI who were enrolled: 87 resulted in positive and 272 in negative. CDI children had a higher number of previous-day hospital admissions (p = 0.024), hospitalizations (p = 0.001), and total hospital admissions (p = 0.008). Chronic comorbidities were more frequent in the CDI group (66.7% vs. 33.3%). Previous use of proton pump inhibitors and antibiotics was associated with CDI (p < 0.001). Among the antibiotics, only fluoroquinolones were significantly associated with CDI. Also, CDI children were more frequently exposed to antibiotics during the episode of hospitalization when children were tested. Our study provides an updated clinical and epidemiological analysis of children with CDI compared with a control group of children who tested negative. Further prospective studies could better define risk factors and preventive methods.

Validation of Two Commercial Multiplex Real-Time PCR Assays for Detection of SARS-CoV-2 in Stool Donors for Fecal Microbiota Transplantation.

Recurrent infection by Clostridioides difficile has recently been treated by fecal microbiota transplantation (FMT). As viable SARS-CoV-2 was recovered from stool of asymptomatic individuals, the FMT procedure could be a potential risk of SARS-CoV-2 transmission, thus underlying the need to reliably detect SARS-CoV-2 in stool. Here, we performed a multicentric study to explore performances of two commercially available assays for detection of SARS-CoV-2 RNA in stool of potential FMT donors. In three hospitals, 180 stool samples were spiked with serial 10-fold dilutions of a SARS-CoV-2 inactivated lysate to evaluate the Seegene Allplex™ SARS-CoV-2 (SC2) and SARS-CoV-2/FluA/FluB/RSV (SC2FABR) Assays for the detection of viral RNA in stool of FMT donors. The results revealed that both assays detected down to 2 TCID50/mL with comparable limit of detection values, SC2 showing more consistent target positivity rate than SC2FABR. Beyond high amplification efficiency, correlation between CT values and log concentrations of inactivated viral lysates showed R2 values ranging from 0.88 to 0.90 and from 0.87 to 0.91 for the SC2 and SC2FABR assay, respectively. The present results demonstrate that both methods are highly reproducible, sensitive, and accurate for SARS-CoV-2 RNA detection in stool, suggesting a potential use in FMT-donor screening.

Use of Faecal Transplantation with a Novel Diet for Mild to Moderate Active Ulcerative Colitis: The CRAFT UC Randomised Controlled Trial.

BACKGROUND: We evaluated whether integration of novel diets for donors and patients, in addition to faecal transplantation [FT], could increase FT remission rate in refractory ulcerative colitis [UC]. METHODS: This was a blinded, randomised, controlled trial in adults with active UC, defined by a simple clinical colitis activity index [SCCAI] of ≥5 and ≤11 and endoscopic Mayo score 2-3, refractory to medication. Group 1 received free diet and single donor standard FT by colonoscopy on Day 1and rectal enemas on Days 2 and 14 without dietary conditioning of the donor. Group 2 received FT as above but with dietary pre-conditioning of the donor for 14 days and a UC Exclusion Diet [UCED] for the patients. Group 3 received the UCED alone. The primary endpoint was Week 8 clinical steroid-free remission, defined as SCCAI <3. RESULTS: Of 96 planned patients, 62 were enrolled. Remission Week 8 Group 1 was 2/17 [11.8%], Group 2 was 4/19 [21.1%], Group 3 was 6/15 [40%] [non-significant]. Endoscopic remission Group 1 was 2/17 [12%], Group 2 was 3/19 [16%], Group 3 was 4/15 [27%] [Group 1 vs 3 p = 0.38]. Mucosal healing [Mayo 0] was achieved only in Group 3 [3/15, 20%] vs 0/36 FT patients [p = 0.022]. Exacerbation of disease occurred in 3/17 [17.6%] of Group 1, 4/19 [21.1%] of Group 2, and 1/15 [6.7%] of Group 3 [Group 2 vs 3, p = 0.35]. CONCLUSIONS: UCED alone appeared to achieve higher clinical remission and mucosal healing than single donor FT with or without diet. The study was stopped for futility by a safety monitoring board.