Butyrate enhances Clostridioides difficile sporulation in vitro.

Short-chain fatty acids (SCFAs) are products of bacterial fermentation that help maintain important gut functions such as maintenance of the intestinal barrier, cell signaling, and immune homeostasis. The main SCFAs acetate, propionate, and butyrate have demonstrated beneficial effects for the host, including its importance in alleviating infections caused by pathogens such as Clostridioides difficile. Despite the potential role of SCFAs in mitigating C. difficile infection, their direct effect on C. difficile remains unclear. Through a set of in vitro experiments, we investigated how SCFAs influence C. difficile growth, sporulation, and toxin production. Similar to previous studies, we observed that butyrate decreased growth of C. difficile strain 630 in a dose-dependent manner. The presence of butyrate also increased C. difficile sporulation, with minimal increases in toxin production. RNA-Seq analysis validated our experimental results, demonstrating increased expression of sporulation-related genes in conjunction with changes in metabolic and regulatory genes, such as a putative carbon starvation protein, CstA. Collectively, these data suggest that butyrate may induce alternative C. difficile survival pathways, modifying its growth ability and virulence to persist in the gut environment. IMPORTANCE Several studies suggest that butyrate may modulate gut infections, such as reducing inflammation caused by the healthcare-associated Clostridioides difficile. While studies in both animal models and human studies correlate high levels of butyrate with reduced C. difficile burden, the direct impact of butyrate on C. difficile remains unclear. Our study demonstrates that butyrate directly influences C. difficile by increasing its sporulation and modifying its metabolism, potentially using butyrate as a biomarker to shift survival strategies in a changing gut environment. These data point to additional therapeutic approaches to combat C. difficile in a butyrate-directed manner.

Butyrate enhances Clostridioides difficile sporulation in vitro.

Short chain fatty acids (SCFAs) are products of bacterial fermentation that help maintain important gut functions such as the intestinal barrier, signaling, and immune homeostasis. The main SCFAs acetate, propionate, and butyrate have demonstrated beneficial effects for the host, including importance in combatting infections caused by pathogens such as Clostridioides difficile . Despite the potential role of SCFAs in mitigating C. difficile infection, their direct effect on C. difficile remains unclear. Through a set of in vitro experiments, we investigated how SCFAs influence C. difficile growth, sporulation, and toxin production. Similar to previous studies, we observed that butyrate decreased growth of C. difficile strain 630 in a dose-dependent manner. The presence of butyrate also increased C. difficile sporulation, with minimal increases in toxin production. RNA-Seq analysis validated our experimental results, demonstrating increased expression of sporulation-related genes in conjunction with alternative metabolic and related C. difficile regulatory pathways, such as the carbon catabolite repressor, CcpA. Collectively, these data suggest that butyrate may signal alternative C. difficile metabolic pathways, thus modifying its growth and virulence to persist in the gut environment. IMPORTANCE: Several studies suggest that butyrate may be important in alleviating gut infections, such as reducing inflammation caused by the healthcare-associated Clostridioides difficile . While studies in both animal models and human studies correlate high levels of butyrate with reduced C. difficile burden, the direct impact of butyrate on C. difficile remains unclear. Our study demonstrates that butyrate directly influences C. difficile by increasing its sporulation and modifying its metabolism, potentially using butyrate as a biomarker to shift survival strategies in a changing gut environment. These data point to additional therapeutic approaches to combat C. difficile in a butyrate-directed manner.

Beneficial effects of fecal microbiota transplantation in recurrent Clostridioides difficile infection.

Fecal microbiota transplantation (FMT) is highly effective in preventing recurrent Clostridioides difficile infection (rCDI). However, the mechanisms underpinning its clinical efficacy are incompletely understood. Herein, we provide an overview of rCDI pathogenesis followed by a discussion of potential mechanisms of action focusing on the current understanding of trans-kingdom microbial, metabolic, immunological, and epigenetic mechanisms. We then outline the current research gaps and offer methodological recommendations for future studies to elevate the quality of research and advance knowledge translation. By combining interventional trials with multiomics technology and host and environmental factors, analyzing longitudinally collected biospecimens will generate results that can be validated with animal and other models. Collectively, this will confirm causality and improve translation, ultimately to develop targeted therapies to replace FMT.

Diversity and Prevalence of Clostridium innocuum in the Human Gut Microbiota.

Clostridia are a polyphyletic group of Gram-positive, spore-forming anaerobes in the Firmicutes phylum that significantly impact metabolism and functioning of the human gastrointestinal tract. Recently, Clostridia were divided into two separate classes, Clostridia and Erysipelotrichia, based on phenotypic and 16S rRNA gene-based differences. While Clostridia include many well-known pathogenic bacteria, Erysipelotrichia remain relatively uncharacterized, particularly regarding their role as a pathogen versus commensal. Despite wide recognition as a commensal, the erysipelotrichial species Clostridium innocuum has recently been associated with various disease states. To further understand the ecological and potential virulent role of C. innocuum, we conducted a genomic comparison across 38 C. innocuum isolates and 194 publicly available genomes. Based on colony morphology, we isolated multiple C. innocuum cultivars from the feces of healthy human volunteers (n = 5). Comparison of the 16S rRNA gene of our isolates against publicly available microbiota data sets in healthy individuals suggests a high prevalence of C. innocuum across the human population (>80%). Analysis of single nucleotide polymorphisms (SNPs) across core genes and average nucleotide identify (ANI) revealed the presence of four clades among all available genomes (n = 232 total). Investigation of carbohydrate and protein utilization pathways, including comparison against the carbohydrate-activating enzyme (CAZyme) database, demonstrated inter- and intraclade differences that were further substantiated in vitro. Collectively, these data indicate genetic variance within the C. innocuum species that may help clarify its role in human disease and health. IMPORTANCE Clostridia are a group of medically important anaerobes as both commensals and pathogens. Recently, a new class of Erysipelotrichia containing a number of reassigned clostridial species has emerged, including Clostridium innocuum. Recent studies have implicated C. innocuum as a potential causative agent of diarrhea in patients from whom Clostridioides difficile could not be isolated. Using genomic and in vitro comparison, this study sought to characterize C. innocuum in the healthy human gut. Our analyses suggest that C. innocuum is a highly prevalent and diverse species, demonstrating clade-specific differences in metabolism and potential virulence. Collectively, this study is the first investigation into a broader description of C. innocuum as a human gut inhabitant.

Clostridioides difficile: innovations in target discovery and potential for therapeutic success.

INTRODUCTION: Clostridioides difficile infection (CDI) remains a worldwide clinical problem. Increased incidence of primary infection, occurrence of hypertoxigenic ribotypes, and more frequent occurrence of drug resistant, recurrent, and non-hospital CDI, emphasizes the urgent unmet need of discovering new therapeutic targets. AREAS COVERED: We searched PubMed and Web of Science databases for articles identifying novel therapeutic targets or treatments for C. difficile from 2001 to 2021. We present an updated review on current preclinical efforts on designing inhibitory compounds against these drug targets and indicate how these could become the focus of future therapeutic approaches. We also evaluate the increasing exploitability of gut microbial-derived metabolites and host-derived therapeutics targeting VEGF-A, immune targets and pathways, ion transporters, and microRNAs as anti-C. difficile therapeutics, which have yet to reach clinical trials. Our review also highlights the therapeutic potential of re-purposing currently available agents . We conclude by considering translational hurdles and possible strategies to mitigate these problems. EXPERT OPINION: Considerable progress has been made in the development of new anti-CDI drug candidates. Nevertheless, a greater comprehension of CDI pathogenesis and host-microbe interactions is beginning to uncover potential novel therapeutic targets, which can be exploited to plug gaps in the CDI drug discovery pipeline.

Multiomics Profiling Reveals Signatures of Dysmetabolism in Urban Populations in Central India.

BACKGROUND: Non-communicable diseases (NCDs) have become a major cause of morbidity and mortality in India. Perturbation of host-microbiome interactions may be a key mechanism by which lifestyle-related risk factors such as tobacco use, alcohol consumption, and physical inactivity may influence metabolic health. There is an urgent need to identify relevant dysmetabolic traits for predicting risk of metabolic disorders, such as diabetes, among susceptible Asian Indians where NCDs are a growing epidemic. METHODS: Here, we report the first in-depth phenotypic study in which we prospectively enrolled 218 adults from urban and rural areas of Central India and used multiomic profiling to identify relationships between microbial taxa and circulating biomarkers of cardiometabolic risk. Assays included fecal microbiota analysis by 16S ribosomal RNA gene amplicon sequencing, quantification of serum short chain fatty acids by gas chromatography-mass spectrometry, and multiplex assaying of serum diabetic proteins, cytokines, chemokines, and multi-isotype antibodies. Sera was also analysed for N-glycans and immunoglobulin G Fc N-glycopeptides. RESULTS: Multiple hallmarks of dysmetabolism were identified in urbanites and young overweight adults, the majority of whom did not have a known diagnosis of diabetes. Association analyses revealed several host-microbe and metabolic associations. CONCLUSIONS: Host-microbe and metabolic interactions are differentially shaped by body weight and geographic status in Central Indians. Further exploration of these links may help create a molecular-level map for estimating risk of developing metabolic disorders and designing early interventions.

Fecal Microbiota Transplantation for Recurrent Clostridioides difficile Infection Associates With Functional Alterations in Circulating microRNAs.

BACKGROUND AND AIMS: The molecular mechanisms underlying successful fecal microbiota transplantation (FMT) for recurrent Clostridioides difficile infection (rCDI) remain poorly understood. The primary objective of this study was to characterize alterations in microRNAs (miRs) following FMT for rCDI. METHODS: Sera from 2 prospective multicenter randomized controlled trials were analyzed for miRNA levels with the use of the Nanostring nCounter platform and quantitative reverse-transcription (RT) polymerase chain reaction (PCR). In addition, rCDI-FMT and toxin-treated animals and ex vivo human colonoids were used to compare intestinal tissue and circulating miRs. miR inflammatory gene targets in colonic epithelial and peripheral blood mononuclear cells were evaluated by quantitative PCR (qPCR) and 3'UTR reporter assays. Colonic epithelial cells were used for mechanistic, cytoskeleton, cell growth, and apoptosis studies. RESULTS: miRNA profiling revealed up-regulation of 64 circulating miRs 4 and 12 weeks after FMT compared with screening, of which the top 6 were validated in the discovery cohort by means of RT-qPCR. In a murine model of relapsing-CDI, RT-qPCR analyses of sera and cecal RNA extracts demonstrated suppression of these miRs, an effect reversed by FMT. In mouse colon and human colonoids, C difficile toxin B (TcdB) mediated the suppressive effects of CDI on miRs. CDI dysregulated DROSHA, an effect reversed by FMT. Correlation analyses, qPCR ,and 3'UTR reporter assays revealed that miR-23a, miR-150, miR-26b, and miR-28 target directly the 3'UTRs of IL12B, IL18, FGF21, and TNFRSF9, respectively. miR-23a and miR-150 demonstrated cytoprotective effects against TcdB. CONCLUSIONS: These results provide novel and provocative evidence that modulation of the gut microbiome via FMT induces alterations in circulating and intestinal tissue miRs. These findings contribute to a greater understanding of the molecular mechanisms underlying FMT and identify new potential targets for therapeutic intervention in rCDI.

mSphere of Influence: Translating Gut Microbiome Studies To Benefit Human Health.

Anna M. Seekatz works in the field of the gut microbiome as it related to infectious diseases. In this "mSphere of Influence" article, she reflects on how two studies, "The impact of a consortium of fermented milk strains on the gut microbiome of gnotobiotic mice and monozygotic twins" (N. P. McNulty, T. Yatsunenko, A. Hsiao, et al., Sci Transl Med 3:106ra106, 2011) and "High-throughput DNA sequence analysis reveals stable engraftment of gut microbiota following transplantation of previously frozen fecal bacteria" (M. J. Hamilton, A. R. Weingarden, T. Unno, A. Khoruts, and M. J. Sadowsky, Gut Microbes 4:125-135, 2013), shaped how she approaches interpreting microbiome studies.

Dilution as a Solution: Targeting Microbial Populations with a Simplified Dilution Strategy.

The gut microbiota is an integral part of maintaining resistance against infection by Clostridioides (Clostridium) difficile, a pathogen of increasing concern in both health care and community settings. The recent article by J. M. Auchtung, E. C. Preisner, J. Collins, A. I. Lerma, and R. A. Britton (mSphere 5:e00387-20, 2020, https://doi.org/10.1128/mSphere.00387-20) demonstrates an innovative approach to identify microbes that inhibit C. difficile by employing a dilution scheme to test different microbial mixtures in vitro and in vivo This type of approach can advance the identification and validation of specific microbes that elicit functions of interest for many conditions involving the microbiota, of which the complexity and variability can often complicate causality.

Interleukin-22-mediated host glycosylation prevents Clostridioides difficile infection by modulating the metabolic activity of the gut microbiota.

The involvement of host immunity in the gut microbiota-mediated colonization resistance to Clostridioides difficile infection (CDI) is incompletely understood. Here, we show that interleukin (IL)-22, induced by colonization of the gut microbiota, is crucial for the prevention of CDI in human microbiota-associated (HMA) mice. IL-22 signaling in HMA mice regulated host glycosylation, which enabled the growth of succinate-consuming bacteria Phascolarctobacterium spp. within the gut microbiome. Phascolarctobacterium reduced the availability of luminal succinate, a crucial metabolite for the growth of C. difficile, and therefore prevented the growth of C. difficile. IL-22-mediated host N-glycosylation is likely impaired in patients with ulcerative colitis (UC) and renders UC-HMA mice more susceptible to CDI. Transplantation of healthy human-derived microbiota or Phascolarctobacterium reduced luminal succinate levels and restored colonization resistance in UC-HMA mice. IL-22-mediated host glycosylation thus fosters the growth of commensal bacteria that compete with C. difficile for the nutritional niche.

Epidemiological and Microbiome Associations Between Klebsiella pneumoniae and Vancomycin-Resistant Enterococcus Colonization in Intensive Care Unit Patients.

BACKGROUND: Prior colonization by Klebsiella pneumoniae and vancomycin-resistant Enterococci (VRE) is associated with subsequent infection, particularly in intensive care unit (ICU) populations. Screening for VRE colonization, but not K. pneumoniae, is routinely performed in some health care systems. Identification of patient factors associated with K. pneumoniae colonization could enable infection prevention. METHODS: ICU patients were screened for VRE and K. pneumoniae by rectal swab culture over 2 time periods: July-October 2014 (n = 1209) and January-May 2016 (n = 1243). Patient demographics, baseline laboratory data, comorbidities, and outcomes were analyzed. 16S rRNA gene-based analysis was performed on a subset of patients (n = 248) to identify microbiota characteristics associated with VRE and K. pneumoniae colonization. RESULTS: K. pneumoniae colonization (17.3% of patients in the 2014 cohort, 7.3% in 2016) was significantly associated with VRE colonization in multivariable analysis (P = .03 in 2016; P = .08 in 2014). VRE colonization was associated with poor underlying health, whereas K. pneumoniae colonization was associated with advanced age. The most prevalent operational taxonomic units were Escherichia coli /Shigella spp., Klebsiella, and Enterococcus, consistent with high rates of detectable K. pneumoniae and VRE by culture. Microbial community structure in noncolonized patients was significantly different from those with VRE, K. pneumoniae, or both, attributable to differences in the relative abundance of Klebsiella and Enterococcus. CONCLUSIONS: K. pneumoniae co-colonizes with VRE and is a predominant taxon in ICU patients, but colonization was not associated with significant comorbidities. Screening for K. pneumoniae and VRE simultaneously could be an efficient approach for novel infection prevention strategies.

Introduction to the special issue for The anaerobe society of the America's 14th Biennial congress in Las Vegas.

In June 2018, the Anaerobe Society of the America's (ASA) held their 14th Biennial Congress in Las Vegas, Nevada. The Congress was attended by over 200 individuals from many different countries. The focus of the meeting was the fast-growing area of anaerobes in human and animal infectious disease, computational tools to understand basic biology and therapeutic development, the role of anaerobes in the microbiome, and clinical trials of novel bacterial-based therapies. To strengthen the community of researchers working on anaerobes, the congress held two training workshops on clinical bacteriology and anaerobes in the microbiome, several networking events, as well as a dinner which honored the lifetime achievement award given to Ellen Jo Baron. The meeting was also attended by the grandfather of anaerobic bacteriology and the founder of (ASA), Sydney Finegold, at the age of 97. In all, there was a broad diversity of research presented that showed new ways that anaerobes play a important role in health and disease.

Spatial and Temporal Analysis of the Stomach and Small-Intestinal Microbiota in Fasted Healthy Humans.

Although the microbiota in the proximal gastrointestinal (GI) tract have been implicated in health and disease, much about these microbes remains understudied compared to those in the distal GI tract. This study characterized the microbiota across multiple proximal GI sites over time in healthy individuals. As part of a study of the pharmacokinetics of oral mesalamine administration, healthy, fasted volunteers (n = 8; 10 observation periods total) were orally intubated with a four-lumen catheter with multiple aspiration ports. Samples were taken from stomach, duodenal, and multiple jejunal sites, sampling hourly (≤7 h) to measure mesalamine (administered at t = 0), pH, and 16S rRNA gene-based composition. We observed a predominance of Firmicutes across proximal GI sites, with significant variation compared to stool. The microbiota was more similar within individuals over time than between subjects, with the fecal microbiota being unique from that of the small intestine. The stomach and duodenal microbiota displayed highest intraindividual variability compared to jejunal sites, which were more stable across time. We observed significant correlations in the duodenal microbial composition with changes in pH; linear mixed models identified positive correlations with multiple Streptococcus operational taxonomic units (OTUs) and negative correlations with multiple Prevotella and Pasteurellaceae OTUs. Few OTUs correlated with mesalamine concentration. The stomach and duodenal microbiota exhibited greater compositional dynamics than the jejunum. Short-term fluctuations in the duodenal microbiota were correlated with pH. Given the unique characteristics and dynamics of the proximal GI tract microbiota, it is important to consider these local environments in health and disease states.IMPORTANCE The gut microbiota are linked to a variety of gastrointestinal diseases, including inflammatory bowel disease. Despite this importance, microbiota dynamics in the upper gastrointestinal tract are understudied. Our article seeks to understand what factors impact microbiota dynamics in the healthy human upper gut. We found that the upper gastrointestinal tract contains consistently prevalent bacterial OTUs that dominate the overall community. Microbiota variability is highest in the stomach and duodenum and correlates with pH.

Gut Microbiota and Clinical Features Distinguish Colonization With Klebsiella pneumoniae Carbapenemase-Producing Klebsiella pneumoniae at the Time of Admission to a Long-term Acute Care Hospital.

BACKGROUND: Identification of gut microbiota features associated with antibiotic-resistant bacterial colonization may reveal new infection prevention targets. METHODS: We conducted a matched, case-control study of long-term acute care hospital (LTACH) patients to identify gut microbiota and clinical features associated with colonization by Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp), an urgent antibiotic resistance threat. Fecal or rectal swab specimens were collected and tested for KPC-Kp; 16S rRNA gene-based sequencing was performed. Comparisons were made between cases and controls in calibration and validation subsamples using microbiota similarity indices, logistic regression, and unit-weighted predictive models. RESULTS: Case (n = 32) and control (n = 99) patients had distinct fecal microbiota communities, but neither microbiota diversity nor inherent clustering into community types distinguished case and control specimens. Comparison of differentially abundant operational taxonomic units (OTUs) revealed 1 OTU associated with case status in both calibration (n = 51) and validation (n = 80) subsamples that matched the canonical KPC-Kp strain ST258. Permutation analysis using the presence or absence of OTUs and hierarchical logistic regression identified 2 OTUs (belonging to genus Desulfovibrio and family Ruminococcaceae) associated with KPC-Kp colonization. Among clinical variables, the presence of a decubitus ulcer alone was independently and consistently associated with case status. Combining the presence of the OTUs Desulfovibrio and Ruminococcaceae with decubitus ulcer increased the likelihood of KPC-Kp colonization to >38% in a unit-weighted predictive model. CONCLUSIONS: We identified microbiota and clinical features that distinguished KPC-Kp gut colonization in LTACH patients, a population particularly susceptible to KPC-Kp infection. These features may warrant further investigation as markers of risk for KPC-Kp colonization.

Presence of multiple Clostridium difficile strains at primary infection is associated with development of recurrent disease.

Recurrence of Clostridium difficile infection (CDI) places a major burden on the healthcare system. Previous studies have suggested that specific C. difficile strains, or ribotypes, are associated with severe disease and/or recurrence. However, in some patients a new strain is detected in subsequent infections, complicating longitudinal studies focused on strain differences that may contribute to disease outcome. We examined ribotype composition over time in patients who did or did not develop recurrence to examine infection with multiple C. difficile ribotypes (mixed infection), during the course of infection. Using a retrospective patient cohort, we isolated and ribotyped a median of 36 C. difficile colonies from 61 patients (105 total samples) at initial infection, recurrence (a second case of CDI within 15-56 days of initial infection), and reinfection (a second case of CDI after 56 days of initial infection). We observed mixed infection in 78.6% of samples at initial infection in patients who went on to develop recurrence compared to 18.1% of patients who did not, and mixed infection remained associated with subsequent recurrence after adjusting for gender and prior antibiotic exposure (OR 3.5, 95% CI 1.3-9.4, P = .015). In patients who were sampled longitudinally (44 consecutive events in 32 patients), the dominant ribotype changed in 31.8% of consecutive samples and the newly dominant ribotype was not detected in prior samples from that patient. Our results suggest that mixed C. difficile infection is more prevalent than previously demonstrated and potentially a marker of susceptibility to recurrence.

Restoration of short chain fatty acid and bile acid metabolism following fecal microbiota transplantation in patients with recurrent Clostridium difficile infection.

A significant proportion of individuals develop recurrent Clostridium difficile infection (CDI) following initial disease. Fecal microbiota transplantation (FMT), a highly effective treatment method for recurrent CDI, has been demonstrated to induce microbiota recovery. One of the proposed functions associated with restoration of colonization resistance against C. difficile has been recovery of bile acid metabolism. In this study, we aimed to assess recovery of short chain fatty acids (SCFAs) in addition to bile acids alongside microbial community structure in six patients with recurrent CDI following treatment with FMT over time. Using 16S rRNA gene-based sequencing, we observed marked similarity of the microbiota between recipients following FMT (n = 6, sampling up to 6 months post-FMT) and their respective donors. Sustained increases in the levels of the SCFAs butyrate, acetate, and propionate were observed post-FMT, and variable recovery over time was observed in the secondary bile acids deoxycholate and lithocholate. To correlate these changes with specific microbial taxa at an individual level, we applied a generalized estimating equation approach to model metabolite concentrations with the presence of specific members of the microbiota. Metabolites that increased following FMT were associated with bacteria classified within the Lachnospiraceae, Ruminococcaceae, and unclassified Clostridiales families. In contrast, members of these taxa were inversely associated with primary bile acids. The longitudinal aspect of this study allowed us to characterize individualized patterns of recovery, revealing variability between and within patients following FMT.

Comparison of stool versus rectal swab samples and storage conditions on bacterial community profiles.

BACKGROUND: Sample collection for gut microbiota analysis from in-patients can be challenging. Collection method and storage conditions are potential sources of variability. In this study, we compared the bacterial microbiota from stool stored under different conditions, as well as stool and swab samples, to assess differences due to sample storage conditions and collection method. METHODS: Using bacterial 16S rRNA gene sequence analysis, we compared the microbiota profiles of stool samples stored and collected under various conditions. Stool samples (2 liquid, 1 formed) from three different patients at two hospitals were each evaluated under the following conditions: immediately frozen at -80°C, stored at 4°C for 12-48 hours before freezing at -80°C and stored at -20°C with 1-2 thaw cycles before storage at -80°C. Additionally, 8 stool and 30 rectal swab samples were collected from 8 in-patients at one hospital. Microbiota differences were assessed using the Yue and Clayton dissimilarity index (θYC distance) and analysis of molecular variance (AMOVA). RESULTS: Regardless of the storage conditions, the bacterial communities of aliquots from the same stool samples were very similar based on θYC distances (median intra-sample θYC distance: 0.035, IQR: 0.015-0.061) compared to aliquots from different stool samples (median inter-sample θYC distance: 0.93, IQR: 0.85-0.97) (Wilcoxon test p-value: <0.0001). For the stool and rectal swab comparison, samples from different patients, regardless of sample collection method, were significantly different (AMOVA p-values: <0.001-0.029) compared to no significant difference between all stool and swab samples (AMOVA p-value: 0.976). The θYC dissimilarity index between swab and stool samples was significantly lower within individuals (median 0.17, IQR: 0.10-0.27) than between individuals (median 0.93, IQR: 0.85-0.97) (Wilcoxon test p-value: <0.0001), indicating minimal differences between stool and swab samples collected from the same individual over the sampling period. CONCLUSION: For gastrointestinal microbiota studies based on bacterial 16S rRNA gene sequence analysis, interim stool sample storage at 4 °C or -20 °C, rather than immediate storage at -80 °C, does not significantly alter results. Additionally, stool and rectal swab microbiotas from the same subject were highly similar, indicating that these sampling methods could be used interchangeably to assess the community structure of the distal GI tract.

A Dietary Fiber-Deprived Gut Microbiota Degrades the Colonic Mucus Barrier and Enhances Pathogen Susceptibility.

Despite the accepted health benefits of consuming dietary fiber, little is known about the mechanisms by which fiber deprivation impacts the gut microbiota and alters disease risk. Using a gnotobiotic mouse model, in which animals were colonized with a synthetic human gut microbiota composed of fully sequenced commensal bacteria, we elucidated the functional interactions between dietary fiber, the gut microbiota, and the colonic mucus barrier, which serves as a primary defense against enteric pathogens. We show that during chronic or intermittent dietary fiber deficiency, the gut microbiota resorts to host-secreted mucus glycoproteins as a nutrient source, leading to erosion of the colonic mucus barrier. Dietary fiber deprivation, together with a fiber-deprived, mucus-eroding microbiota, promotes greater epithelial access and lethal colitis by the mucosal pathogen, Citrobacter rodentium. Our work reveals intricate pathways linking diet, the gut microbiome, and intestinal barrier dysfunction, which could be exploited to improve health using dietary therapeutics.