The microbiome regulates amygdala-dependent fear recall.

The amygdala is a key brain region that is critically involved in the processing and expression of anxiety and fear-related signals. In parallel, a growing number of preclinical and human studies have implicated the microbiome-gut-brain in regulating anxiety and stress-related responses. However, the role of the microbiome in fear-related behaviours is unclear. To this end we investigated the importance of the host microbiome on amygdala-dependent behavioural readouts using the cued fear conditioning paradigm. We also assessed changes in neuronal transcription and post-transcriptional regulation in the amygdala of naive and stimulated germ-free (GF) mice, using a genome-wide transcriptome profiling approach. Our results reveal that GF mice display reduced freezing during the cued memory retention test. Moreover, we demonstrate that under baseline conditions, GF mice display altered transcriptional profile with a marked increase in immediate-early genes (for example, Fos, Egr2, Fosb, Arc) as well as genes implicated in neural activity, synaptic transmission and nervous system development. We also found a predicted interaction between mRNA and specific microRNAs that are differentially regulated in GF mice. Interestingly, colonized GF mice (ex-GF) were behaviourally comparable to conventionally raised (CON) mice. Together, our data demonstrates a unique transcriptional response in GF animals, likely because of already elevated levels of immediate-early gene expression and the potentially underlying neuronal hyperactivity that in turn primes the amygdala for a different transcriptional response. Thus, we demonstrate for what is to our knowledge the first time that the presence of the host microbiome is crucial for the appropriate behavioural response during amygdala-dependent memory retention.

The influence of rosuvastatin on the gastrointestinal microbiota and host gene expression profiles.

Statins are the most widely prescribed medications worldwide for the treatment of hypercholesterolemia. They inhibit the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-R), an enzyme involved in cholesterol synthesis in higher organisms and in isoprenoid biosynthesis in some bacteria. We hypothesized that statins may influence the microbial community in the gut through either direct inhibition or indirect mechanisms involving alterations to host responses. We therefore examined the impact of rosuvastatin (RSV) on the community structure of the murine gastrointestinal microbiota. RSV was orally administered to mice and the effects on the gut microbiota, host bile acid profiles, and markers of inflammation were analyzed. RSV significantly influenced the microbial community in both the cecum and feces, causing a significant decrease in α-diversity in the cecum and resulting in a reduction of several physiologically relevant bacterial groups. RSV treatment of mice significantly affected bile acid metabolism and impacted expression of inflammatory markers known to influence microbial community structure (including RegIIIγ and Camp) in the gut. This study suggests that a commonly used statin (RSV) leads to an altered gut microbial composition in normal mice with attendant impacts on local gene expression profiles, a finding that should prompt further studies to investigate the implications of statins for gut microbiota stability and health in humans.NEW & NOTEWORTHY This work demonstrates that rosuvastatin administration in mice affects the gastrointestinal microbiota, influences bile acid metabolism, and alters transcription of genes encoding factors involved in gut homeostasis and immunity in the gastrointestinal tract.

The altered gut microbiota in adults with cystic fibrosis.

BACKGROUND: Cystic Fibrosis (CF) is an autosomal recessive disease that affects the function of a number of organs, principally the lungs, but also the gastrointestinal tract. The manifestations of cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction in the gastrointestinal tract, as well as frequent antibiotic exposure, undoubtedly disrupts the gut microbiota. To analyse the effects of CF and its management on the microbiome, we compared the gut microbiota of 43 individuals with CF during a period of stability, to that of 69 non-CF controls using 454-pyrosequencing of the 16S rRNA gene. The impact of clinical parameters, including antibiotic therapy, on the results was also assessed. RESULTS: The CF-associated microbiome had reduced microbial diversity, an increase in Firmicutes and a reduction in Bacteroidetes compared to the non-CF controls. While the greatest number of differences in taxonomic abundances of the intestinal microbiota was observed between individuals with CF and the healthy controls, gut microbiota differences were also reported between people with CF when grouped by clinical parameters including % predicted FEV1 (measure of lung dysfunction) and the number of intravenous (IV) antibiotic courses in the previous 12 months. Notably, CF individuals presenting with severe lung dysfunction (% predicted FEV1 ≤ 40%) had significantly (p < 0.05) reduced gut microbiota diversity relative to those presenting with mild or moderate dysfunction. A significant negative correlation (-0.383, Simpson's Diversity Index) was also observed between the number of IV antibiotic courses and gut microbiota diversity. CONCLUSIONS: This is one of the largest single-centre studies on gut microbiota in stable adults with CF and demonstrates the significantly altered gut microbiota, including reduced microbial diversity seen in CF patients compared to healthy controls. The data show the impact that CF and it's management have on gut microbiota, presenting the opportunity to develop CF specific probiotics to minimise microbiota alterations.

Differential expression of key regulators of Toll-like receptors in ulcerative colitis and Crohn's disease: a role for Tollip and peroxisome proliferator-activated receptor gamma?

The innate immune system is currently seen as the probable initiator of events which culminate in the development of inflammatory bowel disease (IBD) with Toll-like receptors (TLRs) known to be involved in this disease process. Many regulators of TLRs have been described, and dysregulation of these may also be important in the pathogenesis of IBD. The aim of this study was to perform a co-ordinated analysis of the expression levels of both key intestinal TLRs and their inhibitory proteins in the same IBD cohorts, both ulcerative colitis (UC) and Crohn's disease (CD), in order to evaluate the potential roles of these proteins in the pathogenesis of IBD. Of the six TLRs (TLRs 1, 2, 4, 5, 6 and 9) examined, only TLR-4 was increased significantly in IBD, specifically in active UC. In contrast, differential alterations in expression of TLR inhibitory proteins were observed. A20 and suppressor of cytokine signalling 1 (SOCS1) were increased only in active UC while interleukin-1 receptor-associated kinase 1 (IRAK-m) and B cell lymphoma 3 protein (Bcl-3) were increased in both active UC and CD. In contrast, expression of both peroxisome proliferator-activated receptor gamma (PPARγ) and Toll interacting protein (Tollip) was decreased in both active and inactive UC and CD and at both mRNA and protein levels. In addition, expression of both PPARγ and A20 expression was increased by stimulation of a colonic epithelial cell line Caco-2 with both TLR ligands and commensal bacterial strains. These data suggest that IBD may be associated with distinctive changes in TLR-4 and TLR inhibitory proteins, implying that alterations in these may contribute to the pathogenesis of IBD.

Spatial variation of the colonic microbiota in patients with ulcerative colitis and control volunteers.

OBJECTIVES: The relevance of spatial composition in the microbial changes associated with UC is unclear. We coupled luminal brush samples, mucosal biopsies and laser capture microdissection with deep sequencing of the gut microbiota to develop an integrated spatial assessment of the microbial community in controls and UC. DESIGN: A total of 98 samples were sequenced to a mean depth of 31,642 reads from nine individuals, four control volunteers undergoing routine colonoscopy and five patients undergoing surgical colectomy for medically-refractory UC. Samples were retrieved at four colorectal locations, incorporating the luminal microbiota, mucus gel layer and whole mucosal biopsies. RESULTS: Interpersonal variability accounted for approximately half of the total variance. Surprisingly, within individuals, asymmetric Eigenvector map analysis demonstrated differentiation between the luminal and mucus gel microbiota, in both controls and UC, with no differentiation between colorectal regions. At a taxonomic level, differentiation was evident between both cohorts, as well as between the luminal and mucosal compartments, with a small group of taxa uniquely discriminating the luminal and mucosal microbiota in colitis. There was no correlation between regional inflammation and a breakdown in this spatial differentiation or bacterial diversity. CONCLUSIONS: Our study demonstrates a conserved spatial structure to the colonic microbiota, differentiating the luminal and mucosal communities, within the context of marked interpersonal variability. While elements of this structure overlap between UC and control volunteers, there are differences between the two groups, both in terms of the overall taxonomic composition and how spatial structure is ascribable to distinct taxa.

The role of pure iterative reconstruction in conventional dose CT enterography.

PURPOSE: Pure iterative reconstruction (Pure IR) has been proposed as a solution to improve diagnostic quality of low dose CT images. We assess the performance of model based iterative reconstruction (MBIR) in improving conventional dose CT enterography (CTE) images. METHODS: 43 Crohn's patients (27 female) (38.5 ± 12.98 years) referred for CTE were included. Images were reconstructed with pure IR (MBIR, General Electric Healthcare) in addition to standard department protocol (reconstructed with hybrid iterative reconstruction (Hybrid IR) [60% filtered back projection/40% adaptive statistical IR (General Electric Healthcare)]. Image quality was assessed objectively and subjectively at 6 anatomical levels. Clinical interpretation was undertaken in consensus by 2 blinded radiologists along with 2 non-blinded readers ('gold standard'). Results were analyzed using Statistical Package for Social Scientists. RESULTS: Mean effective radiation dose was 6.05 ± 2.84 mSv (size specific dose estimates 9.25 ± 2.9 mGy). Objective and subjective assessment yielded 6106 data points. Pure IR images significantly outperformed those using standard reconstruction techniques across all subjective (p < 0.001 for all comparisons) (noise, contrast resolution, spatial resolution, streak artifact, axial diagnostic acceptability, coronal diagnostic acceptability) and objective (p < 0.004) (noise, signal-to-noise ratio) parameters. Clinical reads of the pure IR images agreed more closely with the gold standard reads than the hybrid IR image reads in terms of overall Crohn's activity grade (κ = 0.630, 0.308) and detection of acute complications (κ = 1.0, 0.896). Results were comparable for bowel wall disease severity assessment (κ = 0.523, 0.593). CONCLUSIONS: Pure IR considerably improves image quality of conventional dose CTE images and therefore its use should be expanded beyond low dose protocols to improving image quality at conventional dose CT imaging.

Cognitive performance in irritable bowel syndrome: evidence of a stress-related impairment in visuospatial memory.

BACKGROUND: Central nervous system (CNS) dysfunction is a prominent feature of the functional gastrointestinal (GI) disorder, irritable bowel syndrome (IBS). However, the neurobiological and cognitive consequences of key pathophysiological features of IBS, such as stress-induced changes in hypothalamic-pituitary-adrenal (HPA)-axis functioning, is unknown. Our aim was to determine whether IBS is associated with cognitive impairment, independently of psychiatric co-morbidity, and whether cognitive performance is related to HPA-axis function. METHOD: A cross-sectional sample of 39 patients with IBS, a disease control group of 18 patients with Crohn's disease (CD) in clinical remission and 40 healthy age- and IQ-matched control participants were assessed using the Paired Associates Learning (PAL), Intra-Extra Dimensional Set Shift (IED) and Spatial Working Memory (SWM) tests from the Cambridge Neuropsychological Test Automated Battery (CANTAB) and a computerized Stroop test. HPA-axis function was determined by measuring the cortisol awakening response (CAR). RESULTS: IBS patients exhibited a subtle visuospatial memory deficit at the PAL six- pattern stage (p = 0.03), which remained after psychiatric co-morbidity was controlled for (p = 0.04). Morning cortisol levels were lower in IBS (p = 0.04) and significantly associated with visuospatial memory performance within IBS only (p = 0.02). CONCLUSIONS: For the first time, altered cognitive function on a hippocampal-mediated test of visuospatial memory, which was related to cortisol levels and independent of psychiatric co-morbidity, has been identified in IBS. Visuospatial memory impairment may be a common, but currently neglected, component of IBS. Further elucidation of the nature of this impairment may lead to a greater understanding of the underlying pathophysiology of IBS, and may provide novel therapeutic approaches.

The microbiome-gut-brain axis during early life regulates the hippocampal serotonergic system in a sex-dependent manner.

Bacterial colonisation of the intestine has a major role in the post-natal development and maturation of the immune and endocrine systems. These processes are key factors underpinning central nervous system (CNS) signalling. Regulation of the microbiome-gut-brain axis is essential for maintaining homeostasis, including that of the CNS. However, there is a paucity of data pertaining to the influence of microbiome on the serotonergic system. Germ-free (GF) animals represent an effective preclinical tool to investigate such phenomena. Here we show that male GF animals have a significant elevation in the hippocampal concentration of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid, its main metabolite, compared with conventionally colonised control animals. Moreover, this alteration is sex specific in contrast with the immunological and neuroendocrine effects which are evident in both sexes. Concentrations of tryptophan, the precursor of serotonin, are increased in the plasma of male GF animals, suggesting a humoral route through which the microbiota can influence CNS serotonergic neurotransmission. Interestingly, colonisation of the GF animals post weaning is insufficient to reverse the CNS neurochemical consequences in adulthood of an absent microbiota in early life despite the peripheral availability of tryptophan being restored to baseline values. In addition, reduced anxiety in GF animals is also normalised following restoration of the intestinal microbiota. These results demonstrate that CNS neurotransmission can be profoundly disturbed by the absence of a normal gut microbiota and that this aberrant neurochemical, but not behavioural, profile is resistant to restoration of a normal gut flora in later life.

Bcl-3 deficiency protects against dextran-sodium sulphate-induced colitis in the mouse.

Bcl-3 is a member of the IκB family of proteins and is an essential negative regulator of Toll-like receptor-induced responses. Recently, a single nucleotide polymorphism associated with reduced Bcl-3 gene expression has been identified as a potential risk factor for Crohn's disease. Here we report that in contrast to the predictions of single nucleotide polymorphism (SNP) analysis, patients with Crohn's disease and ulcerative colitis demonstrate elevated Bcl-3 mRNA expression relative to healthy individuals. To explore further the potential role of Bcl-3 in inflammatory bowel disease (IBD), we used the dextran-sodium sulphate (DSS)-induced model of colitis in Bcl-3(-/-) mice. We found that Bcl-3(-/-) mice were less sensitive to DSS-induced colitis compared to wild-type controls and demonstrated no significant weight loss following treatment. Histological analysis revealed similar levels of oedema and leucocyte infiltration between DSS-treated wild-type and Bcl-3(-/-) mice, but showed that Bcl-3(-/-) mice retained colonic tissue architecture which was absent in wild-type mice following DSS treatment. Analysis of the expression of the proinflammatory cytokines interleukin (IL)-1ß, tumour necrosis factor (TNF)-α and IL-6 revealed no significant differences between DSS-treated Bcl-3(-/-) and wild-type mice. Analysis of intestinal epithelial cell proliferation revealed enhanced proliferation in Bcl-3(-/-) mice, which correlated with preserved tissue architecture. Our results reveal that Bcl-3 has an important role in regulating intestinal epithelial cell proliferation and sensitivity to DSS-induced colitis which is distinct from its role as a negative regulator of inflammation.

Bifidobacterium infantis suppression of Peyer's patch MIP-1α and MIP-1ß secretion during Salmonella infection correlates with increased local CD4+CD25+ T cell numbers.

The outcome following infection depends on the generation of an immune response that results in control of the pathogenic microorganism, while limiting inflammatory collateral damage to the host. Bifidobacterium infantis 35624 was shown to be protective against Salmonella associated host injury via a Treg-dependent mechanism. In this study, we further examined the mechanisms by which B. infantis-induced Tregs protect against Salmonella-associated inflammation. B. infantis 35624 feeding to Salmonella-infected mice significantly reduced Peyer's patch MIP-1α and MIP-1ß secretion. Chemokine secretion was significantly inversely correlated with Peyer's patch CD4+CD25+ cell numbers. In vitro, CD25+ T cells, but not CD25- T cells, specifically inhibited TNF-α and IFN-γ secretion. However, both CD25+ and CD25- T cells suppressed MIP-1α and MIP-1ß secretion to the same extent. This study suggests that although B. infantis 35624-induced Tregs correlate with inhibition of chemokine secretion within the mucosa of pathogen infected animals, indirect cellular mechanisms may play a role.

Plain abdominal radiographs in patients with Crohn's disease: radiological findings and diagnostic value.

AIM: To determine the diagnostic yield and clinical value of plain film of the abdomen (PFA) in Crohn's disease (CD) patients and to determine whether performance of PFA yields definitive diagnostic information or whether additional imaging examinations are required. MATERIALS AND METHODS: One hundred and seventy-seven CD patients underwent 643 PFAs during the period September 1992 to August 2008. Two radiologists blinded to the clinical details independently evaluated individual PFAs and/or their reports for abnormal findings using the following criteria: normal, small bowel (SB) findings; colonic findings, acute CD complications, extra-colonic findings; global assessment/impression. The results of additional imaging studies performed within 5 days of PFA were recorded and findings were analysed. RESULTS: A mean of 3.6 (range 1-22) PFAs was performed per patient during the study period. Almost 70% of films were normal (n = 449). SB abnormalities were detected in 21.8% (n = 140) PFAs; most commonly dilated loops (18.8%, n = 121) and mucosal oedema (5%, n = 32). Colonic abnormalities were present in 11.4% (n = 73); most commonly mucosal oedema (7.5%, n = 48) and dilated loops (5%, n = 32). Four cases of pneumoperitoneum were detected. There was no case of toxic megacolon. There was one case in which intra-abdominal abscess/collection was suspected and two cases of obstruction/ileus. Extracolonic findings (renal calculi, sacro-iliitis, etc.) were identified in 7.5% (n = 48). PFAs were followed by additional abdominal imaging within 5 days of PFA in 273/643 (42.5%) of cases. CONCLUSION: Despite the high rates of utilization of PFA in CD patients, there is a low incidence of abnormal findings (32.5%). Many of the findings are non-specific and clinically irrelevant and PFA is frequently followed by additional abdominal imaging examinations.

Safety of Bifidobacterium breve, Bif195, employing a human exercise-induced intestinal permeability model: a randomised, double-blinded, placebo-controlled, parallel group trial.

We have previously shown that the probiotic Bifidobacterium breve strain Bif195 alleviates mucosal injury including ulcer formation in the upper intestine induced by non-steroid anti-inflammatory drugs (NSAIDs). Here, we report additional safety use of Bif195 in 126 healthy humans undergoing an exercise-induced intestinal permeability challenge in a double-blinded, placebo-controlled randomised 6-week intervention trial. Intestinal permeability was assessed by urinary lactulose/rhamnose (L/R) ratio. L/R ratio, plasma intestinal fatty acid binding protein (I-FABP) and gastrointestinal symptom rating scale (GSRS) questionnaire were measured resting and after a 1 h treadmill challenge, prior to and at the end of the intervention. To be able to compare the equivalence of resting state at baseline, of this cohort of well-trained subjects, to non-trained subjects, a cohort of 63 healthy and non-trained subjects (<2 h/week of endurance sports) was included. Study subjects (well-trained) were 35.7% women with a mean age and body mass index (in kg/m2) of 35.0 years and 24.8, respectively. There were no differences between the Bif195 and placebo groups in effects on L/R ratio, I-FABP and GSRS questionnaire score. In addition, there were no differences between Bif195 and placebo in number of adverse events and change in cytokines, liver or kidney biomarkers. The exercise model successfully induced intestinal permeability by statistically significantly increasing L/R ratio by ~100% (P<0.0001) and cytokines after the exercise challenge. No significant difference was found between well-trained and non-trained subjects in baseline resting L/R ratio. In conclusion, the reported cytoprotective effects of Bif195 are unlikely to be primarily related to small bowel permeability, and the safety of Bif195 in individuals with increased permeability is supported by the present data. ClinicalTrials.gov: NCT03027583.

Metagenomic assembled plasmids of the human microbiome vary across disease cohorts.

We compiled a human metagenome assembled plasmid (MAP) database and interrogated differences across multiple studies that were originally designed to investigate the composition of the human microbiome across various lifestyles, life stages and events. This was performed as plasmids enable bacteria to rapidly expand their functional capacity through mobilisation, yet their contribution to human health and disease is poorly understood. We observed that inter-sample ß-diversity differences of plasmid content (plasmidome) could distinguish cohorts across a multitude of conditions. We also show that reduced intra-sample plasmidome α-diversity is consistent amongst patients with inflammatory bowel disease (IBD) and Clostridioides difficile infections. We also show that faecal microbiota transplants can restore plasmidome diversity. Overall plasmidome diversity, specific plasmids, and plasmid-encoded functions can all potentially act as biomarkers of IBD or its severity. The human plasmidome is an overlooked facet of the microbiome and should be integrated into investigations regarding the role of the microbiome in promoting health or disease. Including MAP databases in analyses will enable a greater understanding of the roles of plasmid-encoded functions within the gut microbiome and will inform future human metagenome analyses.

Modulation, microbiota and inflammation in the adult CF gut: A prospective study.

BACKGROUND: Cystic Fibrosis (CF) has prominent gastrointestinal and pancreatic manifestations. The aim of this study was to determine the effect of Cystic fibrosis transmembrane conductance regulator (CFTR) modulation on, gastrointestinal inflammation, pancreatic function and gut microbiota composition in people with cystic fibrosis (CF) and the G551D-CFTR mutation. METHODS: Fourteen adult patients with the G551D-CFTR mutation were assessed clinically at baseline and for up to 1 year after treatment with ivacaftor. The change in gut inflammatory markers (calprotectin and lactoferrin), exocrine pancreatic status and gut microbiota composition and structure were assessed in stool samples. RESULTS: There was no significant change in faecal calprotectin nor lactoferrin in patients with treatment while all patients remained severely pancreatic insufficient. There was no significant change in gut microbiota diversity and richness following treatment. CONCLUSION: There was no significant change in gut inflammation after partial restoration of CFTR function with ivacaftor, suggesting that excess gut inflammation in CF is multi-factorial in aetiology. In this adult cohort, exocrine pancreatic function was irreversibly lost. Longer term follow-up may reveal more dynamic changes in the gut microbiota and possible restoration of CFTR function.

Immune privilege or inflammation? Insights into the Fas ligand enigma.

Fas ligand (FasL) has become an enigmatic molecule: some evidence indicates that it contributes to immune privilege in tissues and tumors, whereas other data demonstrates that FasL can elicit inflammation. New findings may begin to reconcile the paradoxical effects of FasL.

Composition and energy harvesting capacity of the gut microbiota: relationship to diet, obesity and time in mouse models.

BACKGROUND AND AIMS: Increased efficiency of energy harvest, due to alterations in the gut microbiota (increased Firmicutes and decreased Bacteroidetes), has been implicated in obesity in mice and humans. However, a causal relationship is unproven and contributory variables include diet, genetics and age. Therefore, we explored the effect of a high-fat (HF) diet and genetically determined obesity (ob/ob) for changes in microbiota and energy harvesting capacity over time. METHODS: Seven-week-old male ob/ob mice were fed a low-fat diet and wild-type mice were fed either a low-fat diet or a HF-diet for 8 weeks (n=8/group). They were assessed at 7, 11 and 15 weeks of age for: fat and lean body mass (by NMR); faecal and caecal short-chain fatty acids (SCFA, by gas chromatography); faecal energy content (by bomb calorimetry) and microbial composition (by metagenomic pyrosequencing). RESULTS: A progressive increase in Firmicutes was confirmed in both HF-fed and ob/ob mice reaching statistical significance in the former, but this phylum was unchanged over time in the lean controls. Reductions in Bacteroidetes were also found in ob/ob mice. However, changes in the microbiota were dissociated from markers of energy harvest. Thus, although the faecal energy in the ob/ob mice was significantly decreased at 7 weeks, and caecal SCFA increased, these did not persist and faecal acetate diminished over time in both ob/ob and HF-fed mice, but not in lean controls. Furthermore, the proportion of the major phyla did not correlate with energy harvest markers. CONCLUSION: The relationship between the microbial composition and energy harvesting capacity is more complex than previously considered. While compositional changes in the faecal microbiota were confirmed, this was primarily a feature of high-fat feeding rather than genetically induced obesity. In addition, changes in the proportions of the major phyla were unrelated to markers of energy harvest which changed over time. The possibility of microbial adaptation to diet and time should be considered in future studies.

The fecal mycobiome in patients with Irritable Bowel Syndrome.

Alterations of the gut microbiota have been reported in various gastrointestinal disorders, but knowledge of the mycobiome is limited. We investigated the gut mycobiome of 80 patients with Irritable Bowel Syndrome (IBS) in comparison with 64 control subjects. The fungal-specific internal transcribed spacer 1 (ITS-1) amplicon was sequenced, and mycobiome zero-radius operational taxonomic units (zOTUs) were defined representing known and unknown species and strains. The fungal community was sparse and individual-specific in all (both IBS and control) subjects. Although beta-diversity differed significantly between IBS and controls, no difference was found among clinical subtypes of IBS or in comparison with the mycobiome of subjects with bile acid malabsorption (BAM), a condition which may overlap with IBS with diarrhoea. The mycobiome alterations co-varied significantly with the bacteriome and metabolome but were not linked with dietary habits. As a putative biomarker of IBS, the predictive power of the fecal mycobiome in machine learning models was significantly better than random but insufficient for clinical diagnosis. The mycobiome presents limited therapeutic and diagnostic potential for IBS, despite co-variation with bacterial components which do offer such potential.

Mapping the colorectal tumor microbiota.

The gut microbiome in patients with colorectal cancer (CRC) is different than that of healthy controls. Previous studies have profiled the CRC tumor microbiome using a single biopsy. However, since the morphology and cellular subtype vary significantly within an individual tumor, the possibility of sampling error arises for the microbiome within an individual tumor. To test this hypothesis, seven biopsies were taken from representative areas on and off the tumor in five patients with CRC. The microbiome composition was strikingly similar across all samples from an individual. The variation in microbiome alpha-diversity was significantly greater between individuals' samples then within individuals. This is the first study, to our knowledge, that shows that the microbiome of an individual tumor is spatially homogeneous. Our finding strengthens the assumption that a single biopsy is representative of the entire tumor, and that microbiota changes are not limited to a specific area of the neoplasm.

The gut virome in Irritable Bowel Syndrome differs from that of controls.

Irritable Bowel Syndrome (IBS), the most common gastrointestinal disorder, is diagnosed solely on symptoms. Potentially diagnostic alterations in the bacterial component of the gut microbiome (the bacteriome) are associated with IBS, but despite the known role of the virome (particularly bacteriophages), in shaping the gut bacteriome, few studies have investigated the virome in IBS. We performed metagenomic sequencing of fecal Virus-Like Particles (VLPs) from 55 patients with IBS and 51 control individuals. We detected significantly lower alpha diversity of viral clusters comprising both known and novel viruses (viral 'dark matter') in IBS and a significant difference in beta diversity compared to controls, but not between IBS symptom subtypes. The three most abundant bacteriophage clusters belonged to the Siphoviridae, Myoviridae, and Podoviridae families (Order Caudovirales). A core virome (defined as a cluster present in at least 50% of samples) of 5 and 12 viral clusters was identified in IBS and control subjects, respectively. We also identified a subset of viral clusters that showed differential abundance between IBS and controls. The virome did not co-vary significantly with the bacteriome, with IBS clinical subtype, or with Bile Acid Malabsorption status. However, differences in the virome could be related back to the bacteriome as analysis of CRISPR spacers indicated that the virome alterations were at least partially related to the alterations in the bacteriome. We found no evidence for a shift from lytic to lysogenic replication of core viral clusters, a phenomenon reported for the gut virome of patients with Inflammatory Bowel Disease. Collectively, our data show alterations in the virome of patients with IBS, regardless of clinical subtype, which may facilitate development of new microbiome-based therapeutics.