Microbiota composition and mucosal immunity in patients with asymptomatic diverticulosis and controls.

INTRODUCTION: The etiology of diverticulosis is still poorly understood. However, in patients with diverticulitis, markers of mucosal inflammation and microbiota alterations have been found. The aim of this study was to evaluate potential differences of the gut microbiota composition and mucosal immunity between patients with asymptomatic diverticulosis and controls. METHODS: We performed a prospective study on patients who underwent routine colonoscopy for causes not related to diverticular disease or inflammatory bowel disease. Participants were grouped based on the presence or absence of diverticula. Mucosal biopsies were obtained from the sigmoid and transverse colon. Microbiota composition was analyzed with IS-pro, a 16S-23S based bacterial profiling technique. To predict if patients belonged to the asymptomatic diverticulosis or control group a partial least squares discriminant analysis (PLS-DA) regression model was used. Inflammation was assessed by neutrophil and lymphocyte counts within the taken biopsies. RESULTS: Forty-three patients were enrolled. Intestinal microbiota profiles were highly similar within individuals for all phyla. Between individuals, microbiota profiles differed substantially but regardless of the presence (n = 19) of absence (n = 24) of diverticula. Microbiota diversity in both sigmoid and transverse colon was similar in all participants. We were not able to differentiate between diverticulosis patients and controls with a PLS-DA model. Mucosal lymphocyte counts were comparable among both groups; no neutrophils were detected in any of the studied biopsies. CONCLUSIONS: Microbiota composition and inflammatory markers were comparable among asymptomatic diverticulosis patients and controls. This suggests that the gut microbiota and mucosal inflammation do not play a major role in the pathogenesis of diverticula formation.

Intestinal Barrier Breakdown and Mucosal Microbiota Disturbance in Neuromyelitis Optical Spectrum Disorders.

Background and Purpose: The mechanism underlying the pathology of neuromyelitis optica spectrum disorders (NMOSD) remains unclear even though antibodies to the water channel protein aquaporin-4 (AQP4) on astrocytes play important roles. Our previous study showed that dysbiosis occurred in the fecal microbiota of NMOSD patients. In this study, we further investigated whether the intestinal barrier and mucosal flora balance are also interrupted in NMOSD patients. Methods: Sigmoid mucosal biopsies were collected by endoscopy from six patients with NMOSD and compared with samples from five healthy control (HC) individuals. These samples were processed for electron microscopy and immunohistochemistry to investigate changes in ultrastructure and in the number and size of intestinal inflammatory cells. Changes in mucosal flora were also analyzed by high-throughput 16S ribosomal RNA gene amplicon sequencing. Results: The results from bacterial rRNA gene sequencing showed that bacterial diversity was decreased, but Streptococcus and Granulicatella were abundant in the colonic mucosa specimens of NMOSD patients compared to the HC individuals. The intercellular space between epithelia of the colonic mucosa was wider in NMOSD patients compared to the HC subjects (p < 0.01), and the expression of tight junction proteins [occludin, claudin-1 and zonula occludens-1 (ZO-1)] in NMOSD patients significantly decreased compared to that in the HC subjects. We also found numerous activated macrophages with many inclusions within the cytoplasm, mast cells with many particles in their cytoplasm, and enlarged plasma cells with rich developed rough endoplasmic reticulum in the lamina propria of the mucosa of the patients with NMOSD. Quantitative analysis showed that the percentages of small CD38+ and CD138+ cells (plasma cells) were lower, but the percentage of larger plasma cells was higher in NMOSD patients. Conclusion: The present study demonstrated that the intestinal barrier was disrupted in the patients with NMOSD, accompanied by dysbiosis and inflammatory activation of the gut. The mucosal microbiota imbalance and inflammatory responses might allow pathogens to cross the damaged intestinal barrier and participate in pathological process in NMOSD. However, further study on the pathological mechanism of NMOSD underlying gut dysbiosis is warranted in the future.

Sialylation and fucosylation modulate inflammasome-activating eIF2 Signaling and microbial translocation during HIV infection.

An emerging paradigm suggests that gut glycosylation is a key force in maintaining the homeostatic relationship between the gut and its microbiota. Nevertheless, it is unclear how gut glycosylation contributes to the HIV-associated microbial translocation and inflammation that persist despite viral suppression and contribute to the development of several comorbidities. We examined terminal ileum, right colon, and sigmoid colon biopsies from HIV-infected virally-suppressed individuals and found that gut glycomic patterns are associated with distinct microbial compositions and differential levels of chronic inflammation and HIV persistence. In particular, high levels of the pro-inflammatory hypo-sialylated T-antigen glycans and low levels of the anti-inflammatory fucosylated glycans were associated with higher abundance of glycan-degrading microbial species (in particular, Bacteroides vulgatus), a less diverse microbiome, higher levels of inflammation, and higher levels of ileum-associated HIV DNA. These findings are linked to the activation of the inflammasome-mediating eIF2 signaling pathway. Our study thus provides the first proof-of-concept evidence that a previously unappreciated factor, gut glycosylation, is a force that may impact the vicious cycle between HIV infection, microbial translocation, and chronic inflammation.

An Aberrant Microbiota is not Strongly Associated with Incidental Colonic Diverticulosis.

Colonic diverticula are protrusions of the mucosa through weak areas of the colonic musculature. The etiology of diverticulosis is poorly understood, but could be related to gut bacteria. Using mucosal biopsies from the sigmoid colon of 226 subjects with and 309 subjects without diverticula during first-time screening colonoscopy, we assessed whether individuals with incidental colonic diverticulosis have alternations in the adherent bacterial communities in the sigmoid colon. We found little evidence of substantial associations between the microbial community and diverticulosis among cases and controls. Comparisons of bacterial abundances across all taxonomic levels showed differences for phylum Proteobacteria (p = 0.038) and family Comamonadaceae (p = 0.035). The r-squared values measuring the strength of these associations were very weak, however, with values ~2%. There was a similarly small association between the abundance of each taxa and total diverticula counts. Cases with proximal only diverticula and distal only diverticula likewise showed little difference in overall microbiota profiles. This large study suggests little association between diverticula and the mucosal microbiota overall, or by diverticula number and location. We conclude that the mucosal adherent microbiota community composition is unlikely to play a substantial role in development of diverticulosis.

DNA Methylation and Transcription Patterns in Intestinal Epithelial Cells From Pediatric Patients With Inflammatory Bowel Diseases Differentiate Disease Subtypes and Associate With Outcome.

BACKGROUND & AIMS: We analyzed DNA methylation patterns and transcriptomes of primary intestinal epithelial cells (IEC) of children newly diagnosed with inflammatory bowel diseases (IBD) to learn more about pathogenesis. METHODS: We obtained mucosal biopsies (N = 236) collected from terminal ileum and ascending and sigmoid colons of children (median age 13 years) newly diagnosed with IBD (43 with Crohn's disease [CD], 23 with ulcerative colitis [UC]), and 30 children without IBD (controls). Patients were recruited and managed at a hospital in the United Kingdom from 2013 through 2016. We also obtained biopsies collected at later stages from a subset of patients. IECs were purified and analyzed for genome-wide DNA methylation patterns and gene expression profiles. Adjacent microbiota were isolated from biopsies and analyzed by 16S gene sequencing. We generated intestinal organoid cultures from a subset of samples and genome-wide DNA methylation analysis was performed. RESULTS: We found gut segment-specific differences in DNA methylation and transcription profiles of IECs from children with IBD vs controls; some were independent of mucosal inflammation. Changes in gut microbiota between IBD and control groups were not as large and were difficult to assess because of large amounts of intra-individual variation. Only IECs from patients with CD had changes in DNA methylation and transcription patterns in terminal ileum epithelium, compared with controls. Colon epithelium from patients with CD and from patients with ulcerative colitis had distinct changes in DNA methylation and transcription patterns, compared with controls. In IECs from patients with IBD, changes in DNA methylation, compared with controls, were stable over time and were partially retained in ex-vivo organoid cultures. Statistical analyses of epithelial cell profiles allowed us to distinguish children with CD or UC from controls; profiles correlated with disease outcome parameters, such as the requirement for treatment with biologic agents. CONCLUSIONS: We identified specific changes in DNA methylation and transcriptome patterns in IECs from pediatric patients with IBD compared with controls. These data indicate that IECs undergo changes during IBD development and could be involved in pathogenesis. Further analyses of primary IECs from patients with IBD could improve our understanding of the large variations in disease progression and outcomes.

The Microbial Ecosystem Distinguishes Chronically Diseased Tissue from Adjacent Tissue in the Sigmoid Colon of Chronic, Recurrent Diverticulitis Patients.

Diverticular disease is commonly associated with the older population in the United States. As individual's age, diverticulae, or herniation of the mucosa through the colonic wall, develop. In 10-25% of individuals, the diverticulae become inflamed, resulting in diverticulitis. The gut ecosystem relies on the interaction of bacteria and fungi to maintain homeostasis. Although bacterial dysbiosis has been implicated in the pathogenesis of diverticulitis, associations between the microbial ecosystem and diverticulitis remain largely unstudied. This study investigated how the cooperative network of bacteria and fungi differ between a diseased area of the sigmoid colon chronically affected by diverticulitis and adjacent non-affected tissue. To identify mucosa-associated microbes, bacterial 16S rRNA and fungal ITS sequencing were performed on chronically diseased sigmoid colon tissue (DT) and adjacent tissue (AT) from the same colonic segment. We found that Pseudomonas and Basidiomycota OTUs were associated with AT while Microbacteriaceae and Ascomycota were enriched in DT. Bipartite co-occurrence networks were constructed for each tissue type. The DT and AT networks were distinct for each tissue type, with no microbial relationships maintained after intersection merge of the groups. Our findings indicate that the microbial ecosystem distinguishes chronically diseased tissue from adjacent tissue.

The Potential Role of Gut-Derived Inflammation in Multiple System Atrophy.

BACKGROUND: Recent evidence suggests that Parkinson's disease (PD) is associated with intestinal microbiota dysbiosis, abnormal intestinal permeability, and intestinal inflammation. OBJECTIVE: Our study aimed to determine if these gut abnormalities are present in another synucleinopathy, multiple system atrophy (MSA). METHODS: In six MSA and 11 healthy control subjects, we performed immunohistochemistry studies of colonic sigmoid mucosa to evaluate the intestinal barrier marker Zonula Occludens-1 and the endotoxin-related inflammation marker Toll-like-receptor-4 expression. We also assessed colonic sigmoid mucosal and fecal microbiota compositions using high-throughput 16S ribosomal RNA gene amplicon sequencing. RESULTS: MSA subjects showed disrupted tight junction protein Zonula Occludens-1 structure in sigmoid mucosa tissue suggesting intestinal barrier dysfunction. The lipopolysaccharide specific inflammatory receptor Toll-like-receptor-4 was significantly higher in the colonic sigmoid mucosa in MSA relative to healthy controls. Microbiota analysis suggested high relative abundance of gram-negative, putative "pro-inflammatory" bacteria in various family and genus level taxa, from the phylum Bacteroidetes and Proteobacteria, in MSA feces and mucosa. At the taxonomic level of genus, putative "anti-inflammatory" butyrate-producing bacteria were less abundant in MSA feces. Predictive functional analysis indicated that the relative abundance of a number of genes involved in metabolism were lower in MSA feces, whereas the relative abundance of genes involved in lipopolysaccharide biosynthesis were higher in both MSA feces and mucosa compared to healthy controls. CONCLUSIONS: This proof-of-concept study provides preliminary evidence that like PD, MSA subjects display evidence of disrupted intestinal barrier integrity, increased marker of endotoxin-related intestinal inflammation, and pro-inflammatory colonic microbiota.

Colonic bacterial composition in Parkinson's disease.

INTRODUCTION: We showed that Parkinson's disease (PD) patients have alpha-synuclein (α-Syn) aggregation in their colon with evidence of colonic inflammation. If PD patients have altered colonic microbiota, dysbiosis might be the mechanism of neuroinflammation that leads to α-Syn misfolding and PD pathology. METHODS: Sixty-six sigmoid mucosal biopsies and 65 fecal samples were collected from 38 PD patients and 34 healthy controls. Mucosal-associated and feces microbiota compositions were characterized using high-throughput ribosomal RNA gene amplicon sequencing. Data were correlated with clinical measures of PD, and a predictive assessment of microbial community functional potential was used to identify microbial functions. RESULTS: The mucosal and fecal microbial community of PD patients was significantly different than control subjects, with the fecal samples showing more marked differences than the sigmoid mucosa. At the taxonomic level of genus, putative, "anti-inflammatory" butyrate-producing bacteria from the genera Blautia, Coprococcus, and Roseburia were significantly more abundant in feces of controls than PD patients. Bacteria from the genus Faecalibacterium were significantly more abundant in the mucosa of controls than PD. Putative, "proinflammatory" Proteobacteria of the genus Ralstonia were significantly more abundant in mucosa of PD than controls. Predictive metagenomics indicated that a large number of genes involved in metabolism were significantly lower in the PD fecal microbiome, whereas genes involved in lipopolysaccharide biosynthesis and type III bacterial secretion systems were significantly higher in PD patients. CONCLUSION: This report provides evidence that proinflammatory dysbiosis is present in PD patients and could trigger inflammation-induced misfolding of α-Syn and development of PD pathology.

Clostridium difficile infection after ileostomy closure mimicking anastomotic leak.

Clostridium difficile infection is linked to antibiotic exposure, with elderly and immunocompromised hospitalised patients being particularly at risk. The symptoms range from mild diarrhoea to life-threatening fulminant colitis. We describe an unusual presentation of C. difficile infection after closure of ileostomy in a healthy 60-year-old man with a history of low anterior resection and defunctioning ileostomy for rectal tumour. On the third day postoperatively, the patient developed left lower abdominal pain and profuse diarrhoea. With worsening symptoms and steadily increasing inflammatory markers over the following few days, concerns were raised about an anastomotic leak with pelvic abscess. CT of the abdomen/pelvis on day 7 surprisingly showed colitis in the neorectum/sigmoid colon. A stool test confirmed C. difficile infection.

Phylum level change in the cecal and fecal gut communities of rats fed diets containing different fermentable substrates supports a role for nitrogen as a factor contributing to community structure.

Fermentation differs between the proximal and distal gut but little is known regarding how the bacterial communities differ or how they are influenced by diet. In order to investigate this, we compared community diversity in the cecum and feces of rats by 16S rRNA gene content and DNA shot gun metagenomics after feeding purified diets containing different fermentable substrates. Gut community composition was dependent on the source of fermentable substrate included in the diet. Cecal communities were dominated by Firmicutes, and contained a higher abundance of Lachnospiraceae compared to feces. In feces, community structure was shifted by varying degrees depending on diet towards the Bacteroidetes, although this change was not always evident from 16S rRNA gene data. Multi-dimensional scaling analysis (PCoA) comparing cecal and fecal metagenomes grouped by location within the gut rather than by diet, suggesting that factors in addition to substrate were important for community change in the distal gut. Differentially abundant genes in each environment supported this shift away from the Firmicutes in the cecum (e.g., motility) towards the Bacteroidetes in feces (e.g., Bacteroidales transposons). We suggest that this phylum level change reflects a shift to ammonia as the primary source of nitrogen used to support continued microbial growth in the distal gut.

Gonorrhoea of the sigmoid neovagina in a male-to-female transgender.

A 33-year-old male-to-female transgender consulted our outpatient clinic with perneovaginal bleeding during and following coitus. Four years before, she underwent a total laparoscopic sigmoid neovaginoplasty. Physical, histological and endoscopic examination revealed neither focus of active bleeding nor signs of active inflammation. A polymerase chain reaction test performed on a neovaginal swab showed gonococcal infection. Treatment consisted of 500 mg intramuscular ceftriaxone. Three weeks later, our patient reported resolution of symptoms, consistent with eradication of the infection demonstrated by a follow-up neovaginal swab polymerase chain reaction. To our knowledge, this is the first case report of gonococcal infection of the sigmoid neovagina.

Postoperative bacteriuria, pyuria and urinary tract infection in patients with an orthotopic sigmoid colon neobladder replacement.

The purpose of this study is to investigate the prevalence of postoperative bacteriuria, pyuria and urine culture in patients with an orthotopic sigmoid colon neobladder replacement. Urine samples for bacteriuria, pyuria and urine culture, if necessary, were collected at 1, 3, 6, 9 and 12 months after surgery and the presence of blood culture and antibiotic-resistant strains, and their treatments on positive urine culture cases were investigated. Of 209 for bacteriuria and 207 for pyuria urine samples with evaluable data, 95 (45.5%) were positive for bacteriuria and 76 (36.7%) had pyuria (10 or more white blood cells per high-power field). Totally, 30 bacteria were isolated from urine culture of urinary tract infection (UTI) and Klebisiella pneumoniae, Escherichia coli, Staphylococcos aureus and Enterococcus spp. strains were representatively isolated. The incidence of pyuria significantly decreased over time (P=0.041) but that of bacteriuria did not (P=0.107). In them, there were six bacteria (20.7%) with antibiotic-resistant strains. The antibiotics used for their treatments representatively were levofloxacin in five cases, tazobactam/piperacillin in three cases and sulfamethoxazole/trimethoprim and cefepime, meropenem in two cases, respectively. In conclusion, these findings suggest that physicians taking care of sigmoid colon neobladder patients need to be aware of these high ratios of bacteriuria, pyuria and UTI, including bacteremia.

Host-microbe relationships in inflammatory bowel disease detected by bacterial and metaproteomic analysis of the mucosal-luminal interface.

BACKGROUND: Host-microbe interactions at the intestinal mucosal-luminal interface (MLI) are critical factors in the biology of inflammatory bowel disease (IBD). METHODS: To address this issue, we performed a series of investigations integrating analysis of the bacteria and metaproteome at the MLI of Crohn's disease, ulcerative colitis, and healthy human subjects. After quantifying these variables in mucosal specimens from a first sample set, we searched for bacteria exhibiting strong correlations with host proteins. This assessment identified a small subset of bacterial phylotypes possessing this host interaction property. Using a second and independent sample set, we tested the association of disease state with levels of these 14 "host interaction" bacterial phylotypes. RESULTS: A high frequency of these bacteria (35%) significantly differentiated human subjects by disease type. Analysis of the MLI metaproteomes also yielded disease classification with exceptional confidence levels. Examination of the relationships between the bacteria and proteins, using regularized canonical correlation analysis (RCCA), sorted most subjects by disease type, supporting the concept that host-microbe interactions are involved in the biology underlying IBD. Moreover, this correlation analysis identified bacteria and proteins that were undetected by standard means-based methods such as analysis of variance, and identified associations of specific bacterial phylotypes with particular protein features of the innate immune response, some of which have been documented in model systems. CONCLUSIONS: These findings suggest that computational mining of mucosa-associated bacteria for host interaction provides an unsupervised strategy to uncover networks of bacterial taxa and host processes relevant to normal and disease states. (Inflamm Bowel Dis 2012;).

MRSA peritonitis secondary to perforation of sigmoid diverticulitis.

The occurrence of methicillin-resistant Staphylococcus aureus (MRSA) is well documented, but the pathology is usually associated with post surgical infections or long-term peritoneal dialysis. We report the case of a 50-year-old Caucasian man who presented with a one week history of left lower quadrant abdominal pain, poor appetite and nausea due to MRSA peritonitis secondary to perforated sigmoid diverticulitis. Despite a thorough search of the medical literature, we could not find that this problem has been previously described. We report this case to demonstrate the robust nature of MRSA, which has generally not been considered to be a normal colonizing bacterium of the sigmoid colon.

A case of perforative peritonitis caused by a piece of bamboo in a patient on peritoneal dialysis.

We report a case of peritonitis resulting from colon perforation caused by ingestion of a rare foreign body in a patient on peritoneal dialysis (PD). A 72-year-old woman on PD was hospitalized with abdominal pain and cloudy PD fluid (PDF). Although conventional antibiotic therapy was started because of a diagnosis of infectious peritonitis, low-grade fever, abdominal pain and a high number of white blood cells in PDF persisted. On day 3, anaerobic bacteria were recognized on bacterial culture of PDF, suggesting a gastrointestinal etiology. During exploratory laparotomy, sigmoidal perforation by a piece of bamboo, probably resulting from ingestion of contaminated food, was found.

Twin study indicates loss of interaction between microbiota and mucosa of patients with ulcerative colitis.

BACKGROUND & AIMS: Interactions between genetic and environmental factors are believed to be involved in onset and initiation of inflammatory bowel disease. We analyzed the interaction between gastrointestinal mucosal microbiota and host genes in twin pairs discordant for ulcerative colitis (UC) to study the functional interaction between microbiota and mucosal epithelium. METHODS: Biopsy were collected from sigmoid colon of UC patients and their healthy twins (discordant twin pairs) and from twins without UC. Microbiota profiles were determined from analysis of 16S ribosomal DNA libraries; messenger RNA profiles were determined by microarray analysis. RESULTS: Patients with UC had dysbiotic microbiota, characterized by less bacterial diversity and more Actinobacteria and Proteobacteria than that of their healthy siblings; healthy siblings from discordant twins had more bacteria from the Lachnospiraceae and Ruminococcaceae families than twins who were both healthy. In twins who were both healthy, 34 mucosal transcripts correlated with bacterial genera, whereas only 25 and 11 correlated with bacteria genera in healthy individuals and their twins with UC, respectively. Transcripts related to oxidative and immune responses were differentially expressed between patients with UC and their healthy twins. CONCLUSIONS: The transcriptional profile of the mucosa appears to interact with the colonic microbiota; this interaction appears to be lost in colon of patients with UC. Bacterial functions, such as butyrate production, might affect mucosal gene expression. Patients with UC had different gene expression profiles and lower levels of biodiversity than their healthy twins, as well as unusual aerobic bacteria. Patients with UC had lower percentages of potentially protective bacterial species than their healthy twins.