Spatial Organization and Composition of the Mucosal Flora in Patients with Inflammatory Bowel Disease

Abstract

The composition and spatial organization of the mucosal flora in biopsy specimens from patients with inflammatory bowel disease (IBD; either Crohn's disease or ulcerative colitis), self-limiting colitis, irritablebowel syndrome (IBS), and healthy controls were investigated by using a broad range of fluorescent bacterial group-specific rRNA-targeted oligonucleotide probes. Each group included 20 subjects. Ten patients who had IBD and who were being treated with antibiotics were also studied. Use of nonaqueous Carnoy fixative to preserve the mucus layer was crucial for detection of bacteria adherent to the mucosal surface (mucosal bacteria). No biofilm was detectable in formalin-fixed biopsy specimens. Mucosal bacteria were found at concentrations greater than 10 9 /ml in 90 to 95% of IBD patients, 95% of patients with self-limiting colitis, 65% of IBS patients, and 35% of healthy controls. The mean density of the mucosal biofilm was 2 powers higher in IBD patients than in patients with IBS or controls, and bacteria were mostly adherent. Bacteroides fragilis was responsible for >60% of the biofilm mass in patients with IBD but for only 30% of the biofilm mass in patients with self-limiting colitis and <15% of the biofilm mass in patients with IBS. In contrast, bacteria which positively hybridized with the probe specific for Eubacterium rectale-Clostridium coccoides accounted for >40% of the biofilm in IBS patients but for <15% of the biofilm in IBD patients. In patients treated with (5-ASA) or antibiotics, the biofilm could be detected with 4,6-diamidino-2-phenylindole but did not hybridize with fluorescence in situ hybridization probes. A Bacteroides fragilis biofilm is the main feature of IBD. This was not previously recognized due to a lack of appropriate tissue fixation. Both 5-ASA and antibiotics suppress but do not eliminate the adherent biofilm.

Gram-Negative Bacteria Aggravate Murine Small Intestinal Th1-Type Immunopathology following Oral Infection with Toxoplasma gondii

“…These findings are well in line with the amelioration of ileitis developing in SAMP1/ YitFc mice by antibiotic treatment (29 -31, 60) and the abundance of E. coli and Bacteroides/Prevotella spp. detected in experimental colitis (61,62) and in patients with IBD (13). Furthermore, the colitogenic potential of both bacterial groups was determined earlier (18,19,21,63) and the contributions of Gram-negative bacteria to the severity of intestinal inflammation were supported by successful antibiotic treatment of experimental colitis and in human IBD (22,64,65), as well as in GvHD (8).…”

“…IBD patients display immunoreactivity against bacterial Ags (10 -12) and intestinal immunopathology is accompanied by accumulation of Escherichia coli or Bacteroides spp. at inflamed tissue sites (13)(14)(15)(16). In experimental colitis (17)(18)(19)(20)(21)(22)(23)(24), inflammation was suppressed in germfree animals or animals treated with antibiotics.…”

Mucosal biofilms: challenges and future directions

“…The presence of biofilms has been associated with the formation of inflammatory lesions in several mucosal locations in animals and humans [4,5,15], which suggests a direct role in pathogenesis. However, in the case of inflammatory bowel disease or chronic rhinosinusitis, where biofilms have been identified on the respective mucosal surfaces, the possibility exists that biofilm formation is secondary to an aberrant immune response to non-microbial antigens [16,17]. Thus, it is plausible that a chronic mucosal inflammatory response to selfantigens or allergens provides a tissue environment conducive to biofilm growth, since locally released inflammatory cytokines or excess mucus may promote biofilm growth [18].…”

“…Perhaps the biggest challenge in recognizing tissue microbial communities as biofilms is the demonstration of ECM presence, since it is easily destroyed by standard histologic processing. Thus, specialized fixation techniques or the use of fresh tissues for ECM analysis are frequently required [5,16]. Lectins, such as concanavalin A (with mannopyranosyl and glucopyranosyl specificity) or wheat germ agglutinin (with sialic acid and N-acetylglucosamine specificity), with affinities to carbohydrate targets that resemble those for antigen-antibody interactions, have been traditionally used to identify the ECM components in abiotic surface biofilms [23,24].…”

Microflora in Crohn’s disease: the emergence of adherent and invasiveEscherichia coli

“…CEACAM6 is expressed in the colonic mucosa of patients with CD and UC [8,42], and could be a receptor for colonic E. coli strains that display AIEC-like activity in cultured epithelial cells. However, this possibility has to be reconciled with the results of in situ analysis that show marked increases in mucosally adherent Bacteroides rather than E. coli in CD and UC colonic mucosa [44].…”