Search for Localized Dysbiosis in Crohn's Disease Ulcerations by Temporal Temperature Gradient Gel Electrophoresis of 16S rRNA

Seksik, Philippe; Lepage, Patricia; Cochetière, Marie-France de la; Bourreille, Arnaud; Sutren, Malène; Galmiche, Jean–Paul; Doré, Joël; Marteau, Philippe

doi:10.1128/jcm.43.9.4654-4658.2005

Cited by 74 publications

(50 citation statements)

References 32 publications

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“…This is in agreement with the recent findings of Eckburg and colleagues (3) but is somewhat surprising since several of our analyses compared inflamed and healthy tissues. A previous study of CD patients by use of temporal temperature gradient gel electrophoresis has also shown that ulcerated and nonulcerated tissues are similar in dominant-species composition (20). These findings suggest that it is unlikely that inflammation is directly caused by a mucosa-associated pathogen, a conclusion also reached by another group using a different methodology (22).…”

Section: Resultssupporting

confidence: 69%

Differences between Tissue-Associated Intestinal Microfloras of Patients with Crohn's Disease and Ulcerative Colitis

et al. 2006

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A leading hypothesis for the role of bacteria in inflammatory bowel diseases is that an imbalance in normal gut flora is a prerequisite for inflammation. Testing this hypothesis requires comparisons between the microbiota compositions of ulcerative colitis and Crohn's disease patients and those of healthy individuals. In this study, we obtained biopsy samples from patients with Crohn's disease and ulcerative colitis and from healthy controls. Bacterial DNA was extracted from the tissue samples, amplified using universal bacterial 16S rRNA gene primers, and cloned into a plasmid vector. Insert-containing colonies were picked for highthroughput sequencing, and sequence data were analyzed, yielding species-level phylogenetic data. The clone libraries yielded 3,305 sequenced clones, representing 151 operational taxonomical units. There was no significant difference between floras from inflamed and healthy tissues from within the same individual. Proteobacteria were significantly (P ‫؍‬ 0.0007) increased in Crohn's disease patients, as were Bacteroidetes (P < 0.0001), while Clostridia were decreased in that group (P < 0.0001) in comparison with the healthy and ulcerative colitis groups, which displayed no significant differences. Thus, the bacterial flora composition of Crohn's patients appears to be significantly altered from that of healthy controls, unlike that of ulcerative colitis patients. Imbalance in flora in Crohn's disease is probably not sufficient to cause inflammation, since microbiotas from inflamed and noninflamed tissues were of similar compositions within the same individual.

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Section: Resultssupporting

confidence: 69%

Differences between Tissue-Associated Intestinal Microfloras of Patients with Crohn's Disease and Ulcerative Colitis

et al. 2006

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“…Some authors reported increased numbers of aerobic Gram-negative rods (E. coli), while others found increased numbers of non-spore forming anaerobes (Proteobacteria and Bacteroides) and decreased numbers of spore-forming anaerobes like Clostridium [20][21][22][23][24] . There are also papers in which the authors found no differences between the bacterial flora in inflamed and non-inflamed tissue in the same patient [25,26] . On the contrary, Sepehri et al [27] using a PCR method showed significant differences in bacterial flora between the inflamed and non-inflamed mucosa, and an increase in microbial diversity in controls and the non-inflamed tissue from adult IBD patients.…”

Section: Discussionmentioning

confidence: 99%

Mucosal bacterial microﬂora and mucus layer thickness inadolescents with inﬂammatory bowel disease

Fyderek¹,

Strus²,

Kowalska-Duplaga³

et al. 2009

WJG

125

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AIM:To assess the mucosa-associated bacterial microflora and mucus layer in adolescents with inflammatory bowel disease (IBD). METHODS:Sixty-one adolescents (mean age 15 years, SD ± 4.13) were included in the study. Intestinal biopsies from inflamed and non-inflamed mucosa of IBD patients and from controls with functional abdominal pain were cultured under aerobic and anaerobic conditions. The number of microbes belonging to the same group was calculated per weight of collected tissue. The mucus thickness in frozen samples was measured under a fluorescent microscope. RESULTS:The ratios of different bacterial groups in inflamed and non-inflamed mucosa of IBD patients and controls were specific for particular diseases.Streptococcus spp . were predominant in the inflamed mucosa of Crohn's disease (CD) patients (80% of all bacteria), and Lactobacillus spp . were predominant in ulcerative colitis patients (90%). The differences were statistically significant (P = 0.01-0.001). Lower number of bifidobacteria was observed in the whole IBD group. A relation was also found between clinical and endoscopic severity and decreased numbers of Lactobacillus and, to a lesser extent, of Streptococcus in biopsies from CD patients. The mucus layer in the inflamed sites was significantly thinner as compared to controls (P = 0.0033) and to non-inflamed areas in IBD patients (P = 0.031). CONCLUSION:The significantly thinner mucosa of IBD patients showed a predominance of some aerobes specific for particular diseases, their numbers decreased in relation to higher clinical and endoscopic activity of the disease.

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“…TTGE is a culture-independent molecular method that has proven most appropriate in dynamic studies of dominant species diversity within complex ecosystems like the colon (8). The repeatability of amplifications with the universal primers used, and TTGE runs using the same starting sample, has already been shown (16,19). Furthermore, in all cases, a clear limit of detection was observed when the minority species accounted for 1:100 or less of the total DNA concentration (15,17).…”

Section: Discussionmentioning

confidence: 99%

Resilience of the Dominant Human Fecal Microbiota upon Short-Course Antibiotic Challenge

et al. 2005

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Recent studies have shown that the human fecal microbiota is composed of a consortium of species specific to the host and resistant to modifications over time. Antibiotics are known to affect the intestinal microflora, and ensuing changes may result in antibiotic-associated diarrhea. It is therefore important to characterize the nature and amplitude of these modifications and the ability of this ecosystem to return to its original profile-i.e., its resilience. Six healthy volunteers received oral amoxicillin (1.5 g/day) for 5 days. Fecal samples were collected at day 0 (D0) before antibiotic treatment and at set intervals until 60 days thereafter. Fecal DNA was isolated, and V6-to-V8 regions of the 16S rRNA genes were amplified by PCR with general primers and analyzed by temporal temperature gradient gel electrophoresis. Dominant species profiles were compared on the basis of similarity (Pearson correlation coefficient). Dominant species profiles at D0 were used as a reference. The fecal microbiota showed a major shift in dominant species upon antibiotic treatment, starting 24 h after treatment initiation and reaching an average similarity of only 74% after 4 days. Within 30 days following antibiotic treatment, the fecal microbiota tended to reach an average similarity of 88% to the D0 value; within 60 days, the average similarity to the D0 value was 89%. However, in one subject, important modifications persisted for at least 2 months, with similarity to the D0 value remaining below 70%. We demonstrated the resilience of the dominant human fecal microbiota upon short-course antibiotic challenge. Yet the persistence of long-term alterations in some subjects may explain susceptibilities to antibioticassociated diarrhea. Furthermore, these findings suggest that strategies reinforcing the ability of the fecal microbiota to resist modifications would be of clinical relevance.The human intestinal tract harbors a large, active, and complex community of microbes (22). The intestinal microbiota plays several significant roles in the digestion of food, metabolism of endogenous and exogenous compounds, immunopotentiation, and prevention of colonization by pathogens in the gastrointestinal tract and hence is involved in maintaining human health (12). Recent culture-independent molecular studies on healthy individuals have shown that the intestinal microbiota is specific to the host and resistant to modifications over time (26).Molecular analysis of the bacterial microbiota based on the 16S rRNA genes have attracted attention as reliable methods for detection and identification of bacterial species (9,13,14).

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Search for Localized Dysbiosis in Crohn's Disease Ulcerations by Temporal Temperature Gradient Gel Electrophoresis of 16S rRNA

Cited by 74 publications

References 32 publications

Differences between Tissue-Associated Intestinal Microfloras of Patients with Crohn's Disease and Ulcerative Colitis

Differences between Tissue-Associated Intestinal Microfloras of Patients with Crohn's Disease and Ulcerative Colitis

Mucosal bacterial microﬂora and mucus layer thickness inadolescents with inﬂammatory bowel disease

Resilience of the Dominant Human Fecal Microbiota upon Short-Course Antibiotic Challenge

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