Alterations of the dominant faecal bacterial groups in patients with Crohn's disease of the colon - PubMed (original) (raw)
Alterations of the dominant faecal bacterial groups in patients with Crohn's disease of the colon
P Seksik et al. Gut. 2003 Feb.
Abstract
Background and aims: The colonic microflora is involved in the pathogenesis of Crohn's disease (CD) but less than 30% of the microflora can be cultured. We investigated potential differences in the faecal microflora between patients with colonic CD in remission (n=9), patients with active colonic CD (n=8), and healthy volunteers (n=16) using culture independent techniques.
Methods: Quantitative dot blot hybridisation with six radiolabelled 16S ribosomal ribonucleic acid (rRNA) targeting oligonucleotide probes was used to measure the proportions of rRNA corresponding to each phylogenetic group. Temporal temperature gradient gel electrophoresis (TTGE) of 16S rDNA was used to evaluate dominant species diversity.
Results: Enterobacteria were significantly increased in active and quiescent CD. Probe additivity was significantly lower in patients (65 (11)% and 69 (6)% in active CD and quiescent CD) than in healthy controls (99 (7)%). TTGE profiles varied markedly between active and quiescent CD but were stable in healthy conditions.
Conclusion: The biodiversity of the microflora remains high in patients with CD. Enterobacteria were observed significantly more frequently in CD than in health, and more than 30% of the dominant flora belonged to yet undefined phylogenetic groups.
Figures
Figure 1
Composition of the dominant faecal flora, as assessed by dot blot hybridisation with specific probes in healthy volunteers, patients with inactive Crohn's disease, and independent patients with active Crohn's disease. Ribosomal ribonucleic acid (rRNA) indexes correspond to specific 16S rRNA as a proportion of total bacterial 16S rRNA (means (SEM) of triplicate measurements). Comparison versus healthy subjects : *p=0.0001, **p=0.006, ***p=0.010, ****p=0.013, *****p=0.034.
Figure 2
Temporal temperature gradient gel electrophoresis of 16S rDNA amplicons (obtained using primers for the V6-V8 region of the gene) of samples obtained from independent patients during active and inactive colonic Crohn's disease. M, Marker, composed of a polymerase chain reaction amplification mix of cloned rDNA from Clostridium coccoides (Nos 157, 93, and 40), Clostridium leptum (Nos 365 and 296), and Bacteroides (Nos 303 and 73)23. I, inactive disease; A, active disease.
Figure 3
Temporal temperature gradient gel electrophoresis of 16S rDNA amplicons of the dominant flora in a healthy control subject sampled over a two year period. S1, sample collected in 1997; S2–S5, samples collected in 1999 on days 1, 23, 58, and 78, respectively. The dendrogram gives a statistically optimal representation of similarities between temporal temperature gradient gel electrophoresis profiles based on the matrix of Pearson correlation coefficients and applying the Unweighted Pair Group Method using Arithmetic averages (UPGMA).
Figure 4
Temporal temperature gradient gel electrophoresis of 16S rDNA amplicons of paired faecal samples from four patients who were studied during both active and inactive Crohn's disease. The dendrogram was obtained as indicated in fig 3 ▶. IX, sample from patient X during inactive disease; AX, sample from patient X during active disease.
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