P462. Terminal restriction fragment length polymorphism analysis of the diversity of fecal microbiota in patients with inflammatory bowel disease


C.H. Choi1, B.K. Cha2, K. Kim3

1Chung-Ang University College of Medicine, Seoul, South Korea; 2Chung-Ang University College of Medicine, Internal Medicine, Seoul, South Korea; 3Chung-Ang University College of Medicine, Department of Microbiology, Seoul, South Korea



Background: To perform terminal restriction fragment length polymorphism (T-RFLP) analyses of fecal microbiota in inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS) patients and to investigate the potential alterations in fecal bacterial communities.

Methods: A total of 104 patients with ulcerative colitis (UC, n = 54), Crohn's disease (CD, n = 24) and IBS (n = 26) and 37 healthy subjects (HS) were enrolled. DNA was extracted from their stool samples, and the 16S rRNA genes were amplified by PCR. The PCR products were then digested with MspI and HinP1I restriction enzymes, and the length of the T‑RF was determined. The sizes of T‑RFs were rounded to the nearest number between samples to produce operational taxonomic units (OTUs). A one-way analysis of similarity (ANOSIM) was used to compare the microbial communities. Analysis of similarity percentages (SIMPER) was done to determine the overall average dissimilarity and the OTUs significantly contributing the dissimilarity in microbial community compositions among the groups. The bacterial species of the significantly different OTUs were predicted from the database we developed (http://microbiology.cau.ac.kr) based on the silico PCR amplification and restriction of 16S rRNA sequences.

Results: The composition of the fecal bacterial community was significantly different between the patient groups and the HS (P < 0.05). It was also significantly different among all disease groups except between UC and IBS. Dissimilarities between the patient groups and the HS were as following: UC, 62.7%; CD, 68.4%; IBS, 61.5%. In UC patients, streptococci, Megasphaera and Bacteroides species were significantly abundant, and Bacillus, Megasphaera elsdenii, Selenomonas ruminantium, Ruminococcus, Clostridium, Selenomonas, Enterobacter species were deficient compared to the HS. In CD patients, Megasphaera, Streptomyces and Sphingomonas were significantly abundant, and Bacillus, Megasphaera elsdenii, Selenomonas ruminantium, Lactobacillus, Ruminococcus, Clostridium, Bacteroides and Sphingobacterium were deficient. In IBS patients, Actinobacillus species were abundant and other bacteria were also detected abundantly similar to those of UC patients. Bacillus, Clostridium, Megasphaera elsdenii and Selenomonas ruminantium species were commonly deficient in all groups of patients.

Conclusions: The composition of fecal microbiota in patients with UC, CD, or IBS significantly differs from that of HS, and also different from each other patient group except between UC and IBS.