Mucosa-associated bacteria in ulcerative colitis before and after new antibiotic combination therapy : DNA polymorphism analysis

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We examined the efficacy of three-antibiotics combination therapy against F. varium, and investigated the mucosa-associated bacteria before and after the therapy using a molecular approach. Twenty patients with ulcerative colitis were randomly assigned into the antibiotic treatment group (amoxicillin, tetracycline, and metronidazole for 2 weeks) of 12 patients and placebo group of 8 patients. Colonoscopic biopsy was performed at 0, 3 and 12 months after the treatment. DNA from mucosal bacteria was isolated from biopsy specimens. We investigated the mucosa-associated bacterial components by terminal restriction fragment length polymorphism (T-RFLP) with the restriction enzyme HhaI, and quantified the change in the number of Fusobacterium varium by real-time PCR. After the treatment, profile changes of T-RFLP were found after treatment in 10 of 12 patients in the antibiotic group and none of 8 patients in the placebo group (p=0.0007). Based on the real time PCR study, F. varium/actin ratio was reduced after treatment in 8 of 12 patients (67%) in the antibiotic group and 4 of 8patients (50%) in the placebo group. Mucosal RNA extracts were also analyzed by Nextera RNA sequencing system. The number of Bacteroides, Acidaminococcus, Prevotella, Parabacteroides were decreased in the antibiotic group. Some number of mucosa-associated bacteria significantly decreased after the antibiotics combination therapy.