ORIGINAL ARTICLE Bacterial Community Diversity in Cultures Derived from Healthy and Inflamed Ileal Pouches After Restorative Proctocolectomy Matt W. Johnson, MRCP,* ,† Geraint B. Rogers, PhD, ‡ Kenneth D. Bruce, PhD, ‡ Andrew K. Lilley, PhD, ‡ Axel von Herbay, MD, § Alastair Forbes, FRCS, k Paul J. Ciclitira, FRCP, † and R. John Nicholls, FRCS* ,¶ Background: Pouchitis is believed to occur as a reaction to dys- biosis. In this study we assessed differences between mucosal bacterial communities cultured from noninflamed and inflamed ileal pouches. Methods: Thirty-two ileal pouch patients, 22 with ulcerative co- litis (UC) and 10 with familial adenomatous polyposis (FAP), underwent symptomatic, endoscopic, and histological assessment. The Objective Pouchitis Score (OPS) and the Pouch Disease Ac- tivity Index (PDAI) were used to diagnose pouchitis. Seven UC patients had pouchitis (UCþ), 15 had a noninflamed pouch (UC), 9 had a noninflamed pouch (FAP), and 1 FAP patient had pouchitis (FAPþ). Biopsies taken from the ileal mucosa of the pouch were cultured under aerobic and anaerobic conditions. Following standardized DNA extraction a polymerase chain reac- tion (PCR) was performed to generate 16S rRNA gene products. A ‘‘fingerprint’’ of the bacterial community within each sample was created using terminal-restriction fragment length polymor- phism (T-RFLP) profiling. Species richness and evenness were determined using T-RF band lengths and relative band intensities. Results: From the 64 DNA samples, 834 bands were detected, of which 179 represented different species (operational taxonomic units [OTUs]). The average species richness for the FAP, FAPþ, UC, and UCþ groups was 26, 35, 23.9, and 29.6 per patient, with the average species diversity within the groups of 10.6, 29, 8.3, and 11.4, respectively. Similar trends were observed when the anaerobic and aerobic-derived bacterial groups were an- alyzed separately. Conclusions: No significant differences were found between the bacterial cultures derived from any of the clinical groups or between pouchitis and nonpouchitis patients. (Inflamm Bowel Dis 2009;000:000–000) Key Words: bacteria, familial adenomatous polyposis, inflammatory bowel disease, pathogenesis, pouchitis, ulcerative colitis T he prevalence of ulcerative colitis (UC) has been esti- mated at over 30 cases per 10,000 in the UK. 1 Approx- imately 20% of these patients will at some point require colectomy for medically refractory disease or dysplasia. 2 Restorative procto-colectomy (RPC) with ileal pouch anal anastomosis has become the most common elective surgi- cal procedure for UC patients and in selected patients with familial adenomatous polyposis (FAP). The ileal pouch mucosa can become acutely inflamed. This condition, termed ‘‘pouchitis,’’ has a prevalence of 10% 3 and a cumu- lative incidence of 50% over a 5–10-year period. 4 Pou- chitis was thought to be rare in FAP patients but recent studies have suggested a prevalence of 5%. 5,6 After RPC the ileal pouch mucosa undergoes mor- phological changes characterized by villous shortening and increased leukocyte infiltration. This process, termed ‘‘colo- nization,’’ may be a reflection of ileal mucosal adaptation to the new fecal environment within the ileal reservoir. 7 There is a million-fold increase in the number of bacteria present and a transformation of the flora toward a spectrum more typical of the large bowel. 8,9 The cause of pouchitis is unknown but the effective- ness of antibiotic treatment to provide symptomatic, 10 en- doscopic, and histological 11–13 benefit suggests that bacteria are involved. Dysbiosis, defined as an abnormality of the host’s gastrointestinal microflora, is thought to play a key role in the pathogenesis of inflammatory bowel disease 14,15 and this partly explains the clinical improvements seen with antibiotic therapy in UC 16–21 and Crohn’s disease. 21–24 Received for publication April 2, 2009; Accepted May 11, 2009. From the *Gastroenterology Surgical Department, St. Mark’s Hospital, Harrow, Middlesex, UK, † Department of Gastroenterology, Rayne Institute, St Thomas’ Hospital, London, UK, ‡ King’s College London, Molecular Microbiology Research Laboratory, Pharmaceutical Sciences Division, Franklin-Wilkins Building, London, UK, § Academic Department of Pathology, St Mark’s Hospital, Harrow, Middlesex, UK, k Intestinal Failure Unit, University College Hospital, London, UK, ¶ Department of Biosurgery and Surgical Technology Imperial College, London, UK. Supported by St. Mark’s Hospital Foundation. Reprints: R. John Nicholls, FRCS, Gastroenterology Surgical Department, St. Mark’s Hospital, Northwick Park, Harrow, Middlesex, UK. HA13UJ (e-mail: j.nicholls@imperial.ac.uk) Copyright V C 2009 Crohn’s & Colitis Foundation of America, Inc. DOI 10.1002/ibd.21022 Published online in Wiley InterScience (www.interscience. wiley.com). Inflamm Bowel Dis 1