Background & Aims: Pouch inflammation after surgery for ulcerative colitis can significantly alter quality of life and thus ideally should be prevented. Dysbiosis or altered microflora is suspected to be the key pathogenic factor for pouchitis. However, dysbiosis in pouchitis has not been characterized carefully because of a lack of available sensitive microbiological technology suitable for in vivo studies in human beings. Thus, the aims of our study were as follows: (1) to show the use of the length heterogeneity polymerase chain reaction (LH-PCR) technique for studying microflora in human beings, and (2) to use the technique to characterize the microfloral patterns in the ileal pouch of patients with pouchitis. Methods: Microfloral patterns initially were assessed using a 16S ribosomal RNA technique (LH-PCR) to determine the qualitative changes in the luminal and mucosal intestinal flora. We subsequently cloned and sequenced the LH-PCR amplification products from the community 16S ribosomal RNA found in patients with pouchitis and in control pouch to identify the microbial species involved in pouchitis. Results: We have shown unique microfloral patterns in pouchitis. Through cloning and sequencing of the LH-PCR amplicons, we have shown the persistence of Fusobacter and Enteric species associated with the disease state and the absence of specific bacteria such as Streptococcus species in the inflamed pouch. Conclusions: We have shown that the LH-PCR technique is suitable for studying microflora in human beings. By using this technique and the clone sequences, we have shown dysbiosis in the microbial biofilm adherent to the mucosa in pouchitis. Our data provide direct evidence of the role of bacteria in the pathogenesis of pouchitis.
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