Diversity of Microflora in Colonic Mucus from Severe Ulcerative Colitis Patients Analyzed by Terminal Restriction Fragment Length Polymorphism and Clone Libraries of Bacterial 16S rRNA Gene Sequences

I-Nung Huang,Takeshi Naitoh,Junko Nishimura,Akira Horii,Chikashi Shibata,Tadao Saito,Moriya Ohkuma,Haruki Kitazawa,Mitsuo Sakamoto,Yuri Sato,Shinobu Ohnuma

doi:10.4236/aim.2014.413095

Abstract

Although the gut microflora is thought to be an essential factor in the development of ulcerative colitis (UC), the entire gut microflora occurring in UC remains unknown. Most studies use feces to represent the microflora distribution; however, here we analyzed the bacterial diversity in colonic mucus from UC patients receiving colectomy surgery and control patients. The diversity of microflora was investigated using a combination of terminal restriction fragment length polymorphism (T-RFLP) and clone library analyses of the 16S rRNA gene sequences. In the T-RFLP analysis, the number of terminal restriction fragments (T-RFs) decreased significantly in UC patients when compared to control samples. Also in the clone library analysis, the number of operational taxonomic units (OTU) and the Shannon diversity index were reduced significantly in UC patients. These molecular analyses reveal an overall dysbiosis in UC patients. No specific pathogen was found, and a strong negative correlation in relative abundance of bacterial populations was observed between the phyla Bacteroidetes and Firmicutes in the UC patients. This is the first report showing a significant correlation between these two phyla, which may be important characteristics in the pathogenesis of UC.

Highlights

Ulcerative colitis (UC) is an intractable disease where erosion and ulcers occur in the colon with chronic inflammation
Computer-Simulated terminal restriction fragment length polymorphism (T-RFLP) Analysis Based on the results from the 16S rRNA gene clone library analysis, we enabled to match the bacterial genus and/or species to terminal restriction fragments (T-RFs) in the T-RFLP patterns of the four UC samples and six controls
It is evident that there is a strong relationship between UC and gut microflora, the entire picture remains unclear

Summary

Introduction

Ulcerative colitis (UC) is an intractable disease where erosion and ulcers occur in the colon with chronic inflammation. The etiology of UC is associated with genetic and environmental factors causing immunological disorders and chronic inflammation; the mechanism remains unclear. Genetic factors, such as ECM1 [1] and HLA-DR [2] were reported to be strongly associated with an excessive immune response in UC. The symptoms disappeared when the mice were bred in sterile conditions. This supports the hypothesis that the presence of microflora is necessary for development of UC symptoms

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