儿童近端小肠粘膜相关菌群分布研究

目的研究儿童近端小肠粘膜相关菌群种类和分布。方法取儿童近端小肠粘膜活检组织,提取总DNA,经PCR扩增后对16SrRNA基因扩增片段克隆,并对克隆片段进行DNA序列测定,根据序列结果分析细菌组成。同时在不同培养条件下进行细菌培养及鉴定。结果16SrRNA基因测序结果表明儿童近端小肠粘膜相关菌群包括6个细菌门类,厚壁菌门、变形菌门和放线菌门占总细菌的88.8%;拟杆菌门、梭杆菌门和TM7占11.2%。链球菌属和奈瑟菌属为儿童近端小肠粘膜相关菌群的主要菌属;韦荣菌属、孪生菌属、放线菌属、罗氏球菌属、嗜血菌属、普雷沃菌属和颗粒链菌属占较高比例。细菌培养结果与16SrRNA基因测序结果类似。结论儿童近端小肠粘膜具有独特的菌群定殖分布。

Study on the characterization of the mucosa-associated bacteria in proximal small intestine in children

Objective To characterize the microbiota of proximal small intestine in children and identify the mucosa-associated bacteria. Methods Collected the proximal small intestine biopsies from 15 children with clinical controls and extracted the whole DNA. Bacteria were identified by 16S rDNA sequencing in DNA extracted from biopsies washed with PBS buffer containing dithiothreitol to enrich for bacteria adhering to the epithelial lining. Three consecutive biopsies were investigated using selective conditions for anaerobic and aerobic growth. Bacteria were subjected to quantitative cultivation and characterized by standard bacteriological methods. Results The normal mucosa-associated microbiota of proximal small intestine in children is constructed by six different phyla, including Firmicutes, Proteobacteria and Actinohacteria dominated the bacterial community which accounted for the 88. 8%. Bacteriodetes, Fusobacteria and TM7 constituted small populations which accounted for the 11.2%. The most diverse phyla were Firmicutes and Proteobacteria. The Genus Level Operational Taxonomic Units （GELOTUs） were identified. It was dominated by the genera Streptococcus and Neisseria also containing VeiUonella, Gemella, Actinomyces, Rothia, Haemophilus, Prevotella and Granulicatella. Live bacteria were identified in all three biopsies by culture methods. Streptococcus, Neisseria, Haemophilus, Veillonella, Fusobacterium, Rothia and Actinomyces species were found in most, or all biopsies, confirming the results from 16S rDNA sequencing. Cenclusion The composition of the bacterial biota in children proximal small intestine is different from those of the other sites indicating that proximal small intestine is a distinct ecological niche.