GROWTH AND COLONIZATION OF THE INTESTINE BY E COLI

细菌要在动物宿主体内定植，必须获得生长所需的营养。然而，尽管进行了广泛的研究，但人们对生活在哺乳动物大肠中的400-500种动物的营养物质和代谢途径几乎一无所知。本研究利用功能基因组学技术研究肠道细菌在定植过程中对营养物质的获取。具体来说，我们将验证一个假设，即诱导黏液衍生糖的分解代谢途径对大肠杆菌的定植至关重要。全球转录测定将用于确定肠道粘液中存在的复杂营养混合物诱导生长的调控。从含有盲肠粘液的最小培养基上培养的细胞制备的探针将与所有大肠杆菌基因的DNA阵列杂交。通过分析实验定植无菌小鼠盲肠中大肠杆菌细胞的全局转录模式，将确定在粘液上原位诱导生长的途径。大肠杆菌成功与大肠内其他400-500种细菌竞争的途径将被研究。在特定的分解代谢途径中被选择性阻断的突变菌株将被测试其定植小鼠的能力，绿色荧光蛋白报告融合将被用来确定这些调控在定植期间在单个细菌细胞中的时间和空间表达。这些实验将确定单个粘膜糖在大肠杆菌定殖中的独特生理作用，为解决代谢多样性是否允许致病性大肠杆菌菌株在预定殖菌株存在的情况下定殖从而导致其致病能力的更普遍问题提供重要的知识。确切地知道哪些营养物质对定植是必不可少的，应该有助于改进对抗感染的策略。

To colonize an animal host, bacteria must acquire nutrients for growth. However, despite extensive research, next to nothing is known about the nutrients and metabolic pathways used in situ by the 400-500 species which inhabit the mammalian large intestine. The proposed research uses functional genomics technology to investigate the acquisition of nutrients by enteric bacteria during the colonization process. Specifically, we will test the hypothesis that induction of the pathways used for catabolism of mucus-derived sugars is essential for colonization of E. coli. Global transcription assays will be used to identify regulons induced for growth on the complex mixture of nutrients present in intestinal mucus. Probes prepared from cells grown on minimal media containing cecal mucus will be hybridized to DNA arrays of all E. coli genes. Pathways induced in situ for growth on mucus will be identified by analyzing global transcription patterns of E. coli cells in the ceca of experimentally colonized germ-free mice. Pathways which contribute to the ability of E. coli to successfully compete with the 400-500 other species in the large intestine will be examined. Mutant strains selectively blocked in specific catabolic pathways will be tested for their ability to colonize mice and green fluorescent protein reporter fusions will be used to determine the temporal and spatial expression of these regulons in individual bacterial cells during colonization. These experiments will identify the distinct physiological roles of individual mucosal sugars in colonization of the large intestine by E. coli, providing knowledge important for addressing the more general question of whether or not metabolic diversity allows pathogenic E. coli strains to colonize in the presence of precolonized strains, thus leading to their ability to cause disease. Knowing precisely which nutrients are essential for colonization should lead to improved strategies for combating infection.