Adhesion of Escherichia coli under flow conditions reveals potential novel effects of FimH mutations.
Adhesion of Escherichia coli under flow conditions reveals potential novel effects of FimH mutations.
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DOI：
10.1007/s10096-016-2820-8
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发表时间：
2017-03
期刊：
影响因子：
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通讯作者：
Kain R
中科院分区：
文献类型：
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作者：
Feenstra T;Thøgersen MS;Wieser E;Peschel A;Ball MJ;Brandes R;Satchell SC;Stockner T;Aarestrup FM;Rees AJ;Kain R
fimh介导的大肠杆菌对膀胱上皮的粘附是尿路感染的先决条件。FimH对血源性细菌传播也是必不可少的，但其机制尚不清楚。本研究的目的是利用一种新型的粘附试验来评估不同的FimH突变对细菌粘附的影响，该试验模拟了细菌所暴露的生理流动条件。我们在仅表达1型菌毛的大肠杆菌菌株（MSC95-FimH）中引入了12个不同的点突变。我们通过几种常用的粘附试验比较了每个突变体的细菌粘附，包括酵母的凝集，对单甘露糖基化和三甘露糖基化底物的粘附，以及对膀胱上皮细胞和内皮细胞的静态粘附。我们将这些实验与我们开发的一种新方法进行了比较，该方法用于研究细菌在流动条件下对哺乳动物细胞的粘附。我们发现大肠杆菌MSC95-FimH在微血管内皮上的粘附比在膀胱上皮上的粘附更有效，并且只有内皮在生理剪切应力下支持粘附。结果证实甘露糖结合口袋突变消除了粘附。我们证明了FimH残基E50和T53对流动条件下的粘附至关重要。内皮细胞在生物芯片上的涂层和生理流动条件的建模使我们能够识别对粘附至关重要的FimH残基。这些结果为确定FimH突变体和潜在的FimH拮抗剂的作用的筛选方法提供了新的见解。本文的在线版本（doi:10.1007/s10096-016-2820-8）包含补充资料，仅供授权用户使用。
FimH-mediated adhesion of Escherichia coli to bladder epithelium is a prerequisite for urinary tract infections. FimH is also essential for blood-borne bacterial dissemination, but the mechanisms are poorly understood. The purpose of this study was to assess the influence of different FimH mutations on bacterial adhesion using a novel adhesion assay, which models the physiological flow conditions bacteria are exposed to. We introduced 12 different point mutations in the mannose binding pocket of FimH in an E. coli strain expressing type 1 fimbriae only (MSC95-FimH). We compared the bacterial adhesion of each mutant across several commonly used adhesion assays, including agglutination of yeast, adhesion to mono- and tri-mannosylated substrates, and static adhesion to bladder epithelial and endothelial cells. We performed a comparison of these assays to a novel method that we developed to study bacterial adhesion to mammalian cells under flow conditions. We showed that E. coli MSC95-FimH adheres more efficiently to microvascular endothelium than to bladder epithelium, and that only endothelium supports adhesion at physiological shear stress. The results confirmed that mannose binding pocket mutations abrogated adhesion. We demonstrated that FimH residues E50 and T53 are crucial for adhesion under flow conditions. The coating of endothelial cells on biochips and modelling of physiological flow conditions enabled us to identify FimH residues crucial for adhesion. These results provide novel insights into screening methods to determine the effect of FimH mutants and potentially FimH antagonists. The online version of this article (doi:10.1007/s10096-016-2820-8) contains supplementary material, which is available to authorized users.