Synchrotron imaging of crystalline biofilms in urinary catheters

描述（由申请方提供）：一套基于同步加速器的X射线成像方法将用于评估导尿管中奇异变形杆菌形成的结晶生物膜的发展。变形杆菌是尿路感染中极其常见的病原体，并且在经受长期导尿的患者中特别成问题。奇异变形杆菌水解尿素，导致尿液pH值升高，并在尿路内形成矿物质沉积物。这些沉积物在膀胱和肾脏中形成结石和结壳，以及在插入的装置（例如导管和输尿管支架）上形成结壳。大约有一半的长期导尿管患者由于这些感染而发生导管堵塞。堵塞事件可引起严重的急性症状，是导管插入患者发病率和死亡率的一个促成因素。监测和更换导尿管的需求也给这些患者的长期管理带来了巨大的负担。了解结晶生物膜的发展和相关导管堵塞的一个主要挑战是，矿物沉积物阻止使用光学方法，如共聚焦显微镜。为了克服这一限制，尿导管内的变形杆菌生物膜将使用我们最近开发的用于矿物样品非侵入性研究的基于同步加速器的复杂方法进行成像。同步加速器X射线计算机微断层扫描将用于以近微米级分辨率对变形杆菌生物膜的三维孔隙结构进行成像，同步加速器X射线衍射也将用于评估生物膜内不同矿物相的分布。此外，将使用环境扫描电子显微镜评估生物膜内的细胞分布，并使用能量色散光谱法和电子衍射进行分析电子显微镜检查，以确定结晶沉积物内的化学分布。将对使用模型膀胱系统在体外生长的含有结晶生物膜的完整导管以及从患者体内取出的堵塞导管进行分析。生物膜孔隙结构的定量描述将开发使用图像分析方法，并在每个生物膜内的内部运输环境将通过使用断层扫描数据结合晶格玻尔兹曼模拟孔隙流体流动进行评估。将比较不同变形杆菌菌株和不同生长持续时间获得的结果，以评估孔结构、内部运输条件和结晶生物膜发育的相互作用，以及生物膜生长对导管堵塞的总体影响。公共卫生相关性：变形杆菌感染引起的导管堵塞仍然是患者和长期护理机构的主要、持续和严重问题。对结晶生物膜的结构和组成缺乏了解阻碍了预防导管感染和控制导管堵塞的新治疗方法的开发。该项目的结果将表明晶体结构如何影响尿导管上变形杆菌生物膜的发展，并提出治疗这些感染和预防导管堵塞的改进策略。

DESCRIPTION (provided by applicant): A suite of synchrotron-based x-ray imaging methodologies will be used to assess the development of crystalline biofilms formed by Proteus mirabilis in urinary catheters. Proteus is an extremely common pathogen in urinary tract infections, and is particularly problematic in patients subject to long-term urinary catheterization. P. mirabilis hydrolyzes urea, leading to an increase in the pH of urine and the formation of mineral deposits within the urinary tract. These deposits form stones and encrustations in the bladder and kidneys, as well as encrustations on inserted devices such as catheters and ureteral stents. Approximately half of patients with long-term urinary catheters develop catheter blockage as a result of these infections. Blockage incidents can cause serious acute symptoms and are a contributing factor to morbidity and mortality in catheterized patients. The need to monitor and replace urinary catheters also presents a substantial burden for long-term management of these patients. One major challenge in understanding the development of crystalline biofilms and the associated catheter blockage is that the mineral deposits prevent use of optical methods such as confocal microscopy. To overcome this limitation, Proteus biofilms within urinary catheters will be imaged using sophisticated synchrotron-based methods that we have recently developed for non-invasive investigations of mineral samples. Synchrotron x-ray computed micro-tomography will be used to image the pore structure of Proteus biofilms in three dimensions with near-micron-scale resolution, and synchrotron x- ray diffraction will also be used to assess the distribution of different mineral phases within the biofilms. In addition, environmental scanning electron microscopy will be used to assess cell distributions within biofilms, and analytical electron microscopy will be performed using energy-dispersive spectrometry and electron diffraction to determine chemical distributions within the crystalline deposits. Analyses will be performed on intact catheters containing crystalline biofilms grown in vitro using a model bladder system, as well as on blocked catheters retrieved from patients. Quantitative descriptions of the biofilm pore structure will be developed using image analysis methods, and the internal transport environment within each biofilm will be evaluated by using the tomographic data in conjunction with lattice Boltzmann simulations of pore fluid flow. Results obtained for different Proteus strains and different growth durations will be compared in order to assess the interplay of pore structure, internal transport conditions, and crystalline biofilm development, as well as the overall effects of biofilm growth on catheter blockage. PUBLIC HEALTH RELEVANCE: Catheter blockage from Proteus infections still represents a major, ongoing, and serious problem for patients and for long-term care facilities. A lack of understanding of the structure and composition of crystalline biofilms has hindered the development of new treatments to prevent catheter infections and control catheter blockage. The project results will indicate how the crystalline structure influences the development of Proteus biofilms on urinary catheters and suggest improved strategies for treating these infections and for preventing catheter blockage.