Understanding How Bacteria Sense Mechanics Upon Attaching to Surfaces

生物膜是微生物群落，它们通过聚合物和蛋白质的基质相互结合并与表面结合。它们污染和腐蚀基础设施，并引起慢性感染。目前，制造防止感染的表面的方法只取得了有限的成功。该项目的目标是确定细菌在附着时如何感知表面力学，并通过开始形成生物膜来响应表面力学。这项工作将建立基础知识，可用于设计通过不给细菌开始形成生物膜所需的机械提示来抵抗生物膜发展的表面。因此，科学目标将改善生物膜的预防，从而有利于公共卫生和基础设施，如水处理、航运和石油运输。道德和心态方面的指导和教育将改善STEM专业的本科环境和管道。课堂教育方法的研究将改善本科STEM教育。该项目的研究目标是开发一个预测框架，以了解细菌如何利用细胞包膜中的蛋白质来感知和响应它们所附着的表面的力学。目前的假设是，细菌通过在它们的包膜中传导应力和变形来做到这一点，这取决于细菌和基质的弹性以及与表面粘附的能量。本研究将扩展铜绿假单胞菌的研究，并将其推广到另一种杆状革兰氏阴性菌（大肠杆菌）、一种杆状革兰氏阳性菌（枯草芽孢杆菌）和一种球形革兰氏阳性菌（金黄色葡萄球菌）。原子力显微镜（AFM）将用于测量细菌与具有不同表面化学和弹性的各种测试基质之间的附着力；AFM还将表征基底的弹性和地形。孔板中的高通量培养将用于测量底物上的细菌积累和生长，以及抗生素耐药性的增加，这是一种生物膜表型。有限元模型将描述当不同类型的细菌附着在不同的基质上时所产生的应力和变形。模型显示的趋势将通过测量机械敏感离子通道的活性来验证，当膜应力更大时，这些离子通道更开放。定量共聚焦显微镜和图像分析将测量由于细菌附着在表面而产生的细胞内信号和由于细菌在表面生长而产生的细胞间信号。基因操作（报告菌株和等基因敲除）将用于测量信号和阐明特定基因产物的作用，如候选机械感觉蛋白。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Biofilms are communities of microbes that are bound to each other and to a surface by a matrix of polymers and proteins. They foul and corrode infrastructure and cause chronic infections. At present, approaches to making surfaces that prevent infection have met with only limited success. The goal of this project is to determine how bacteria sense the mechanics of the surface upon attachment and respond to surface mechanics by starting to develop a biofilm. This work will establish basic knowledge that can be used to design surfaces that resist the development of biofilms by not giving bacteria the mechanical cue(s) needed to start forming a biofilm. Thus, the scientific aims will improve biofilm prevention and thereby benefit public health and infrastructure such as water treatment, shipping, and oil transport. Mentoring and education in ethics and mindset will improve the undergraduate environment and pipeline for STEM majors. A study of classroom education methods will improve undergraduate STEM education. The research objective of this project is to develop a predictive framework for understanding how bacteria use proteins in their cell envelopes to sense and respond to the mechanics of the surface to which they attach. The working hypothesis is that bacteria do this by transducing stresses and deformations in their envelopes that depend on the elasticity of both the bacterium and the substrate as well as the energy of adhesion with the surface. This study will extend studies of P. aeruginosa and generalize to include another rod-shaped Gram-negative bacterium (Escherichia coli), a rod-shaped Gram-positive bacterium (Bacillus subtilis), and a spherical Gram-positive bacterium (Staphylococcus aureus). Atomic force microscopy (AFM) will be used to measure the adhesion forces between bacteria and a wide range of test substrates with different surface chemistries and elasticities; AFM will also characterize the elasticity and topography of substrates. High-throughput culturing in well plates will be used to measure bacterial accumulation and growth on substrates, as well as the increase in antibiotic resistance that is a biofilm phenotype. Finite-element modeling will characterize the stresses and deformations that arise when different types of bacteria attach to different substrates. Trends shown by modeling will be validated by measuring the activity of mechanosensitive ion channels that are more open when the membrane stress is greater. Quantitative confocal microscopy and image analysis will measure the intracellular signaling that arises as a result of bacteria attaching to surfaces and the intercellular signaling that arises as a result of bacterial growth on the surface. Genetic manipulation (reporter strains and isogenic knockouts) will be used to measure signaling and to elucidate the role of specific gene products, such as candidate mechanosensory proteins.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.