Microfabricated 3D Environments for Characterizing Bacterial Group Behaviors

描述（由申请人提供）：这项工作的目标是建立一个平台，用于研究受控环境条件下的细菌群体行为，从而研究与毒力和增强的抗生素和免疫抗性相对应的状态的发生和维持有关的基本问题。在这些研究中，一种新的方法，基于多光子光刻（MPL）将开发用于微制造定义，任意地形使用生物积木（蛋白质），这将被调查作为一种手段，以产生能够支持细胞聚集，生长和群体行为的微腔，集中在群体感应和生物膜沉积的机会环境细菌，铜绿假单胞菌，作为一个模型系统。这种细菌与许多人类疾病有关，包括囊性纤维化个体的持续感染。为此，拟议的研究将推进基础MPL技术，重点是材料和微腔特性的表征，并将这种最先进的技术应用于评估铜绿假单胞菌在蛋白质微腔中的行为。该项目代表了Jason Shear教授的研究小组（化学/生物化学系）与UT Austin的Marvin Whiteley教授（微生物学系）之间的合作研究。预计所产生的数据将为未来的研究提供重要信息，旨在确定针对细菌通讯系统的新疗法。虽然我们建议将铜绿假单胞菌作为这些研究的测试细菌，但它们可以很容易地扩展到其他细菌物种，甚至更复杂的多物种群落。具体目标是：（1）优化用于制造具有可调质量传输到外部环境的3D细菌微腔的柔性平台;和（2）评估基于3D蛋白质的微腔用于研究细菌群体行为的效用。公共卫生相关性：拟议研究的目标是开发一个平台，用于研究受控环境条件下的细菌群体行为，从而研究与毒力和增强的抗生素和免疫抗性相对应的状态的发生和维持有关的基本问题。在这些研究中，一种新的方法，用于微制造定义，任意地形使用生物积木将被调查作为一种手段，以产生能够支持细胞聚集，生长和群体行为的微腔。预计所产生的数据将为旨在鉴定靶向细菌通讯系统的新型疗法的研究提供重要信息。

DESCRIPTION (provided by applicant): The goal of this work is to produce a platform for studying bacterial group behavior under controlled environmental conditions, allowing fundamental questions to be investigated regarding the onset and maintenance of states corresponding to virulence and enhanced antibiotic and immune resistance. In these studies, a novel approach based on multiphoton lithography (MPL) will be developed for microfabricating defined, arbitrary topographies using biological building blocks (proteins) which will be investigated as a means to produce microcavities capable of supporting cell aggregration, growth, and group behavior, focusing on quorum sensing and biofilm deposition by the opportunistic environmental bacterium, Pseudomonas aeruginosa, as a model system. This bacterium has relevance to a number of human diseases, including persistant infections in cystic fibrosis individuals. Toward these ends, the proposed studies will advance the underlying MPL technology, focusing on characterization of material and microcavity properties, and will apply this state-of-the-art technology to an evaluation of P. aeruginosa behavior in protein-based microcavities. This project represents collaborative studies between Prof. Jason Shear's research group (Chemistry/Biochemistry Department), and that of Prof. Marvin Whiteley (Microbiology Department) at UT Austin. It is anticipated that the data generated will provide important information for future studies aimed at identifying novel therapeutics targeting bacterial communication systems. Although we are proposing P. aeruginosa as the test bacterium for these studies, they could easily be extended to other bacterial species and even to more complex multi-species communities. The Specific Aims are: (1) To Optimize a flexible platform for fabricating 3D bacterial microcavities having tunable mass-transport to the outside environment; and (2) To assess the utility of 3D protein-based microcavities for studying bacterial group behavior. PUBLIC HEALTH RELEVANCE: The goal of the proposed research is to develop a platform for studying bacterial group behavior under controlled environmental conditions, allowing fundamental questions to be investigated regarding the onset and maintenance of states corresponding to virulence and enhanced antibiotic and immune resistance. In these studies, a novel approach for microfabricating defined, arbitrary topographies using biological building blocks will be investigated as a means to produce microcavities capable of supporting cell aggregration, growth, and group behavior. It is anticipated that the data generated will provide important information for studies aimed at identifying novel therapeutics targeting bacterial communication systems.