Metabolic heterogeneity and antibiotic susceptibility in biofilms

描述（由申请人提供）：生物膜是附着在表面上生长的微生物群落。基于生物膜的感染经常发生，生物膜在留置装置上生长很难根除。生物膜内低速率的抗生素运输、生物膜基质的保护作用以及生物膜内部低速率的代谢活性都被发现有助于这些感染的持续存在，但目前对这些影响的过程知之甚少。虽然生物膜的空间异质性对基于生物膜的感染的治疗选择显然很重要，但关于当地环境条件如何影响生物膜空间模式的发展，以及抗生素的有效性如何根据身体部位和留置装置的类型而变化的信息很少。我们假设生物膜内代谢活动的空间模式受到流动环境中的空间模式的影响，这些相互作用导致生物膜的复杂性随着时间的推移而增加。我们还假设流动环境不仅通过影响生物膜内抗菌素向细胞的传递，而且通过决定群落内的代谢梯度来影响生物膜的抗菌素敏感性。我们建议通过以下具体目标来解决这些假设。目的1：观察单物种生物膜在平面流动细胞中的生长，以评估生物膜形态、运输模式和代谢活性随着环境流动条件空间变异性的增加而发生的变化。目的2：观察抗生素治疗对不同程度空间复杂性生物膜的杀灭效果，并将局部杀灭效率的分布与运输条件和代谢活动的空间格局联系起来。目标3：发展一个改进的数值模型，以便对上述影响进行定量分析。目的4：使用该模型澄清多尺度流动-生物膜相互作用，特别是评估抗生素治疗下生物膜中细胞亚群存活的关键特征。我们建议采用新颖的实验和数值模拟相结合的方法来实现这些目标。我们将在一个新的实验系统中进行生物膜生长和处理的实验，该实验系统提供了在流入和流出模式中施加精确控制程度的空间变异性的能力。生物膜的生长、流量和氧气分布的变化、抗生素的运输以及由此产生的细胞死亡都将直接在原位观察。我们将利用这些新的和独特的观察结果来支持生物膜发展的新数值模型的发展，该模型随后将用于模拟抗生素治疗在不同局部生长条件下根除生物膜的有效性。这种测量和建模的结合将提供对生物膜生长与外部流动相互作用和改变的方式的独特见解，以及最终这种复杂的相互作用如何控制生物膜的整体形成以及引入的抗菌剂对居住在生物膜基质中的细胞的影响。

DESCRIPTION (provided by applicant): Biofilms are microbial communities that grow attached to a surface. Biofilm-based infections occur frequently, and biofilm growth on indwelling devices is very difficult to eradicate. Low rates of antibiotic transport within biofilms, protective effects of the biofilm matrix, and low rates of metabolic activity within the biofilm interior have all been found to contribute to the persistence of these infections, but there is currently little understanding of the processes responsible for these effects. While spatial heterogeneity in biofilms is clearly important to selection of therapy for biofilm-based infections, little information is available on the way in which local environmental conditions influence the development of spatial patterns in biofilms, and hence how the effectiveness of antibiotics varies depending on the body site and type of indwelling device. We hypothesize that spatial patterns of metabolic activity within a biofilm are influenced by spatial patterns in the flow environment, and that these interactions cause biofilm complexity to increase over time. We also hypothesize that the flow environment affects biofilm antimicrobial susceptibility not only by influencing delivery of antimicrobials to cells within the biofilm but also by dictating metabolic gradients within the community. We propose to address these hypotheses through the following specific aims. Aim 1: Observe growth of mono- species biofilms in a planar flow cell in order to assess changes in biofilm morphology, transport patterns, and metabolic activity with increasing spatial variability in environmental flow conditions. Aim 2: Observe the effectiveness of antibiotic treatment in eradicating biofilms having different degrees of spatial complexity, and relate the distribution of local killing efficiency to spatial patterns in transport conditions and metabolic activity. Aim 3: Develop an improved numerical model to allow quantitative analysis of the effects described above. Aim 4: Use the model to clarify multi-scale flow-biofilm interactions, and particularly to evaluate the key features that contribute to the survival of subpopulations of cells in biofilms under antibiotic treatment. We propose to achieve these aims by using a combination of novel experiments and numerical modeling. We will conduct experiments on biofilm growth and treatment in a new experimental system that provides the ability to impose a precisely controlled degree of spatial variability in inflow and outflow patterns. Biofilm growth, changes in flow and oxygen distributions, transport of antibiotic, and the resulting cell death will all be observed directly in situ. We will utilize these new and unique observations to support development of a new numerical model for biofilm development, which will subsequently be used to simulate the effectiveness of antibiotic treatment in eradicating biofilms under different local growth conditions. This combination of measurements and modeling will provide unique insight into the way in which biofilm growth interacts with and modifies the external flow, and ultimately how this complex interaction controls the overall formation of the biofilm and the effects of introduced antimicrobial agents on cells residing in the biofilm matrix. PUBLIC HEALTH RELEVANCE: Metabolic heterogeneity and antibiotic susceptibility in biofilms Summary Narrative Biofilm-based infections of inserted and implanted medical devices such as catheters, neurosurgical devices, and orthopedic devices are difficult to treat. Low rates of antibiotic transport within biofilms, protective effects of the biofilm matrix, and low rates of metabolic activity within the biofilm interior have all been found to contribute to the persistence of these infections, but there is currently little understanding of the processes responsible for these effects. The proposed work will advance understanding of how local environmental conditions influence biofilm growth, and will develop improved tools for assessing the effectiveness of antibiotics against biofilm-based infections.