Characterization of Biofilms by Correlated Mass Spectrometric and Raman Imaging

描述（由申请人提供）：大多数细菌在自然和临床环境中生长为表面附着的生物膜，这是细菌群落，它们自组装成细胞外聚合物物质（EPS）的包膜基质。为了形成这些细菌生物膜群落并感染宿主细胞，一种被称为群体感应（quorum sensing， QS）的细胞间信号传递过程非常重要。对于条件致病菌铜绿假单胞菌，QS调节许多对生物膜起始、EPS产生和毒力重要的基因的表达。虽然在体外和动物模型中已经了解了许多调节生物膜形成的选择因素，但多物种细胞群形成生物膜的具体情况尚不清楚。相关质谱和拉曼成像将用于研究宿主相关微生物群落的多重分析方法。这些方法允许在表面原位生长的混合细菌群落中测定单个细菌种类及其微生物产物。长期目标是通过了解细菌相互作用和特征生物分子的产生来设计检测和诊断策略。为实现这一目标，将首先对机会致病菌铜绿假单胞菌进行实验，具体开发区分铜绿假单胞菌与其他细菌种类的方法，区分不同的铜绿假单胞菌菌株，并确定与毒力相关的四种铜绿假单胞菌分子的空间分布：酰基高丝氨酸内酯（AHL）、喹诺酮类药物，如喹诺酮假单胞菌信号（PQS）、鼠李糖脂和外聚合物质（EPS）。成功实施这一研究计划将导致细菌作图方法具有更高的灵敏度和分辨率，大大提高我们在混合样品中识别特定种类细菌的能力，并识别和绘制这些细菌产生的特定化学产物，这些化学产物对宿主定植、感染和毒力至关重要。

DESCRIPTION (provided by applicant): Most bacteria in natural and clinical settings grow as surface-attached biofilms, which are bacterial communities that have self-assembled into an encased matrix of extracellular polymeric substances (EPS). To form these bacterial biofilm communities and infect host cells, an intercellular signaling process described as quorum sensing (QS) is very important. For the opportunistic pathogen, Pseudomonas aeruginosa, QS regulates the expression of many genes important to biofilm initiation, EPS production, and virulence. While much has been learned about select factors that regulate biofilm formation in vitro and in animal models, the specifics by which multispecies groups of cells form biofilms are not yet clear. Correlated mass spectrometric and Raman imaging will be used to develop a multiplex analysis method for studying host-associated microbial communities. These methods allow the determination of individual bacterial species and their microbial products within a mixed bacterial community growing in situ on surfaces. A long-term goal is the design of detection and diagnostic strategies informed by an understanding of bacterial interactions and signature biomolecule production. Work toward this goal will begin by conducting experiments with the opportunistic pathogen Pseudomonas aeruginosa, specifically developing methods to distinguish P. aeruginosa from other bacterial species, distinguish separate P. aeruginosa strains from each other, and determine the spatial distribution of four types of P. aeruginosa molecules associated with virulence: acyl- homoserine lactones (AHL), quinolones, such as pseudomonas quinolone signal (PQS), rhamnolipids, and exopolymeric substances (EPS). Successfully implementing this research program will result in bacterial mapping methods with improved sensitivity and resolution, substantially enhancing our ability to identify specific species of bacteria in mixed samples, and to identify and map specific chemical products produced by these bacteria that are critical to host colonization, infection, and virulence.