Characterization of Biofilms by Correlated Mass Spectrometric and Raman Imaging

描述(申请人提供)：自然和临床环境中的大多数细菌以表面附着生物膜的形式生长，这些生物膜是自组装成包裹的胞外聚合物(EPS)基质的细菌群落。为了形成这些细菌生物膜群落并感染宿主细胞，被称为群体感应(Quorum Sensing，QS)的细胞间信号传递过程是非常重要的。对于条件致病菌铜绿假单胞菌，QS调节许多基因的表达，这些基因对生物被膜的启动、EPS的产生和毒力至关重要。虽然在体外和动物模型中调节生物膜形成的选定因素已经了解了很多，但多物种细胞组形成生物膜的具体情况尚不清楚。相关的质谱学和拉曼成像将被用来开发一种研究宿主相关微生物群落的多重分析方法。这些方法可以确定在表面就地生长的混合细菌群落中的单个细菌种类及其微生物产品。一个长期目标是设计检测和诊断策略，了解细菌相互作用和标志性生物分子生产。为实现这一目标，将从对条件致病菌铜绿假单胞菌进行实验开始，特别是开发方法将铜绿假单胞菌与其他细菌物种区分开来，将不同的铜绿假单胞菌菌株彼此区分开来，并确定与毒力相关的四种类型的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，如假单胞菌喹诺酮信号(PQS)、鼠李糖脂和胞外多聚体物质(EPS)。成功实施这项研究计划将带来灵敏度和分辨率更高的细菌图谱方法，大大增强我们在混合样本中识别特定细菌种类的能力，以及识别和绘制这些细菌产生的对宿主定植、感染和毒力至关重要的特定化学产物的能力。

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.