Isolation and physiological characterization of cell types from heterogeneous aggregates of Staphylococcus aureus using transcriptomic analysis

使用转录组分析从金黄色葡萄球菌异质聚集体中分离细胞类型并对其进行生理学表征

• 批准号: 217573181
• 负责人: Dr. Daniel López
• 金额: --
• 依托单位: Institut für Molekulare Infektionsbiologie (IMIB)
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2012
• 资助国家: 德国
• 起止时间: 2011-12-31 至 2017-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/217573181?language=en
• 关键词: Isolation; physiological; characterization; cell; types

During the first period of this SPP1617 program, we showed that multicellular aggregates of the bacterial pathogen Staphylococcus aureus are constituted by a heterogeneous population of, at least, two different cell types, biofilm producers and toxin producers. We used a number of biochemical techniques to characterize the molecular mechanism that drives to this cell differentiation. We found a complex genetic regulation triggered by the bimodal behavior of the quorum-sensing cascade agr, which ultimately selects for the progression of acute (higher concentration of toxin producers) or biofilm-associated chronic infections (higher concentration of biofilm producers). In this renewal, we will physiologically characterize these two distinct subpopulations that we find in S. aureus biofilms. To do this, we have setup a new cell-sorting methodology that permits the isolation of each one of these subpopulations to further analyze their transcriptomic profile after isolation. This approach will allow us determining the physiological characteristics of each cell type to better understand their specialization within the biofilm. Moreover, to test the versatility of our approach, we will provide this sorting + RNA-seq technology to any laboratory of this consortium that is interested in characterizing a subpopulation of cells. Furthermore, we will combine our cell sorting + RNA-seq technology with our state-of-the-art microscopic techniques to perform a spatio-temporal localization of the subpopulations within the S. aureus aggregates. The combination of these two approaches will provide a better understanding of the role of specialized cell types within microbial communities and whether this is connected to the specific physiological state of each cell type. The third goal of this proposal will study the differential sensitivity of the subpopulations, biofilm producers and toxin producers, to antimicrobial treatments that are conventionally used to eliminate S. aureus biofilms. Cell types with different transcriptomic profiles and different physiologies may display different sensitivities to antibiotics and may develop into difficult-to-treat staphylococcal infections. We will study the capacity of each cell type to resist an antibiotic treatment based on their physiological profile. Overall, our project will demonstrate that S. aureus communities are constituted by cell types that are physiologically different and will demonstrate that the general physiology of each cell type is related to their role within the biofilm and their capacity to resist antimicrobial treatments. Our approach should be of interest to other research groups of this consortium.

在SPP 1617项目的第一阶段，我们发现细菌病原体金黄色葡萄球菌的多细胞聚集体由至少两种不同细胞类型的异质群体构成，生物膜生产者和毒素生产者。我们使用了一些生物化学技术来表征驱动这种细胞分化的分子机制。我们发现了由群体感应级联agr的双峰行为触发的复杂的遗传调节，其最终选择急性（较高浓度的毒素产生者）或生物膜相关的慢性感染（较高浓度的生物膜产生者）的进展。在这次更新中，我们将从生理学上描述我们在S.金黄色葡萄球菌生物膜。为此，我们建立了一种新的细胞分选方法，允许分离这些亚群中的每一个，以在分离后进一步分析其转录组谱。这种方法将使我们能够确定每种细胞类型的生理特征，以更好地了解它们在生物膜内的专业化。此外，为了测试我们方法的多功能性，我们将向该联盟中任何有兴趣表征细胞亚群的实验室提供这种分选+ RNA-seq技术。此外，我们将联合收割机结合我们的细胞分选+ RNA-seq技术和我们最先进的显微镜技术，对S.金黄色聚集体。这两种方法的结合将更好地了解微生物群落中专门细胞类型的作用，以及这是否与每种细胞类型的特定生理状态有关。本提案的第三个目标将研究亚群生物膜生产者和毒素生产者对常规用于消除S的抗菌治疗的不同敏感性。金黄色葡萄球菌生物膜。具有不同转录组学特征和不同生理学的细胞类型可能对抗生素表现出不同的敏感性，并可能发展为难以治疗的葡萄球菌感染。我们将研究每种细胞类型的能力，以抵抗抗生素治疗的基础上，他们的生理概况。总的来说，我们的项目将证明S。金黄色葡萄球菌群落由生理学上不同的细胞类型构成，并且将证明每种细胞类型的一般生理学与它们在生物膜内的作用和它们抵抗抗微生物处理的能力有关。我们的方法应该对该联盟的其他研究小组感兴趣。