Elucidating stress responses in bacterial biofilms using electromagnetic sensors and transcriptome analysis on single-cell level

使用电磁传感器和单细胞水平的转录组分析阐明细菌生物膜的应激反应

• 批准号: 272139544
• 负责人: Professor Dr.-Ing. Rolf Jakoby
• 金额: --
• 依托单位: Institut für Mikrosystemtechnik (IMTEK)
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/272139544?language=en
• 关键词: Elucidating; stress; responses; bacterial; biofilms

Biofouling is ubiquitous in nature. The formation of these so-called biofilms poses significant challenges for technical systems in aqueous environments. Biofilms are sessile formations of bacteria on substrates which coexist symbiotically mutually profiting metabolically and sharing protection strategies. Bacteria organized in biofilms are significantly better protected against environmental hazards compared to the planktonic state in free solution. Besides being a major economic problem due to the reduction performance of technical systems (membranes, material properties, etc.) biofilms have become a major thread for human health. Their short genetic cycle allows for fast evolutionary adaptability which results, e.g., in the development of resistance to most antibiotics. This is a common problem in hospitals where these persistent biofilms are a source of severe infections and similar diseases.In order to developed more sophisticated anti-biofilm treatment strategies quantitative tools are required which elucidate the physiological and genetic adaptability of bacteria in biofilms when probed with anti-biofilm reagents, e.g., for cleaning or disinfection. The aim of this project is to provide such a tool which allows determining physiological (morphological) and genetic changes quantitatively on a single-cell level. This tool will be used on single-species biofilms which will be subjected to various anti-biofilm treatment procedures (stress) in order to induce physiological and genetic responses. The biofilms will then be dissolved to raw cell solutions using suitable enzymatic cocktails that target the extra-cellular matrix. The cell solutions are then subjected to microfluidic flow-through filtration and mechanical trapping on a single-cell level. Once mechanically immobilized the cells are accessible for electrical characterization using a microwave electrochemical impedance spectroscopy based sensor system. The data acquired from the system will be matched to equivalent models extracting relevant parameters which are compared among bacteria originating from differently treated biofilms. In addition the cells will be extracted for single-cell gene expression analysis allowing correlation of electrical (physiological) changes to genetic changes.This combined approach will allow correlating stress responses of bacteria organized in biofilms with changes in their physiology and gene expression and thus allow the quantitative determination of responses of biofilms to various anti-biofilm treatment strategies. Using this tool a more detailed understanding of the mechanisms underlying the adaptability of bacteria in biofilms to treatment strategies will be enabled which will be the foundation for development of efficient strategies for biofilms removal.

生物污损在自然界中普遍存在。这些所谓的生物膜的形成对水性环境中的技术系统提出了重大挑战。生物被膜是细菌在基质上的固着结构，它们共生共存，在代谢上互惠互利，共享保护策略。组织在生物膜中的细菌与游离溶液中的无菌状态相比，明显更好地保护其免受环境危害。除了由于技术系统（膜、材料特性等）的性能降低而成为主要的经济问题之外，生物膜已成为人类健康的主要线索。它们的短遗传周期允许快速进化适应性，例如，对大多数抗生素产生耐药性的过程。这是医院中的一个常见问题，这些持久性生物膜是严重感染和类似疾病的根源。为了开发更复杂的抗生物膜治疗策略，需要定量工具来阐明生物膜中细菌的生理和遗传适应性。当用抗生物膜试剂进行探测时，例如，用于清洁或消毒。该项目的目的是提供这样一种工具，允许在单细胞水平上定量确定生理（形态）和遗传变化。该工具将用于单物种生物膜，其将经受各种抗生物膜处理程序（应激）以诱导生理和遗传反应。然后使用靶向细胞外基质的合适的酶混合物将生物膜溶解成原始细胞溶液。然后使细胞溶液在单细胞水平上经受微流体流通过滤和机械捕获。一旦机械固定的细胞是可访问的电特性，使用微波电化学阻抗谱为基础的传感器系统。从系统中获得的数据将与提取相关参数的等效模型相匹配，这些参数在来自不同处理的生物膜的细菌之间进行比较。此外，细胞将被提取用于单细胞基因表达分析，允许电（生理）变化与遗传变化的相关性。这种组合方法将允许将生物膜中组织的细菌的应激反应与其生理和基因表达的变化相关联，从而允许定量测定生物膜对各种抗生物膜治疗策略的反应。使用这个工具，更详细地了解生物膜中的细菌对处理策略的适应性的机制，这将是开发有效的生物膜去除策略的基础。