From non-inherited to heritable antimicrobial peptide resistance: priming, aging, tolerance and persistence

从非遗传性到遗传性抗菌肽耐药性：引发、老化、耐受性和持久性

• 批准号: 510239573
• 负责人: Professor Dr. Jens Rolff
• 金额: --
• 依托单位: Arbeitsgruppe Rolff - Evolutionsbiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/510239573?language=en
• 关键词: non; inherited; heritable; antimicrobial; peptide

Antimicrobial resistance evolution is a problem that requires evolutionary research, both, to understand host-microbe interactions and the evolution of drug resistance. Recently it has become clear that the evolution of resistance can be preceded by non-inheritable resistance mechanisms such as bacterial persistence and tolerance. This can be elicited by the exposure to sublethal antimicrobial concentrations. We have previously shown that prior short exposure to low concentrations (priming) of antimicrobial peptides (AMPs), ancient weapons of multicellular organisms, induce increased bacterial tolerance and persistence resulting in increased resistance evolution. Here we want to address three goals. (1) Disentangle AMP-priming mediated tolerance and persistence and their consequences experimentally. (2) Understand the role of AMP-priming under different kinetics: is the response to a continuously increasing concentration of stressors mediated by the same mechanisms as the priming response? (3) To investigate the role of cell age on primability by AMPs. Do older cells get primed to the same degree as younger cells? Goal 1 Bacterial priming by AMPs increases survival under lethal AMP-exposure. The surviving cells are either metabolically active dying at a slower rate or more persistent and metabolically inactive. To understand this biphasic response requires to synchronise cells using a device. Determining bacterial survival phenotypes will allow to explore when cells become either resistant (heritable) or tolerant/persistent (non-heritable). RNAseq on these synchronised cells will allow to identify candidate genes that can be followed up with functional analysis. Goal 2 Inducible immune responses of hosts such as AMPs almost always start from zero: after signalling of an immune insult, the expression of inducible immune effectors is continuously increasing. We here want to study whether bacterial priming (as manifested by increases in bacterial survival, persistence and tolerance) also occurs under the different stressor kinetics represented by continuously increasing stress. We will test this question using a microfluidic setup with E. coli. Goal 3 All cells age, also bacterial cells. Here by synchronizing cells and then use ‘baby’ cells of defined age we will investigate if cell age is related to primability and the duration of the priming response. This will be possible combining a cell synchronizing device with a microfluidic that allows to track the fate of individual cells. Addressing these three goals will allow us to understand the biphasic response, increased tolerance and persistence, after antimicrobial peptide induced priming. This will be essential to under the evolutionary dynamics resulting from priming and leading to antimicrobial resistance evolution.

抗生素耐药性进化是一个需要进化研究的问题，以了解宿主-微生物相互作用和耐药性进化。最近已经清楚的是，耐药性的进化可能先于不可遗传的耐药机制，例如细菌的持久性和耐受性。这可以通过暴露于亚致死浓度的抗菌剂引起。我们之前已经表明，先前短时间接触低浓度（启动）抗菌肽（AMP）（多细胞生物的古老武器）会诱导细菌耐受性和持久性增加，从而导致耐药性进化增加。在这里，我们要实现三个目标。 (1) 通过实验阐明 AMP 引发介导的耐受性和持久性及其后果。 (2)了解不同动力学下AMP启动的作用：对持续增加的应激源浓度的反应是否由与启动反应相同的机制介导？ (3)研究细胞年龄对AMPs引发能力的作用。老年细胞的启动程度是否与年轻细胞相同？目标 1 AMP 引发的细菌可提高致命 AMP 暴露下的存活率。幸存的细胞要么代谢活跃，以较慢的速度死亡，要么更持久且代谢不活跃。要了解这种双相反应，需要使用设备同步细胞。确定细菌存活表型将有助于探索细胞何时变得耐药（可遗传）或耐受/持久（不可遗传）。这些同步细胞上的 RNAseq 将能够识别可进行功能分析的候选基因。目标 2 AMP 等宿主的诱导性免疫反应几乎总是从零开始：发出免疫损伤信号后，诱导性免疫效应物的表达不断增加。我们在这里想要研究细菌启动（表现为细菌存活率、持久性和耐受性的增加）是否也会在以持续增加的压力为代表的不同压力源动力学下发生。我们将使用大肠杆菌的微流体装置来测试这个问题。目标 3 所有细胞都会衰老，细菌细胞也是如此。在这里，通过同步细胞，然后使用特定年龄的“婴儿”细胞，我们将研究细胞年龄是否与启动性和启动反应的持续时间有关。这将有可能将细胞同步装置与微流体结合起来，从而跟踪单个细胞的命运。解决这三个目标将使我们能够了解抗菌肽诱导启动后的双相反应、增强的耐受性和持久性。这对于引发并导致抗菌素耐药性进化所产生的进化动态至关重要。