Through adaptive laboratory evolution (ALE) experiments, it was recently found that when a bacterial population was repetitively treated with antibiotics, they will adapt to the treatment conditions and become tolerant to the drug. In this study, we utilized an ampicillin-tolerant Escherichia coli population isolated from an ALE experiment to study the mechanisms of persistence during ampicillin treatment and resuscitation. Interestingly, the persisters of this population exhibit filamentous morphology upon ampicillin treatment, and the filaments are getting longer over time. Proteomics analysis showed that proteins involved in carbohydrate metabolism are upregulated during antibiotic treatment, in addition to those involved in the oxidative stress response. Bacterial SOS response, which is associated with filamentation, was found to be induced on account of the increasing expression of RecA. Measurement of endogenous reactive oxygen species (ROS) revealed that the population have ∼100-fold less ROS generation under ampicillin treatment than the wild type, leading to a lower mutagenesis rate. Single-cell observations through time-lapse microscopy show that resuscitation of the filaments is stochastic. During resuscitation, proteins involved in the tricarboxylic acid (TCA) cycle, glyoxylate cycle and glycolytic processes, and ATP generation are downregulated, while ribosomal proteins and porins are upregulated in the filaments. One particular protein, ElaB, was upregulated by over 7-fold in the filaments after 3 h of resuspension in fresh medium, but its expression went down after the filaments divided. Knockout of elaB increased persistence on wild-type E. coli, and upon resumption of growth, mutants lacking elaB have a higher fraction of small colony variants (SCVs) than the wild type.

中文翻译：

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蛋白质组学研究从循环氨苄青霉素治疗的不断进化的大肠杆菌种群中丝状体的存活和复苏机制。

通过自适应实验室进化（ALE）实验，最近发现，当用抗生素重复治疗细菌种群时，它们将适应治疗条件并变得对药物耐受。在这项研究中，我们利用了耐氨苄西林的大肠杆菌人群从ALE实验中分离出来，以研究氨苄青霉素治疗和复苏过程中的持久性机制。有趣的是，在氨苄青霉素处理后，该种群的持久性表现出丝状形态，并且随着时间的流逝，这些细丝变得越来越长。蛋白质组学分析表明，除了涉及氧化应激反应的蛋白质外，在抗生素治疗过程中与碳水化合物代谢有关的蛋白质也被上调。由于RecA的表达增加，发现与细丝化有关的细菌SOS反应被诱导。内源性活性氧（ROS）的测量表明，该种群在氨苄西林处理下的ROS生成量比野生型少约100倍，从而导致诱变率降低。通过延时显微镜对单细胞进行观察表明，细丝的复苏是随机的。在复苏过程中，参与三羧酸（TCA）循环，乙醛酸循环和糖酵解过程以及ATP生成的蛋白质被下调，而核糖体蛋白和孔蛋白在细丝中被上调。在新鲜培养基中重悬3小时后，一种特殊的蛋白质ElaB在细丝中上调了7倍以上，但在细丝分裂后其表达却下降了。淘汰赛 在新鲜培养基中重悬3 h后，细丝中的蛋白表达上调了7倍以上，但细丝分裂后其表达下降了。淘汰赛 在新鲜培养基中重悬3 h后，细丝中的蛋白上调了7倍以上，但细丝分裂后其表达下降了。淘汰赛elaB增加了对野生型大肠杆菌的持久性，并且在恢复生长后，缺少elaB的突变体具有比野生型更高的小菌落变体（SCV）比例。