Department of Bioengineering, University of California San Diego, La Jolla, California, USA.
Collaborative to Halt Antibiotic-Resistant Microbes, Department of Pediatrics, University of California San Diego, La Jolla, California, USA.
Appl Environ Microbiol. 2019 Oct 16;85(21). doi: 10.1128/AEM.01773-19. Print 2019 Nov 1.
is a Gram-positive pathogenic bacterium that colonizes an estimated one-third of the human population and can cause a wide spectrum of disease, ranging from superficial skin infections to life-threatening sepsis. The adaptive mechanisms that contribute to the success of this pathogen remain obscure partially due to a lack of knowledge of its metabolic requirements. Systems biology approaches can be extremely useful in predicting and interpreting metabolic phenotypes; however, such approaches rely on a chemically defined minimal medium as a basis to investigate the requirements of the cell. In this study, a chemically defined minimal medium formulation, termed synthetic minimal medium (SMM), was investigated and validated to support growth of three strains: LAC and TCH1516 (USA300 lineage), as well as D592 (USA100 lineage). The formulated SMM was used in an adaptive laboratory evolution experiment to probe the various mutational trajectories of all three strains leading to optimized growth capabilities. The evolved strains were phenotypically characterized for their growth rate and antimicrobial susceptibility. Strains were also resequenced to examine the genetic basis for observed changes in phenotype and to design follow-up metabolite supplementation assays. Our results reveal evolutionary trajectories that arose from strain-specific metabolic requirements. SMM and the evolved strains can also serve as important tools to study antibiotic resistance phenotypes of As researchers try to understand and combat the development of antibiotic resistance in pathogens, there is a growing need to thoroughly understand the physiology and metabolism of the microbes. is a threatening pathogen with increased antibiotic resistance and well-studied virulence mechanisms. However, the adaptive mechanisms used by this pathogen to survive environmental stresses remain unclear, mostly due to the lack of information about its metabolic requirements. Defining the minimal metabolic requirements for growth is a first step toward unraveling the mechanisms by which it adapts to metabolic stresses. Here, we present the development of a chemically defined minimal medium supporting growth of three strains, and we reveal key genetic mutations contributing to improved growth in minimal medium.
是一种革兰氏阳性的致病细菌，估计有三分之一的人类人口定植，可引起广泛的疾病，从浅表皮肤感染到危及生命的败血症。导致这种病原体成功的适应机制仍然不清楚，部分原因是缺乏对其代谢需求的了解。系统生物学方法在预测和解释代谢表型方面非常有用；然而，这些方法依赖于化学定义的最小培养基作为基础来研究细胞的需求。在这项研究中，一种化学定义的最小培养基配方，称为合成最小培养基 (SMM)，被研究和验证以支持三种菌株的生长：LAC 和 TCH1516（USA300 谱系），以及 D592（USA100 谱系）。配方 SMM 用于适应性实验室进化实验，以探测导致最佳生长能力的三种菌株的各种突变轨迹。进化后的菌株在生长速率和抗微生物敏感性方面进行了表型特征分析。还对菌株进行了重测序，以检查表型变化的遗传基础，并设计后续代谢物补充测定。我们的结果揭示了源自菌株特异性代谢需求的进化轨迹。SMM 和进化后的菌株也可以作为研究 的抗生素耐药表型的重要工具，因为研究人员试图了解和对抗病原体中抗生素耐药性的发展，因此越来越需要彻底了解微生物的生理学和代谢。 是一种具有增加的抗生素耐药性和经过充分研究的毒力机制的威胁病原体。然而，这种病原体用于应对环境压力的适应机制仍不清楚，主要是因为缺乏关于其代谢需求的信息。定义 生长的最小代谢需求是揭示其适应代谢应激机制的第一步。在这里，我们提出了一种支持三种 菌株生长的化学定义最小培养基的开发，并揭示了导致在最小培养基中生长改善的关键基因突变。
