Department of Molecular Biology and Microbial Food Safety, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands; Department of Mass Spectrometry of Bio-macromolecules, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands.
Department of Molecular Biology and Microbial Food Safety, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands; Van Leeuwenhoek Centre for Advanced Microscopy, Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands.
Food Microbiol. 2019 Aug;81:89-96. doi: 10.1016/j.fm.2018.03.006. Epub 2018 Mar 14.
Bacterial spores are ubiquitous in nature and can withstand both chemical and physical stresses. Spores can survive food preservation processes and upon outgrowth cause food spoilage as well as safety risks. The heterogeneous germination and outgrowth behavior of isogenic spore populations exacerbates this risk. A major unknown factor of spores is likely to be the inherently heterogeneous spore protein composition. The proteomics methods discussed here help in broadening the knowledge about spore structure and identification of putative target proteins from spores of different spore formers. Approaches to synchronize Bacillus subtilis spore formation, and to analyze spore proteins as well as the physiology of spore germination and outgrowth are also discussed. Live-imaging and fluorescence microscopy techniques discussed here allow analysis, at single cell level, of the 'germinosome', the process of spore germination itself, spore outgrowth and the spore intracellular pH dynamics. For the latter, a recently published improved pHluorin (IpHluorin) under control of the ptsG promoter is applicable. While the data obtained from such tools offers novel insight in the mechanisms of bacterial spore awakening, it may also be used to probe candidate antimicrobial compounds for inhibitory effects on spore germination and strengthen microbial risk assessment.
细菌孢子在自然界中无处不在，能够耐受化学和物理压力。孢子可以在食品保存过程中存活下来，并在生长后导致食品变质和安全风险。同基因孢子群体的异质萌发和生长行为加剧了这种风险。孢子的一个主要未知因素可能是其固有的孢子蛋白组成的异质性。本文讨论的蛋白质组学方法有助于拓宽对孢子结构的认识，并从不同孢子形成体的孢子中鉴定出潜在的靶蛋白。本文还讨论了同步枯草芽孢杆菌孢子形成的方法，以及分析孢子蛋白以及孢子萌发和生长的生理学的方法。本文讨论的活细胞成像和荧光显微镜技术允许在单细胞水平上分析“萌发体”，即孢子萌发本身的过程、孢子生长和孢子细胞内 pH 动力学。对于后者，可以应用在 ptsG 启动子控制下的新型 pHlourin（IpHlourin）。虽然从这些工具获得的数据为细菌孢子苏醒的机制提供了新的见解，但它也可用于探测候选抗菌化合物对孢子萌发的抑制作用，并加强微生物风险评估。
