Laboratory of Microbiology, Institute of Biology, University of Neuchatel, Neuchatel, Switzerland.
Department of Civil, Institute for Environmental Engineering, Environmental and Geomatic Engineering, ETH Zurich, Zurich, Switzerland.
Microbiology (Reading). 2023 Feb;169(2). doi: 10.1099/mic.0.001299.
The production of specialized resting cells is a remarkable survival strategy developed by many organisms to withstand unfavourable environmental factors such as nutrient depletion or other changes in abiotic and/or biotic conditions. Five bacterial taxa are recognized to form specialized resting cells: Firmicutes, forming endospores Actinobacteria forming exospores Cyanobacteria forming akinetes; the δ-Proteobacterial order Myxococcales, forming myxospores; and Azotobacteraceae, forming cysts. All these specialized resting cells are characterized by low-to-absent metabolic activity and higher resistance to environmental stress (desiccation, heat, starvation, etc.) when compared to vegetative cells. Given their similarity in function, we tested the potential existence of a universal morpho-chemical marker for identifying these specialized resting cells. After the production of endospores, exospores, akinetes and cysts in model organisms, we performed the first cross-species morphological and chemical comparison of bacterial sporulation. Cryo-electron microscopy of vitreous sections (CEMOVIS) was used to describe near-native morphology of the resting cells in comparison to the morphology of their respective vegetative cells. Resting cells shared a thicker cell envelope as their only common morphological feature. The chemical composition of the different specialized resting cells at the single-cell level was investigated using confocal Raman microspectroscopy. Our results show that the different specialized cells do not share a common chemical signature, but rather each group has a unique signature with a variable conservation of the signature of the vegetative cells. Additionally, we present the validation of Raman signatures associated with calcium dipicolinic acid (CaDPA) and their variation across individual cells to develop specific sorting thresholds for the isolation of endospores. This provides a proof of concept of the feasibility of isolating bacterial spores using a Raman-activated cell-sorting platform. This cross-species comparison and the current knowledge of genetic pathways inducing the formation of the resting cells highlights the complexity of this convergent evolutionary strategy promoting bacterial survival.
专门化休眠细胞的产生是许多生物为了应对不利的环境因素（如营养物质耗尽或非生物和/或生物条件的其他变化）而发展出来的一种非凡的生存策略。有五个细菌分类群被认为能够形成专门化的休眠细胞：厚壁菌门，形成芽孢；放线菌门，形成内生孢子；蓝细菌门，形成异形胞；δ-变形菌门的黏球菌目，形成黏液孢子；以及固氮菌科，形成胞囊。与营养细胞相比，所有这些专门化的休眠细胞的特征是代谢活性低甚至没有，并且对环境压力（干燥、热、饥饿等）的抵抗力更高。鉴于它们在功能上的相似性，我们测试了是否存在一种通用的形态化学标记物来识别这些专门化的休眠细胞。在模式生物中产生芽孢、内生孢子、异形胞和胞囊后，我们首次对细菌孢子形成进行了跨物种的形态和化学比较。使用玻璃切片的冷冻电子显微镜（CEMOVIS）来描述休眠细胞的近天然形态与它们各自的营养细胞的形态进行比较。休眠细胞仅具有较厚的细胞壁这一共有的形态特征。使用共聚焦拉曼微光谱技术研究了不同专门化休眠细胞在单细胞水平上的化学组成。我们的结果表明，不同的专门化细胞没有共同的化学特征，而是每个组都具有独特的特征，并且营养细胞的特征有一定程度的保留。此外，我们还展示了与钙二吡咯酸（CaDPA）相关的拉曼特征的验证及其在单个细胞中的变化，以开发用于分离芽孢的特定分选阈值。这为使用拉曼激活细胞分选平台分离细菌孢子的可行性提供了概念验证。这种跨物种比较以及目前关于诱导休眠细胞形成的遗传途径的知识突出了这种促进细菌生存的趋同进化策略的复杂性。
