Personnic Nicolas, Striednig Bianca, Hilbi Hubert
Institute of Medical Microbiology, University of Zürich, Zürich, Switzerland.
Methods Mol Biol. 2019;1921:191-204. doi: 10.1007/978-1-4939-9048-1_12.
Legionella pneumophila resides in multispecies biofilms, where it infects and replicates in environmental protozoa such as Acanthamoeba castellanii. Studies on L. pneumophila physiology and host-pathogen interactions are frequently conducted using clonal bacterial populations and population level analysis, overlooking the remarkable differences in single cell behavior. The fastidious nutrient requirements of extracellular L. pneumophila and the extraordinary motility of Acanthamoeba castellanii hamper an analysis at single cell resolution. In this chapter, we describe a method to study L. pneumophila and its natural host A. castellanii at single cell level by using an agarose embedment assay. Agarose-embedded bacteria and infected cells can be monitored over several hours up to several days. Using properly adapted flow chambers, agarose-embedded specimens can be subjected to a wide range of fluctuating conditions.
嗜肺军团菌存在于多物种生物膜中，在其中它会感染环境中的原生动物，如卡氏棘阿米巴，并在其体内进行复制。关于嗜肺军团菌生理学以及宿主-病原体相互作用的研究，通常使用克隆细菌群体以及群体水平分析，而忽略了单细胞行为的显著差异。胞外嗜肺军团菌对营养的苛刻需求以及卡氏棘阿米巴非凡的运动性，阻碍了单细胞分辨率水平上的分析。在本章中，我们描述了一种通过使用琼脂糖包埋测定法在单细胞水平研究嗜肺军团菌及其天然宿主卡氏棘阿米巴的方法。琼脂糖包埋的细菌和受感染细胞可以在数小时至数天的时间内进行监测。使用经过适当改造的流动腔室，琼脂糖包埋的标本可以经受各种波动条件。
