Department of Microbiology, University of Washington, Seattle, WA 98195, USA.
Proc Natl Acad Sci U S A. 2012 Nov 27;109(48):19804-9. doi: 10.1073/pnas.1213963109. Epub 2012 Nov 12.
Interbacterial interaction pathways play an important role in defining the structure and complexity of bacterial associations. A quantitative description of such pathways offers promise for understanding the forces that contribute to community composition. We developed time-lapse fluorescence microscopy methods for quantitation of interbacterial interactions and applied these to the characterization of type VI secretion (T6S) in Pseudomonas aeruginosa. Our analyses allowed a direct determination of the efficiency of recipient cell lysis catalyzed by this intercellular toxin delivery pathway and provided evidence that its arsenal extends beyond known effector proteins. Measurement of T6S apparatus localization revealed correlated activation among neighboring cells, which, taken together with genetic data, implicate the elaboration of a functional T6S apparatus with a marked increase in susceptibility to intoxication. This possibility was supported by the identification of T6S-inactivating mutations in a genome-wide screen for resistance to T6S-mediated intoxication and by time-lapse fluorescence microscopy analyses showing a decreased lysis rate of recipient cells lacking T6S function. Our discoveries highlight the utility of single-cell approaches for measuring interbacterial phenomena and provide a foundation for studying the contribution of a widespread bacterial interaction pathway to community structure.
细菌间相互作用途径在定义细菌群落的结构和复杂性方面起着重要作用。对这些途径进行定量描述有望理解促成群落组成的力量。我们开发了用于定量细菌间相互作用的延时荧光显微镜方法，并将其应用于铜绿假单胞菌中 VI 型分泌系统（T6S）的表征。我们的分析允许直接确定这种细胞间毒素输送途径催化的受体细胞裂解的效率，并提供证据表明其武器库超出了已知的效应蛋白。T6S 装置定位的测量显示出相邻细胞之间的相关性激活，这与遗传数据一起表明，一种功能齐全的 T6S 装置的精心设计会导致对中毒的敏感性明显增加。这一可能性得到了全基因组筛选 T6S 介导中毒抗性的 T6S 失活突变的鉴定以及延时荧光显微镜分析的支持，该分析表明缺乏 T6S 功能的受体细胞的裂解率降低。我们的发现强调了单细胞方法在测量细菌间现象方面的实用性，并为研究广泛存在的细菌相互作用途径对群落结构的贡献提供了基础。
