Moormeier Derek E, Bose Jeffrey L, Horswill Alexander R, Bayles Kenneth W
Center for Staphylococcal Research, Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA.
Department of Microbiology, Roy J. and Lucille Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA.
mBio. 2014 Oct 14;5(5):e01341-14. doi: 10.1128/mBio.01341-14.
Biofilm communities contain distinct microniches that result in metabolic heterogeneity and variability in gene expression. Previously, these niches were visualized within Staphylococcus aureus biofilms by observing differential expression of the cid and lrg operons during tower formation. In the present study, we examined early biofilm development and identified two new stages (designated "multiplication" and "exodus") that were associated with changes in matrix composition and a distinct reorganization of the cells as the biofilm matured. The initial attachment and multiplication stages were shown to be protease sensitive but independent of most cell surface-associated proteins. Interestingly, after 6 h of growth, an exodus of the biofilm population that followed the transition of the biofilm to DNase I sensitivity was demonstrated. Furthermore, disruption of the gene encoding staphylococcal nuclease (nuc) abrogated this exodus event, causing hyperproliferation of the biofilm and disrupting normal tower development. Immediately prior to the exodus event, S. aureus cells carrying a nuc::gfp promoter fusion demonstrated Sae-dependent expression but only in an apparently random subpopulation of cells. In contrast to the existing model for tower development in S. aureus, the results of this study suggest the presence of a Sae-controlled nuclease-mediated exodus of biofilm cells that is required for the development of tower structures. Furthermore, these studies indicate that the differential expression of nuc during biofilm development is subject to stochastic regulatory mechanisms that are independent of the formation of metabolic microniches. Importance: In this study, we provide a novel view of four early stages of biofilm formation by the human pathogen Staphylococcus aureus. We identified an initial nucleoprotein matrix during biofilm development that is DNase I insensitive until a critical point when a nuclease-mediated exodus of the population is induced prior to tower formation. Unlike the previously described dispersal of cells that occurs after tower development, we found that the mechanism controlling this exodus event is dependent on the Sae regulatory system and independent of Agr. In addition, we revealed that the gene encoding the secreted staphylococcal nuclease was expressed in only a subpopulation of cells, consistent with a model in which biofilms exhibit multicellular characteristics, including the presence of specialized cells and a division of labor that imparts functional consequences to the remainder of the population.
生物膜群落包含不同的微环境，这导致了代谢异质性和基因表达的变异性。此前，通过观察金黄色葡萄球菌生物膜形成过程中cid和lrg操纵子的差异表达，在生物膜内可视化了这些微环境。在本研究中，我们研究了生物膜的早期发育，并确定了两个新的阶段（称为“增殖”和“迁出”），这两个阶段与生物膜成熟过程中基质组成的变化以及细胞的明显重组有关。初始附着和增殖阶段对蛋白酶敏感，但独立于大多数细胞表面相关蛋白。有趣的是，生长6小时后，证明生物膜群体在生物膜转变为对DNase I敏感后出现迁出。此外，编码葡萄球菌核酸酶（nuc）的基因的破坏消除了这一迁出事件，导致生物膜过度增殖并破坏正常的塔状结构发育。就在迁出事件之前，携带nuc::gfp启动子融合的金黄色葡萄球菌细胞表现出Sae依赖性表达，但仅在明显随机的细胞亚群中表达。与金黄色葡萄球菌中现有的塔状发育模型相反，本研究结果表明存在一种由Sae控制的核酸酶介导的生物膜细胞迁出，这是塔状结构发育所必需的。此外，这些研究表明，生物膜发育过程中nuc的差异表达受随机调节机制的影响，这些机制独立于代谢微环境的形成。重要性：在本研究中，我们提供了人类病原体金黄色葡萄球菌生物膜形成四个早期阶段的新观点。我们确定了生物膜发育过程中的初始核蛋白基质，该基质对DNase I不敏感，直到在塔状结构形成之前诱导群体核酸酶介导的迁出的关键点。与先前描述的塔状发育后发生的细胞扩散不同，我们发现控制这一迁出事件的机制依赖于Sae调节系统且独立于Agr。此外，我们揭示编码分泌型葡萄球菌核酸酶的基因仅在细胞亚群中表达，这与生物膜表现出多细胞特征的模型一致，包括存在特化细胞和分工，这会给其余群体带来功能后果。
