Complexity in the Bacterial Community

Abstract

Biofilms are organized communities of bacteria that form on surfaces. The bacterial population of a biofilm exhibits enhanced resistance to environmental stress and antibiotics, which has important implications for the clinic and industry. Vlamakis et al . integrated genes for fluorescent markers under the control of various specific promoters into Bacillus subtilis to visualize cellular differentiation within a biofilm, to follow cell lineage, and to show that the formation of extracellular matrix was necessary for sporulation within a biofilm. Bacteria expressing P hag -yfp were used to visualize motile cells, those expressing P yqxM -yfp revealed matrix-producing cells, and those expressing P sspB -yfp revealed sporulating cells. Biofilms containing the labeled cells were allowed to form for a period of time and then the cells were dispersed and fluorescence was measured by flow cytometry. At early time points (12 hours), most cells were motile; by 24 hours, the matrix-expressing cells were at a maximum; and by 48 hours, sporulation was readily detected. Thin-section microscopy of the biofilm revealed that the motile cells were at the base and edge of the biofilm, whereas the matrix-expressing cells were distributed in patches throughout the biofilm, and sporulation occurred preferentially in distinct regions called the aerial structures. Cell lineage was monitored by time-lapse microscopy of dual-labeled cells, in which pairwise combinations of variant fluorescent proteins under the control of different promoters were expressed. These assays revealed that motile cells gave rise to matrix-producing cells, which then differentiated to produce sporulating cells. Regions of motile cells in the biofilm were separated from regions of sporulating cells, whereas the regions containing matrix-producing and sporulating cells tended to overlap. Production of extracellular matrix was necessary for proper organization. Biofilms formed from tasA mutants that could not produce a critical matrix protein exhibited decreased abundance of motile cells, and their location was aberrant compared with a wild-type biofilm. The population of cells expressing P yqxM -yfp was elevated and, despite sporulating in liquid culture, there were no sporulating cells in biofilms formed by tasA mutants. Sporulation in the biofilm was restored if sporulation-deficient but matrix-production-competent cells were mixed with matrix-deficient but sporulation-competent cells. Because nutrient depletion is a stimulus that activates Spo0A, which through direct and indirect mechanisms controls both motility and sporulation, the authors suggested that matrix production may create local regions within the biofilm where nutrients are depleted, thus triggering the sporulation pathway through Spo0A activation. H. Vlamakis, C. Aguilar, R. Losick, R. Kolter, Control of cell fate by the formation of an architecturally complex bacterial community. Genes Dev. 22 , 945-953 (2008). [Abstract] [Full Text]