Matrix production inBacillus subtilisbiofilms is localized to a propagating front

Abstract

Bacterial biofilms are surface attached microbial communities encased in self-produced extracellular polymeric substances (EPS). Development of a mature biofilm must require coordinating cell differentiation and multicellular activity at scales much larger than the single microbial unit. Here we demonstrate that during development of Bacillus subtilis biofilms, EPS matrix production is localized to a front propagating at the periphery. We show that within the front, cells switch off matrix production and transition to sporulation after a set time delay of ∼ 100 min. Correlation analyses of fluctuations in fluorescence reporter activity reveals that the front emerges from a pair of gene expression waves of matrix production and sporulation. The expression waves travel across cells that are immobilized in the biofilm matrix, in contrast to active cell migration or horizontal colony spreading. A single length scale and time scale couples the spatiotemporal propagation of both fronts throughout development, with the front displacement obeying a t 1/2 scaling law. As a result, gene expression patterns within the advancing fronts collapse to self-similar expression profiles. Our results indicate that development of bacterial biofilms may be governed by universal wave-like dynamics localized to a self-similar front.The vast majority of bacteria do not exist as solitary cells but within structurally complex communities known as bacterial biofilms [1]. From dental plaques to the alkaline hot springs of Yellowstone, bacteria in natural aquatic and terrestrial ecosystems [2][3][4][5] exist in these surface-associated aggregates encased in a self-produced matrix, known as the extracellular polymeric substance (EPS) [6][7][8]. The EPS matrix is primarily composed of exopolysachharides and proteins [9]. The production of EPS facilitates the construction of sophisticated threedimensional structures [10]. Additionally, the rigid scaffold supports the immobilization of individual microorganisms, allowing for cellular signaling and the creation of localized homeostatic zones [11]. Thus, the EPS matrix supports the biofilm's robust physiology by facilitating reproducible spatial patterns of gene expression, cellular differentiation, and morphology in a manner analogous to multicellular organisms [12][13][14][15]. As a result, biofilms are capable of performing a plethora of sophisticated functions, including promoting surface adhesion and aggregation [16], enhancing mechanical rigidity [17], advanced architecture for water retention and uptake of nutrients [18], promoting cell-cell communication [19], and conferring enhanced antibiotic resistance [20]. Thus it is key for microorganisms to have a robust collective strategy for regulating biofilm matrix production and spore generation.Bacillus subtilis is a convenient model bacteria to work with in view of the extensive single-cell molecular analyses focused on investigating the lineage of biofilm formation [21]. In the early stages of the B. subtilis biofilm de- * shmuel@seas.harva...