Osmotic spreading of Bacillus subtilis biofilms driven by an extracellular matrix

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General Information

Title: Osmotic spreading of Bacillus subtilis biofilms driven by an extracellular matrix

Authors: Agnese Seminaraa, Thomas E. Angelinic, James N. Wilkingb, Hera Vlamakisd, Senan Ebrahime, Roberto Kolterd, David A. Weitzb, and Michael P. Brenner

Keywords: Thin films, Spreading, Osmosis

Introduction

Bacteria thrive in colonies which form stable biological havens called biofilms on stagnant nutrient-rich substrates. Biofilms are highly organized bio-masses composed of differentiated bacterial cells and their cellular secretions that form a matrix around the cells. The composition and functionalities of these extracellular matrices (ECM) are very intriguing; to name a few, they provide protective environs for bacteria and also serve as nutrient reservoirs. Biofilms are hardly static but given enough nutrients in the host medium, they grow and spread. The spread of biofilms may be linked to active motility and multiplication of bacterial cells themselves or changes in the surrounding extracellular matrix or both. In the paper, the authors for the first time provide evidence that biofilm spreading is largely driven by osmotic forces in the extracellular matrix rather than by cellular motility.

Summary and Discussion

The central hypothesis of the paper is that the exopolysaccharide (EPS) component of ECM is mainly responsible for generation of osmotic forces which lead to swelling and spreading of biofilms. The proposition is elaborated by a mathematical model and supported by experimental measurements on different strains of biofilms with or without EPS. Three different strains of Bacillus subtilis bacteria are cultured on agar plates - flagellated wild type (WT), flagella-lacking mutant (hag) and EPS-deficient mutant (eps). WT and hag can produce EPS in their ECM while eps strain cannot. Their respective biofilms are observed over time (see Fig. 1). While the expansions of WT and hag are comparable, that of eps is greatly reduced (Fig. 1 A and B), suggesting that whereas the presence of flagella (active motility) has little statistical significance to expansion, the absence of EPS hinders expansion significantly. The authors also made sure that this discrepancy is not due to any inherent growth defect in eps strain; they observed comparable growth rates for all three strains in non-biofilm producing "shaking liquid" cultures (Fig. 1 C).

Figure 1: Expansion of bacterial biofilms. Figure taken from [1]

The authors posited that as the colonies grow, WT and hag strains secrete EPS into their ECMs which absorb water from the surrounding agar solution; as a result, their biofilms swell and spread. To quantify this, the authors propose a simple mathematical model based on mass conservation and free energy minimization, calculating the time dependence of biomass fraction (cells + ECM) and water fraction of the biofilm. Their model accurately predicts that after osmosis, WT and hag biofilms will swell vertically first (increase in h), followed by horizontal spreading (increase in R, see Fig. 2), whereas such events are not observed in the eps biofilm. Thus, the findings prove consistent with their hypothesis.

Figure 2: Time dependence of biofilm height and radius: experiment and theory. Figure taken from [1]

The spreading of bacterial biofilms is apparently governed by active hydrodynamic forces interacting between the films and the substrates. In this sense, the phenomenon is very different from those occurring in conventional thin films with Van der Waals interactions. Yet, it would be interesting to know more about these biofilms from the perspective of surface thermodynamics, for example the stability of these biofilms on differing host media. Nonetheless, the findings presented in the paper are nothing less than intriguing.

References

[1] Agnese Seminara et al., Osmotic spreading of Bacillus subtilis biofilms driven by an extracellular matrix, PNAS 1109261108. (2012)