There is a 'coffee ring' in the Bacillus subtilis biofilm center, and the colony biofilm morphologies are distinct inside and outside the 'coffee ring'. In this paper, we study this morphological difference and explain the reasons of the 'coffee ring' formation and further the causes to the morphological variation. We developed a quantitative method to characterize the surface morphology, the outer area is thicker than the inner area of the 'coffee ring', and the thickness amplitude in outer area is larger than inner area of the 'coffee ring'. We adopt a logistic growth model to obtain how the environmental resistance influence the colony biofilm thickness. Dead cells provide gaps for stress release and make folds formation in colony biofilm. we developed a technique for optical imaging and matching cells with the BRISK algorithm to capture the distribution and movement of motile cells and matrix producing cells in the colony biofilm. Matrix producing cells are mainly distribute in the outside of the 'coffee ring', and the extracellular matrix (ECM) prevents the motile cells moving outward from center. Motile cells mainly locate inside the ring, a small amount of dead motile cells outside the 'coffee ring' give rise to radial folds formation. There are no ECM blocking cell movements inside the ring, which result in uniform folds formation. The distribution of ECM and different phenotypes lead to the formation of the 'coffee ring', which is verified by using eps and flagellar mutants.
Keywords: Bacillus subtilis biofilm; Coffee ring; Logistic growth model; Phenotypes; Surface morphology.
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