Cell differentiation and motion determine the Bacillus subtilis biofilm morphological evolution under the competitive growth.

The growth discrepancy of Bacillus subtilis biofilms along different directions under the competitive growth drive the formation of anisotropic biofilm morphology directly. Two biofilms growing from two inoculating positions with different distances exhibit promoting or inhibiting growth behavior. Here we develop an optical imaging technology to observe the cell differentiation and the growth dynamics when the biofilm grows. It shows that the spatiotemporal distribution of different phenotypes affects the biofilm morphological evolution. We develop a program to calculate the velocity of cell motion within the biofilm, which is based on the feature point matching approach. We find the cell differentiation ununiformity in the neighboring region and its opposite region leads to the cell velocity difference in the competitive environment, the different cell motion further influences the biofilm morphology evolution. When biofilms grow with a long inoculating distance, there is always a gap between the them; when biofilms grow with a short inoculating distance, two biofilms gradually merge into a whole. Our work establishes a relationship between microscopic cells and macroscopic biofilm morphological which enables us to study the competitive growth process of biofilms from multiple perspectives.