The competitive growth of Bacillus subtilis biofilms from the perspectives of thickness and self-healing.

Abstract

Under resource-limited conditions, biofilms display highly complex competitive behaviors and distinct growth dynamics. This study investigated Bacillus subtilis strain engineered with three fluorescent reporter genes by integrating microscopic and macroscopic analyses to uncover the intrinsic mechanisms driving heterogeneous growth under competitive constraints. The results show that stress accumulation leads to the formation of a pronounced thickness peak in the interfacial region between two neighboring biofilms under strong competition (inoculation-center distance, d = 2 mm). Under nutrient-rich conditions, a ring-shaped elevation emerges approximately 4 mm from the colony center, providing an expanded spatial niche that facilitates bacterial survival. Self-healing rates of incisions at different locations of biofilms were compared to evaluate the effect of competition. The findings show that the self-healing capacity of biofilms in competitive environments is governed by both biofilm age and competitive intensity. Incorporating nutrient diffusion, environmental pressure, and phenotype-switching probabilities, a cellular automaton model was further developed to simulate the competitive growth of the biofilms.