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Material properties and growth environments affect the surface morphology of biofilms. Taken the biofilm growing in competitive environments as the object, which is compared with the single biofilm, we find that the competitive environment has an impact on the biofilm thickness and wrinkle pattens. Through Diffusion Limited Growth (DLG) theoretical model analysis, it shows that the competitive environment is caused by cell competing for nutrition, and the competitive environment reacts on biofilm which affect the phenotypic differentiation, causing changes in the stiffness of the biofilm. Using the theoretical and finite element simulation, we compare these results of bi-layer and tri-layer film-substrate models with experimental observations, and find that tri-layer film-substrate model is in line with the reality, which means that the layer between the biofilm and substrate plays an import role for wrinkle formation. Based on the above analysis, we further study effects of biofilm stiffness and interlayer thickness on wrinkles under competitive environment.

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The surface morphology of mature biofilms is heterogeneous and can be divided into concentric rings wrinkles (I), labyrinthine networks wrinkles (II), radial ridges wrinkles(III) and branches wrinkles (IV), according to surface wrinkle structure and distribution characteristics. Due to the wrinkle structures, channels are formed between the biofilm and substrate and transport nutrients, water, metabolic products, etc. We find that expansion rate variations of biofilms growing on substrates with high and low agar concentration (1.5wt%, 2.0wt%,2.5wt%) are not in the same phase. In the first three days’ growth, the interaction stress between biofilm and each agar substrate increases, which makes the biofilm expansion rate decreases before wrinkle pattern IV (branches) comes up. After three days, in the later growth stage after wrinkle pattern IV appears, the biofilm has larger expansion rate growing on 2.0wt% agar concentration, which has the larger wrinkle distance in wrinkle pattern IV reducing energy consumption. Our study shows that the stiff substrate does not always inhibit the biofilm expansion, although it does in the earlier stage, after that, mature biofilms acquire larger expansion rate by adjusting the growth mode through the wrinkle evolution even in nutrient extremely depletion.