Substrate stiffness impacts early biofilm formation by modulating twitching motility

Biofilms stem from the surface colonization of a wide variety of substrates, from living tissues to inert materials. The influence of mechanical interactions during this process is still unclear. Here, we explore the role of substrate rigidity on early biofilm development, using Pseudomonas aeruginosa as a model organism.

We embed hydrogel substrates of tunable rigidity in microfluidic channels, and image adhering bacteria in situ under constant flow, until colony formation. Our experiments show a strong dependence of colony phenotype on the underlying substrate stiffness. On soft hydrogels, P. aeruginosa assembles into dense hemispherical colonies, while on stiffer substrates it forms thin spread-out colonies. By tracking individual bacteria, we show that this difference in bacterial organization derives from a difference in bacterial surface motility. We propose a simple mechanistic model for the function of type-4 pili (T4P) on a deformable substrate, that accounts for the observed results. Finally, we show that this modulation of twitching motility by substrate stiffness can have far-reaching consequences for the biofilm, e.g. regarding EPS distribution or gene regulation.