Wrinkles emerge from matrix complementarity in heterogenous biofilms

Abstract Biofilms are dynamic communities of microorganisms encased in a self-produced extracellular matrix. These resilient structures pose challenges across nearly all human activities, from healthcare to industry. The mechanical behaviour of a biofilm is shaped by the heterogenous composition of its matrix, both spatially and chemically. In turn, these mechanics influence the biofilm's architecture at micro- and macroscopic scales, driving its complexity and adaptability. Morphologically, this is reflected in mechanical deformations of the biofilm known as wrinkles. In nature, biofilms often host different species of bacteria, allowing for a great diversity of matrix components. Here, we use a combination of two \textit{Escherichia coli} strains as a model for a multispecies biofilm in which each bacterial strain produces one of two complementary matrix fibres: an amyloid protein (curli) or a polysaccharide (phosphoethanolamine-cellulose). Using fluorescence microscopy, we confirm that the two bacterial strains rapidly segregate into isogenic sectors, decreasing local heterogeneity. Furthermore, we show how wrinkles form, both in the homogenous central region of the biofilm, as well as at the boundary between sectors (i.e. where the two matrix producers co-localize). Finally, we show that increasing strain intermixing via the addition of bacteriophages results in thicker, taller wrinkles, irrespective of whether the two fibres are produced by two different strains or co-produced by the same bacteria.