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Biofilms inhabit a range of environments, such as dental plaques or soil micropores, often characterized by intricate, non-even surfaces. However, the impact of surface irregularities on the population dynamics of biofilms remains elusive as most biofilm experiments are conducted on flat surfaces. Here, we show that the shape of the surface on which a biofilm grows influences genetic drift and selection within the biofilm. We culture E. coli biofilms in micro-wells with an undulating bottom surface and observe the emergence of clonal sectors whose size corresponds to that of the undulations, despite no physical barrier separating different areas of the biofilm. The sectors are remarkably stable over time and do not invade each other; we attribute this stability to the characteristics of the velocity field within the growing biofilm, which hinders mixing and clonal expansion. A microscopically-detailed computer model fully reproduces these findings and highlights the role of mechanical (physical) interactions such as adhesion and friction in microbial evolution. The model also predicts clonal expansion to be severely limited even for clones with a significant growth advantage - a finding which we subsequently confirm experimentally using a mixture of antibiotic-sensitive and antibiotic-resistant mutants in the presence of sub-lethal concentrations of the antibiotic rifampicin. The strong suppression of selection contrasts sharply with the behavior seen in bacterial colonies on agar commonly used to study range expansion and evolution in biofilms. Our results show that biofilm population dynamics can be controlled by patterning the surface, and demonstrate how a better understanding of the physics of bacterial growth can pave the way for new strategies in steering microbial evolution., Comment: 15 pages and 4 figures (main text), 13 pages and 6 figures (supplementary material)

We present Tuna-step, a novel microfluidic module based on step emulsification that allows for reliable generation of droplets of different sizes. Until now, sizes of droplets generated with step emulsification were hard-wired into the geometry of the step emulsification nozzle. To overcome this, we incorporate a thin membrane underneath the step nozzle that can be actuated by pressure, enabling the tuning of the nozzle size on-demand. By controllably reducing the height of the nozzle, we successfully achieved a three-order-of-magnitude variation in droplet volume without any adjustment of the flow rates of the two phases. We further enhanced our system by developing and applying a new hydrophilic surface modification, ensuring long-term stability and preventing swelling of the device when generating Oil-in-Water droplets. We used our system in the manufacturing of functionally graded soft materials with adjustable porosity and material content. By combining our microfluidic device with a custom 3D printer, we generated and extruded Oil-in-Water emulsions in an agarose gel bath, allowing for the creation of unique self-standing 3D hydrogel structures with porosity decoupled from flow rate and with composition gradients of external phases. We upscaled Tuna-step by setting 14 actuatable nozzles in parallel, offering a step-emulsification-based single chip solution that can accommodate various requirements in terms of throughput, droplet volumes, flow rates, and surface chemistry.