Pattern Engineering of Living Bacterial Colonies Using Meniscus-Driven Fluidic Channels

Abstract

Engineering spatially organized biofilms for creating adaptive and sustainable biomaterials is a forthcoming mission of synthetic biology. Existing technologies of patterning biofilm materials suffer limitations associated with the high technical barrier and the requirements of special equipment. Here we present controlled meniscus-driven fluidics, MeniFluidics; an easily implementable technique for patterning living bacterial populations. We demonstrate multiscale patterning of living-colony and biofilm formation with submillimetre resolution. Relying on fast bacterial spreading in liquid channels, MeniFluidics allows controlled anisotropic bacterial colonies expansion both in space and time. The technique has also been applied for studying collective phenomena in confined bacterial swarming and organizing different fluorescently labelled Bacillus subtilis strains into a converged pattern. We believe that the robustness and low technical barrier of MeniFluidics offer a tool for developing living functional materials, bioengineering and bio-art, and adding to fundamental research of microbial interactions.

Biofilm and swarming emergent behaviours controlled through the aid of biophysical understanding and tools

“…While a detailed analysis is yet to be done, a similar pattern has been observed in swarming B. subtilis in a meniscus open channel, named MeniFluidics [72]. Such theoretical models and experimental techniques for engineering dynamic swarming patterns could lead to the developments of active-liquid metamaterials [73] and to identify swarming bacteria in physiological conditions [72]. The latter represents a challenge since swarming is mainly observed on agar surfaces.…”

“…A theoretical model of active fluidics has suggested that the confinement geometry could result in dynamic patterns, including vortex lattices [70,71]. While a detailed analysis is yet to be done, a similar pattern has been observed in swarming B. subtilis in a meniscus open channel, named MeniFluidics [72]. Such theoretical models and experimental techniques for engineering dynamic swarming patterns could lead to the developments of active-liquid metamaterials [73] and to identify swarming bacteria in physiological conditions [72].…”

“…Increasing cell densities enhance bacteria motility [18,50] Inhomogeneous cell density patterns of living and dead cells shape biofilm's wrinkle architecture [58] and EPS production is thickness dependent [60] Light Light drives bulk swarm motility in phototactic bacteria and blue light can slow down cells and lead to local cell accumulation [65,66] Blue light can be used to pattern biofilms in genetically engineered cells or to induce biofilm dispersal through cell hyperpolarization [74][75][76][77]79] Mechanical patterning Geometric confinement induces vortex patterns useful to identify swarming motility [70,71] 3D printing and Menifluidics can direct biofilm growth with submillimetre precision [72,[81][82][83]…”

Novel Tunable Spatio-Temporal Patterns From a Simple Genetic Oscillator Circuit

“…There are a vast number of experimental conditions which could be created to induce different spatio-temporal patterns in such microcolonies. Microfluidics has shown to be of particular help to control mechanical constraints (Ruprecht et al, 2017 ), substrate stiffness (Wang et al, 2018 ), nutrients (Alnahhas et al, 2019 ), chemical inducers (Danino et al, 2010 ), cell-cell signaling (Alnahhas et al, 2019 ), and pattern formation (Kantsler et al, 2020 ). As we showed in Figure 9 , controlling biophysical constraints using different channel layouts and mechanical properties of the substrate could produce different patterns of growth rate that give rise to structures that mimic different stages of the development of organisms (Johnson et al, 2017 ; Toda et al, 2018 ).…”

Novel tunable spatio-temporal patterns from a simple genetic oscillator circuit

“…There are a vast number of experimental conditions which could be created to induce different spatio-temporal patterns in such microcolonies. Microfluidics has shown to be of particular help to control mechanical constraints [65], substrate stiffness [66], nutrients [67], chemical inducers [13], cell-cell signaling [67] and pattern formation [68]. As we showed in figure 9, controlling biophysical constraints using different channel layouts and mechanical properties of the substrate could produce different patterns of growth rate that give rise to structures that mimic different stages of the development of organisms [25, 69].…”