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

Creating adaptive, sustainable, and dynamic biomaterials is a forthcoming mission of synthetic biology. Engineering spatially organized bacterial communities has a potential to develop such bio-metamaterials. However, generating living patterns with precision, robustness, and a low technical barrier remains as a challenge. Here we present an easily implementable technique for patterning live bacterial populations using a controlled meniscus-driven fluidics system, named as MeniFluidics. We demonstrate multiscale patterning of biofilm colonies and swarms with submillimeter resolution. Utilizing the faster bacterial spreading in liquid channels, MeniFluidics allows controlled bacterial colonies both in space and time to organize fluorescently labeled Bacillus subtilis strains into a converged pattern and to form dynamic vortex patterns in confined bacterial swarms. The robustness, accuracy, and low technical barrier of MeniFluidics offer a tool for advancing and inventing new living materials that can be combined with genetically engineered systems, and adding to fundamental research into ecological, evolutional, and physical interactions between microbes.

Electrically induced bacterial membrane-potential dynamics correspond to cellular proliferation capacity.

Membrane-potential dynamics mediate bacterial electrical signaling at both intra- and intercellular levels. Membrane potential is also central to cellular proliferation. It is unclear whether the cellular response to external electrical stimuli is influenced by the cellular proliferative capacity. A new strategy enabling electrical stimulation of bacteria with simultaneous monitoring of single-cell membrane-potential dynamics would allow bridging this knowledge gap and further extend electrophysiological studies into the field of microbiology. Here we report that an identical electrical stimulus can cause opposite polarization dynamics depending on cellular proliferation capacity. This was demonstrated using two model organisms, namely Bacillus subtilis and Escherichia coli, and by developing an apparatus enabling exogenous electrical stimulation and single-cell time-lapse microscopy. Using this bespoke apparatus, we show that a 2.5-second electrical stimulation causes hyperpolarization in unperturbed cells. Measurements of intracellular K+ and the deletion of the K+ channel suggested that the hyperpolarization response is caused by the K+ efflux through the channel. When cells are preexposed to 400 ± 8 nm wavelength light, the same electrical stimulation depolarizes cells instead of causing hyperpolarization. A mathematical model extended from the FitzHugh-Nagumo neuron model suggested that the opposite response dynamics are due to the shift in resting membrane potential. As predicted by the model, electrical stimulation only induced depolarization when cells are treated with antibiotics, protonophore, or alcohol. Therefore, electrically induced membrane-potential dynamics offer a reliable approach for rapid detection of proliferative bacteria and determination of their sensitivity to antimicrobial agents at the single-cell level.