This paper describes a system to study how small physical perturbations can affect bacterial community behavior in unexpected ways through modulation of diffusion and convective transport of chemical communication molecules and resources. A culture environment that mimics the chemically open characteristic of natural bacterial habitats but with user-defined spatiotemporal control of bacteria microcolonies is realized through use of an aqueous two phase system (ATPS). The ATPS is formulated with nontoxic dextran (DEX) and poly(ethylene glycol) (PEG) dissolved in cell culture media. DEX-phase droplets formed within a bulk PEG-phase stably confine the bacteria within it while small molecules diffuse relatively freely. Bacteria-containing DEX droplets can also be magnetically relocated, without loss of its bacterial content, when DEX-conjugated magnetic particles are included. We found that decreasing the distance between quorum-sensing (QS)-coupled microcolonies increased green fluorescent protein (GFP) expression due to increased inter-colony chemical communication but with upper limits. Periodic relocation of the chemical signal receiver colony, however, increased GFP expression beyond these typical bounds predicted by quorum sensing concepts alone by maintaining inter-colony chemical communication while also relieving the colony of short-range resource depletion effects. Computer simulations suggest that such increased productive output in response to periodic nonlethal physical perturbations is a common feature of chemically activated interactive cell systems where there is also a short-range inhibitory effect. In addition to providing insights on the effect of bacteria relocation, the magnetic ATPS droplet manipulation capability should be broadly useful for bioanalyses applications where selective partitioning at the microscale in fully aqueous conditions is needed.

Using an aqueous two-phase system comprised of dextran and polyethylene glycol, this article describes the stable spatial patterning of sub-microlitre droplets of bacterial suspensions. Microdroplets of different types of bacterial populations are positioned and maintained adjacent to each other without significant dispersion even though the bacteria are in suspension and not surface bound. Small molecules, in contrast, diffuse relatively freely between the two aqueous phases. The usefulness of these capabilities is demonstrated by generating a small array of suspensions containing different Escherichia coli strains engineered to respond fluorescently or luminescently to different chemical stimuli. When a chemical stimulus is presented, this droplet array produces a pattern of bacterial "illumination" that reflects the type of chemical to which the array was exposed.

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We describe a microfluidic concentrator array device that is integrated with microfabricated ratchet structures to concentrate motile bacterial cells in desired destinations with required cell densities. The device consists of many pairs of concentrators with a wide range of spacing distances on a chip, and allows cells in one concentrator to be physically separated from but chemically connected to cells in the other concentrator. Therefore, the device facilitates quantification of the effect of spacing distance on the cell-to-cell communication of synthetically engineered bacterial cells. In addition, the device enables us to control the cell number density in each concentrator unit by adjusting the concentration time and the density of cell suspensions, and the basic concentrator unit of the device can be repeatedly duplicated on a chip. Hence, the device not only facilitates an investigation of the effect of cell densities on cell-to-cell communication, but it can also be further applied to an investigation of cellular communication among multiple types of cells. Lastly, the device can be easily fabricated using a single-layered soft-lithography technology so that we believe it would provide a simple but robust means for many synthetic and systems biologists to simplify and speed up their investigations of the synthetic genetic circuits in bacterial cells.

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