Quorum sensing and chemotaxis both affect bacterial behavior on the population level. Chemotaxis shapes the spatial distribution of cells, while quorum sensing realizes a cell-density dependent gene regulation. An interesting question is if these mechanisms interact on some level: Does quorum sensing, a density dependent process, affect cell density itself via chemotaxis? Since quorum sensing often spans across species, such a feedback mechanism may also exist between multiple species. We constructed a microfluidic platform to study these questions. A flow-free, stable linear chemical gradient is formed in our device within a few minutes that makes it suitable for sensitive testing of chemoeffectors: we showed that the amino acid lysine is a weak chemoattractant for Escherichia coli, while arginine is neutral. We studied the effect of quorum sensing signal molecules of Pseudomonas aeruginosa on E. coli chemotaxis. Our results show that N-(3-oxododecanoyl)-homoserine lactone (oxo-C12-HSL) and N-(butryl)-homoserine lactone (C4-HSL) are attractants. Furthermore, we tested the chemoeffector potential of pyocyanin and pyoverdine, secondary metabolites under a quorum sensing control. Pyocyanin is proved to be a weak attractant while pyoverdine are repellent. We demonstrated the usability of the device in co-culturing experiments, where we showed that various factors released by P. aeruginosa affect the dynamic spatial rearrangement of a neighboring E. coli population, while surface adhesion of the cells is also modulated.

Quorum sensing (QS) is a communication form between bacteria via small signal molecules that enables global gene regulation as a function of cell density. We applied a microfluidic mother machine to study the kinetics of the QS response of Pseudomonas aeruginosa bacteria to additions and withdrawals of signal molecules. We traced the fast buildup and the subsequent considerably slower decay of a population-level and single-cell-level QS response. We applied a mathematical model to explain the results quantitatively. We found significant heterogeneity in QS on the single-cell level, which may result from variations in quorum-controlled gene expression and protein degradation. Heterogeneity correlates with cell lineage history, too. We used single-cell data to define and quantitatively characterize the population-level quorum state. We found that the population-level QS response is well-defined. The buildup of the quorum is fast upon signal molecule addition. At the same time, its decay is much slower following signal withdrawal, and the quorum may be maintained for several hours in the absence of the signal. Furthermore, the quorum sensing response of the population was largely repeatable in subsequent pulses of signal molecules.

Chlamydomonas reinhardtii is a model organism of increasing biotechnological importance, yet, the evaluation of its life cycle processes and photosynthesis on a single-cell level is largely unresolved. To facilitate the study of the relationship between morphology and photochemistry, we established microfluidics in combination with chlorophyll a fluorescence induction measurements. We developed two types of microfluidic platforms for single-cell investigations: (i) The traps of the “Tulip” device are suitable for capturing and immobilizing single cells, enabling the assessment of their photosynthesis for several hours without binding to a solid support surface. Using this “Tulip” platform, we performed high-quality non-photochemical quenching measurements and confirmed our earlier results on bulk cultures that non-photochemical quenching is higher in ascorbate-deficient mutants (Crvtc2-1) than in the wild-type. (ii) The traps of the “Pot” device were designed for capturing single cells and allowing the growth of the daughter cells within the traps. Using our most performant “Pot” device, we could demonstrate that the F_V/F_M parameter, an indicator of photosynthetic efficiency, varies considerably during the cell cycle. Our microfluidic devices, therefore, represent versatile platforms for the simultaneous morphological and photosynthetic investigations of C. reinhardtii on a single-cell level.

Zero-Mode Waveguides were first introduced for Fluorescence Correlation Spectroscopy at micromolar dye concentrations. We show that combining zero-mode waveguides with fluorescence correlation spectroscopy in a continuous flow mixer avoids the compression of the FCS signal due to fluid transport at channel velocities up to ~17 mm/s. We derive an analytic scaling relationship $\mid \frac{\delta k_{\mathrm{ON}}}{k_{\mathrm{ON}}}\mid =\mid \frac{\delta k_{\mathrm{OFF}}}{k_{\mathrm{OFF}}}\mid ~\frac{k_{\mathrm{ON}}+k_{\mathrm{OFF}}}{k_{\mathrm{ON}}}\frac{0.1D_B}{D_F-D_B}\frac{e}{\sqrt{\mathrm{SNR}}}$ converting this flow velocity insensitivity to improved kinetic rate certainty in time-resolved mixing experiments. Thus zero-mode waveguides make FCS suitable for direct kinetics measurements in rapid continuous flow.

Microfluidics-based protoplast isolation, which enabled intracellular uptake of singlet oxygen sensor fluorophore and DNA oligonucleotides in Symbiodinium spp.

Spatiotemporal structures and heterogeneities are common in natural habitats, yet their role in the evolution of antibiotic resistance is still to be uncovered. We applied a microfluidic gradient generator device to study the emergence of resistant bacteria in spatial ciprofloxacin gradients. We observed biofilm formation in regions with sub-inhibitory concentrations of antibiotics, which quickly expanded into the high antibiotic regions. In the absence of an explicit structure of the habitat, this multicellular formation led to a spatial structure of the population with local competition and limited migration. Therefore, such structures can function as amplifiers of selection and aid the spread of beneficial mutations. We found that the physical environment itself induces stress-related mutations that later prove beneficial when cells are exposed to antibiotics. This shift in function suggests that exaptation occurs in such experimental scenarios. The above two processes pave the way for the subsequent emergence of highly resistant specific mutations.