Organoids and organ-on-chip technology for investigating host-microorganism interactions.

Recent advances in organoid and organ-on-chip (OoC) technologies offer an unprecedented level of tissue mimicry. These models can recapitulate the diversity of cellular composition, 3D organization, and mechanical stimulation. These approaches are intensively used to understand complex diseases. This review focuses on the latest advances in this field to study host-microorganism interactions.

Aggregation Temperature of Escherichia coli Depends on Steepness of the Thermal Gradient.

Bacterial chemotaxis, the directed migration of bacteria in a gradient of chemoattractant, is one of the most well-studied and well-understood processes in cell biology. On the other hand, bacterial thermotaxis, the directed migration of bacteria in a gradient of temperature, is understood relatively poorly, with somewhat conflicting reports by different groups. One of the reasons for that is the relative technical difficulty of the generation of well-defined gradients of temperature that are sufficiently steep to elicit readily detectable thermotaxis. Here, we used a specially designed microfluidic device to study thermotaxis of Escherichia coli in a broad range of thermal gradients with a high rate of data collection. We found that in shallow temperature gradients with narrow temperature ranges, E. coli tended to aggregate near a sidewall of the gradient channel at either the lowest or the highest temperature. On the other hand, in sufficiently steep gradients with wide temperature ranges, E. coli aggregated at intermediate temperatures, with maximal cell concentrations found away from the sidewalls. We observed this intermediate temperature aggregation in a motility buffer that did not contain any major chemoattractants of E. coli, in contradiction to some previous reports, which suggested that this type of aggregation required the presence of at least one major chemoattractant in the medium. Even more surprisingly, the aggregation temperature strongly depended on the gradient steepness, decreasing by ∼10° as the steepness was increased from 27 to 53°C/mm. Our experiments also highlight the fact that assessments of thermal gradients by changes in fluorescence of temperature-sensitive fluorescent dyes need to account for thermophoresis of the dyes.

Chemotaxis as a navigation strategy to boost range expansion.

Bacterial chemotaxis, the directed movement of cells along gradients of chemoattractants, is among the best-characterized subjects in molecular biology1-10, but much less is known about its physiological roles11. It is commonly seen as a starvation response when nutrients run out, or as an escape response from harmful situations12-16. Here we identify an alternative role of chemotaxis by systematically examining the spatiotemporal dynamics of Escherichia coli in soft agar12,17,18. Chemotaxis in nutrient-replete conditions promotes the expansion of bacterial populations into unoccupied territories well before nutrients run out in the current environment. Low levels of chemoattractants act as aroma-like cues in this process, establishing the direction and enhancing the speed of population movement along the self-generated attractant gradients. This process of navigated range expansion spreads faster and yields larger population gains than unguided expansion following the canonical Fisher-Kolmogorov dynamics19,20 and is therefore a general strategy to promote population growth in spatially extended, nutrient-replete environments.

Glider soaring via reinforcement learning in the field.

Soaring birds often rely on ascending thermal plumes (thermals) in the atmosphere as they search for prey or migrate across large distances1-4. The landscape of convective currents is rugged and shifts on timescales of a few minutes as thermals constantly form, disintegrate or are transported away by the wind5,6. How soaring birds find and navigate thermals within this complex landscape is unknown. Reinforcement learning7 provides an appropriate framework in which to identify an effective navigational strategy as a sequence of decisions made in response to environmental cues. Here we use reinforcement learning to train a glider in the field to navigate atmospheric thermals autonomously. We equipped a glider of two-metre wingspan with a flight controller that precisely controlled the bank angle and pitch, modulating these at intervals with the aim of gaining as much lift as possible. A navigational strategy was determined solely from the glider's pooled experiences, collected over several days in the field. The strategy relies on on-board methods to accurately estimate the local vertical wind accelerations and the roll-wise torques on the glider, which serve as navigational cues. We establish the validity of our learned flight policy through field experiments, numerical simulations and estimates of the noise in measurements caused by atmospheric turbulence. Our results highlight the role of vertical wind accelerations and roll-wise torques as effective mechanosensory cues for soaring birds and provide a navigational strategy that is directly applicable to the development of autonomous soaring vehicles.

Exploring the function of bacterial chemotaxis.

Bacterial chemotaxis is a classical subject: our knowledge of its molecular pathway has grown very detailed, and experimental observations, as well as mathematical models of the dynamics of chemotactic populations, have a history of several decades. This should not lead to the conclusion that only minor details are left to be understood. Indeed, it is believed that bacterial chemotaxis is under selection for efficiency, yet the underlying functional forces remain largely unknown. These aspects are discussed here by the presentation of illustrative examples related to the role of adaptation and signal integration. Both are expected to be important in ecologically relevant conditions, where chemotaxis should be strongly coupled with metabolism and growth, due to the presence of diverse chemoattractant cues and their active consumption by multiple types of bacteria competing for growth.

The Role of Adaptation in Bacterial Speed Races.

Evolution of biological sensory systems is driven by the need for efficient responses to environmental stimuli. A paradigm among prokaryotes is the chemotaxis system, which allows bacteria to navigate gradients of chemoattractants by biasing their run-and-tumble motion. A notable feature of chemotaxis is adaptation: after the application of a step stimulus, the bacterial running time relaxes to its pre-stimulus level. The response to the amino acid aspartate is precisely adapted whilst the response to serine is not, in spite of the same pathway processing the signals preferentially sensed by the two receptors Tar and Tsr, respectively. While the chemotaxis pathway in E. coli is well characterized, the role of adaptation, its functional significance and the ecological conditions where chemotaxis is selected, are largely unknown. Here, we investigate the role of adaptation in the climbing of gradients by E. coli. We first present theoretical arguments that highlight the mechanisms that control the efficiency of the chemotactic up-gradient motion. We discuss then the limitations of linear response theory, which motivate our subsequent experimental investigation of E. coli speed races in gradients of aspartate, serine and combinations thereof. By using microfluidic techniques, we engineer controlled gradients and demonstrate that bacterial fronts progress faster in equal-magnitude gradients of serine than aspartate. The effect is observed over an extended range of concentrations and is not due to differences in swimming velocities. We then show that adding a constant background of serine to gradients of aspartate breaks the adaptation to aspartate, which results in a sped-up progression of the fronts and directly illustrate the role of adaptation in chemotactic gradient-climbing.

Gene inactivation of a chemotaxis operon in the pathogen Leptospira interrogans.

Chemotaxis may have an important role in the infection process of pathogenic Leptospira spp.; however, little is known about the regulation of flagellar-based motility in these atypical bacteria. We generated a library of random transposon mutants of the pathogen L. interrogans, which included a mutant with insertion in the first gene of an operon containing the chemotaxis genes cheA, cheW, cheD, cheB, cheY and mcp. The disrupted gene encodes a putative histidine kinase (HK). The HK mutant was motile and virulent, but swarm plate and capillary assays suggested that chemotaxis was reduced in this mutant. Further analysis of bacterial trajectories by videomicroscopy showed that the ability of this mutant to reverse was significantly impaired in comparison to wild-type strain. Our data therefore show that this operon is required for full chemotaxis of Leptospira spp.

Noninvasive inference of the molecular chemotactic response using bacterial trajectories.

The quality of sensing and response to external stimuli constitutes a basic element in the selective performance of living organisms. Here we consider the response of Escherichia coli to chemical stimuli. For moderate amplitudes, the bacterial response to generic profiles of sensed chemicals is reconstructed from its response function to an impulse, which then controls the efficiency of bacterial motility. We introduce a method for measuring the impulse response function based on coupling microfluidic experiments and inference methods: The response function is inferred using Bayesian methods from the observed trajectories of bacteria swimming in microfluidically controlled chemical fields. The notable advantages are that the method is based on the bacterial swimming response, it is noninvasive, without any genetic and/or mechanical preparation, and assays the behavior of the whole flagella bundle. We exploit the inference method to measure responses to aspartate and α-methylaspartate--measured previously by other methods--as well as glucose, leucine, and serine. The response to the attractant glucose is shown to be biphasic and perfectly adapted, as for aspartate. The response to the attractant serine is shown to be biphasic yet imperfectly adapted, that is, the response function has a nonzero (positive) integral. The adaptation of the response to the repellent leucine is also imperfect, with the sign of the two phases inverted with respect to serine. The diversity in the bacterial population of the response function and its dependency upon the background concentration are quantified.

Plasmid copy number noise in monoclonal populations of bacteria.

Plasmids are extra chromosomal DNA that can confer to their hosts' supplementary characteristics such as antibiotic resistance. Plasmids code for their copy number through their own replication frequency. Even though the biochemical networks underlying the plasmid copy number (PCN) regulation processes have been studied and modeled, no measurement of the heterogeneity in PCN within a whole population has been done. We have developed a fluorescent-based measurement system, which enables determination of the mean and noise in PCN within a monoclonal population of bacteria. Two different fluorescent protein reporters were inserted: one on the chromosome and the other on the plasmid. The fluorescence of these bacteria was measured with a microfluidic flow cytometry device. We show that our measurements are consistent with known plasmid characteristics. We find that the partitioning system lowers the PCN mean and standard deviation. Finally, bacterial populations were allowed to grow without selective pressure. In this case, we were able to determine the plasmid loss rate and growth inhibition effect.

Inference of plasmid-copy-number mean and noise from single-cell gene expression data.

Plasmids are extrachromosomal DNA molecules which code for their own replication. We previously reported a setup using genes coding for fluorescent proteins of two colors that allowed us, using a simple model, to extract the plasmid-copy-number noise in a monoclonal population of bacteria [J. Wong Ng, Phys. Rev. E 81, 011909 (2010)]. Here we present a detailed calculation relating this noise to the measured levels of fluorescence, taking into account all sources of fluorescence fluctuations: not only the fluctuation of gene expression as in the simple model but also the growth and division of bacteria, the nonuniform distribution of their ages, the random partition of proteins at divisions, and the replication and partition of plasmids and chromosome. We show how to use the chromosome as a reference, which helps extracting the plasmid-copy-number noise in a self-consistent manner.