Microbiology seminars

To survive in the wild small rodents evolved specialized retinas. To escape predators, looming shadows need to be detected with speed and precision. To evade starvation, small seeds, grass, nuts and insects need to also be detected quickly. Some of these succulent seeds and insects may be camouflaged offering only low contrast targets.Moreover, these challenging tasks need to be accomplished continuously at dusk, night, dawn and daytime. Crossover inhibition is thought to be involved in enhancing contrast detectionin the microcircuits of the inner plexiform layer of the mammalian retina. The AII amacrine cells are narrow field cells that play a key role in crossover inhibition. Our lab studies the synaptic physiology that regulates glycine release from AII amacrine cellsin mouse retina. These interneurons receive excitation from rod and conebipolar cells and transmit excitation to ON-type bipolar cell terminals via gap junctions. They also transmit inhibition via multiple glycinergic synapses onto OFF bipolar cell terminals.AII amacrine cells are thus a central hub of synaptic information processing that cross links the ON and the OFF pathways. What are the functions of crossover inhibition? How does it enhance contrast detection at different ambient light levels? How is the dynamicrange, frequency response and synaptic gain of glycine release modulated by luminance levels and circadian rhythms? How is synaptic gain changed by different extracellular neuromodulators, like dopamine, and by intracellular messengers like cAMP, phosphateand Ca2+ ions from Ca2+ channels and Ca2+ stores? My talk will try to answer some of these questions and will pose additional ones. It will end with further hypothesis and speculations on the multiple roles of crossover inhibition.

A major unresolved question in microbiome research is whether the complex ecological patterns observed in surveys of natural communities can be explained and predicted by fundamental, quantitative principles. Bridging theory and experiment is hampered by the multiplicity of ecological processes that simultaneously affect community assembly and a lack of theoretical tools for modeling diverse ecosystems. Here, I will present a simple ecological model of microbial communities that reproduces large-scale ecological patterns observed across multiple natural and experimental settings including compositional gradients, clustering by environment, diversity/harshness correlations, and nestedness. Surprisingly, our model works despite having a “random metabolisms” and “random consumer preferences”. This raises the natural of question of why random ecosystems can describe real-world experimental data. In the second, more theoretical part of the talk, I will answer this question by showing that when a community becomes diverse enough, it will always self-organize into a stable state whose properties are well captured by a “typical random ecosystems”.

Macromolecular protein complexes perform essential biological functions across life forms. A fundamental, though yet unsolved question in biology is how the function of such complexes is regulated by intracellular or extracellular signals. For instance, we have little understanding of how forces affect multi-protein machines whose function is often mechanical in nature. We address this question by studying the bacterial flagellar motor, a large complex that powers swimming motility in many bacteria. This rotary motor autonomously adapts to changes in mechanical load by adding or removing force-generating ‘stator’ units that power rotation. In the bacterium Escherichia coli, up to 11 units drive the motor at high load while all the units are released at low load. We manipulate motor load using electrorotation, a technique in which a rapidly rotating electric field applies an external torque on the motor. This allows us to change motor load at will and measure the resulting stator dynamics at single-unit resolution. We found that the force generated by the stator units controls their unbinding, forming a feedback loop that leads to autoregulation of the assembly. We complemented our experiments with theoretical models that provide insight into the underlying molecular interactions. Torque-dependent remodeling takes place within seconds, making it a highly responsive control mechanism, one that is mediated by the mechano-chemical tuning of protein interactions.

The human gut microbiome has emerged as a key player in the bidirectional communication of the gut-brain axis, affecting various aspects of homeostasis and pathophysiology. Until recently, the majority of studies that seek to explore the mechanisms underlying the microbiome-gut-brain axis cross-talk relied almost exclusively on animal models, and particularly gnotobiotic mice. Despite the great progress made with these models, various limitations, including ethical considerations and interspecies differences that limit the translatability of data to human systems, pushed researchers to seek for alternatives. Over the past decades, the field of in vitro modelling of tissues has experienced tremendous growth, thanks to advances in 3D cell biology, materials, science and bioengineering, pushing further the borders of our ability to more faithfully emulate the in vivo situation. Organ-on-chip technology and bioengineered tissues have emerged as highly promising alternatives to animal models for a wide range of applications. In this talk I’ll discuss our progress towards generating a complete platform of the human microbiota-gut-brain axis with integrated monitoring and sensing capabilities. Bringing together principles of materials science, tissue engineering, 3D cell biology and bioelectronics, we are building advanced models of the GI and the BBB /NVU, with real-time and label-free monitoring units adapted in the model architecture, towards a robust and more physiologically relevant human in vitro model, aiming to i) elucidate the role of microbiota in the gut-brain axis communication, ii) to study how diet and impaired microbiota profiles affect various (patho-)physiologies, and iii) to test personalised medicine approaches for disease modelling and drug testing.

Biological systems can self-organize in complex structures, able to evolve and adapt to widely varying environmental conditions. Despite the importance of fluid flow for transporting and organizing populations, few laboratory systems exist to systematically investigate the impact of advection on their spatial evolutionary dynamics. In this talk, I will discuss how we can address this problem by studying the morphology and genetic spatial structure of microbial colonies growing on the surface of a viscous substrate. When grown on a liquid, I will show that S. cerevisiae (baker’s yeast) can behave like “active matter” and collectively generate a fluid flow many times larger than the unperturbed colony expansion speed, which in turn produces mechanical stresses and fragmentation of the initial colony. Combining laboratory experiments with numerical modeling, I will demonstrate that the coupling between metabolic activity and hydrodynamic flows can produce positive feedbacks and drive preferential growth phenomena leading to the formation of microbial jets. Our work provides rich opportunities to explore the interplay between hydrodynamics, growth and competition within a versatile system.

Cerebral infection by the larvae of the cestode, Taenia solium (neurocysticercosis), is thought to be the leading cause of adult-acquired epilepsy worldwide. Despite this, little is known about the cellular mechanisms that underlie seizure development in this condition. In this talk I will present our recent data exploring multiple interactions between cestode larvae, neuroinflammatory processes and network excitability. We find that viable cestode larvae are able to strongly suppress microglial activation and inflammatory cytokine release with consequences for the modulation host neuroinflammatory responses and seizure development in vivo. At the same time, larvae produce and release glutamate, with acute excitatory effects on neuronal circuits. We hope that an improved understanding of epileptogenic mechanisms in neurocysticercosis will one day improve the management of this condition as well as other inflammatory causes of epilepsy.

Individual bacteria transported in viscous flows, show complex interactions with flows and bounding surfaces resulting from their complex shape as well as their activity. Understanding these transport dynamics is crucial, as they impact soil contamination, transport in biological conducts or catheters, and constitute thus a serious health threat. Here we investigate the trajectories of individual E-coli bacteria in confined geometries under flow, using microfluidic model systems in bulk flows as well as close to surfaces using a novel Langrangian 3D tracking method. Combining experimental observations and modelling we elucidate the origin of upstream swimming, lateral drift or persistent transport along corners. [1] Junot et al, EPL, 126 (2019) 44003 [2] Mathijssen et al. 10:3 (2019) Nature Comm. [3] Figueroa-Morales et al., Soft Matter, 2015,11, 6284-6293 [4] Darnige et al. Review of Scientific Instruments 88, 055106 (2017) [5] Jing et al, Science Advances, 2020; 6 : eabb2012 [6] Figueroa-Morales et al, Sci. Adv. 2020; 6 : eaay0155, 2020, 10.1126/sciadv.aay0155

While the motion and collective behavior of cells are well-studied on flat surfaces or in unconfined liquid media, in most natural settings, cells thrive in complex 3D environments. Bioprinting processes are capable of structuring cells in 3D and conventional bioprinting approaches address this challenge by embedding cells in bio-degradable polymer networks. However, heterogeneity in network structure and biodegradation often preclude quantitative studies of cell behavior in specified 3D architectures. Here, I will present a new approach to 3D bioprinting of cellular communities that utilizes jammed, granular polyelectrolyte microgels as a support medium. The self-healing nature of this medium allows the creation of highly precise cellular communities and tissue-like structures by direct injection of cells inside the 3D medium. Further, the transparent nature of this medium enables precise characterization of cellular behavior. I will describe two examples of my work using this platform to study the behavior of two different classes of cells in 3D. First, I will describe how we interrogate the growth, viability, and migration of mammalian cells—ranging from epithelial cells, cancer cells, and T cells—in the 3D pore space. Second, I will describe how we interrogate the migration of E. coli bacteria through the 3D pore space. Direct visualization enables us to reveal a new mode of motility exhibited by individual cells, in stark contrast to the paradigm of run-and-tumble motility, in which cells are intermittently and transiently trapped as they navigate the pore space; further, analysis of these dynamics enables prediction of single-cell transport over large length and time scales. Moreover, we show that concentrated populations of E. coli can collectively migrate through a porous medium—despite being strongly confined—by chemotactically “surfing” a self-generated nutrient gradient. Together, these studies highlight how the jammed microgel medium provides a powerful platform to design and interrogate complex cellular communities in 3D—with implications for tissue engineering, microtissue mechanics, studies of cellular interactions, and biophysical studies of active matter.

Active matter systems are composed of constituents, each one in nonequilibrium, that consume energy in order to move [1]. A characteristic feature of active matter is collective motion leading to nonequilibrium phase transitions or large scale directed motion [2]. A number of recent works have featured active particles interacting with obstacles, either moving or fixed [3,4,5]. When an active particle encounters an asymmetric obstacle, different behaviours are detected depending on the nature of its active motion. On the one side, rectification effects arise in a suspension of run-and-tumble particles interacting with a wall of funnelled-shaped openings, caused by particles persistence length [6]. The same trapping mechanism could be responsible for the intake of microorganisms in the underground leaves [7] of Carnivorous plants [8]. On the other side, for aligning particles [9] interacting with a wall of funnelled-shaped openings, trapping happens on the (opposite) wider opening side of the funnels [10,11]. Interestingly, when funnels are located on a circular array, trapping is more localised and depends on the nature of the Vicsek model. Active particles can be synthetic (such as synthetic active colloids) or alive (such as living bacteria). A prototypical model to study living microswimmers is P. fluorescens, a rod shaped and biofilm forming bacterium. Biofilms are microbial communities self-assembled onto external interfaces. Biofilms can be described within the Soft Matter physics framework [12] as a viscoelastic material consisting of colloids (bacterial cells) embedded in a cross-linked polymer gel (polysaccharides cross-linked via proteins/multivalent cations), whose water content vary depending on the environmental conditions. Bacteria embedded in the polymeric matrix control biofilm structure and mechanical properties by regulating its matrix composition. We have recently monitored structural features of Pseudomonas fluorescens biofilms grown with and without hydrodynamic stress [13,14]. We have demonstrated that bacteria are capable of self-adapting to hostile hydrodynamic stress by tailoring the biofilm chemical composition, thus affecting both the mesoscale structure of the matrix and its viscoelastic properties that ultimately regulate the bacteria-polymer interactions. REFERENCES [1] C. Bechinger et al. Rev. Mod. Phys. 88, 045006 (2016); [2] T. Vicsek, A. Zafeiris Phys. Rep. 517, 71 (2012); [3] C. Bechinger, R. Di Leonardo, H. Lowen, C. Reichhardt, G. Volpe, and G. Volpe, Reviews of Modern Physics 88, 045006 (2016); [4] R Martinez, F Alarcon, DR Rodriguez, JL Aragones, C Valeriani The European Physical Journal E 41, 1 (2018); [5] DR Rodriguez, F Alarcon, R Martinez, J Ramírez, C Valeriani, Soft matter 16 (5), 1162 (2020); [6] C. O. Reichhardt and C. Reichhardt, Annual Review of Condensed Matter Physics 8, 51 (2017); [7] W Barthlott, S Porembski, E Fischer, B Gemmel Nature 392, 447 (1998); [8] C B. Giuliano, R Zhang, R.Martinez Fernandez, C.Valeriani and L.Wilson (in preparation, 2021); [9] R Martinez, F Alarcon, JL Aragones, C Valeriani Soft matter 16 (20), 4739 (2020); [10] P. Galajada, J. Keymer, P. Chaikin and R.Austin, Journal of bacteriology, 189, 8704 (2007); [11] M. Wan, C.O. Reichhardt, Z. Nussinov, and C. Reichhardt, Physical Review Letters 101, 018102 (2008); [12] J N. Wilking , T E. Angelini , A Seminara , M P. Brenner , and D A. Weitz MRS Bulletin 36, 385 (2011); [13]J Jara, F Alarcón, A K Monnappa, J Ignacio Santos, V Bianco, P Nie, M Pica Ciamarra, A Canales, L Dinis, I López-Montero, C Valeriani, B Orgaz, Frontiers in microbiology 11, 3460 (2021); [14] P Nie, F Alarcon, I López-Montero, B Orgaz, C Valeriani, M Pica Ciamarra

Three complementary lectures explain the development and applications of optogenetics. Gero Miesenböck traces optical control of neuronal activity from early experiments to studies of sleep, including neurons that respond to accumulated sleep need and molecular mechanisms governing sleep pressure. Peter Hegemann connects microbial photobiology to channelrhodopsins that control neuronal membrane voltage. He discusses their use in studying development and learning, and prospects for extending optical control to enzymes, transcription and translation. Georg Nagel follows the discovery and expression of microbial rhodopsins with light-sensitive transport and enzymatic functions. His examples include Channelrhodopsin-2 and its H134R variant, halorhodopsin, the more light-sensitive ChR2/XXL, the rhodopsin guanylyl cyclase Cyclop, and anion-channelrhodopsins in tobacco plants. Together, the talks connect molecular mechanisms with experimental control of neural and plant systems.

In 1676, using candle light and a small glass sphere as the lens, van Leeuwenhoek discovered the microscopic world of living microorganisms. Today, using lasers, spatial light modulators, digital cameras and computers, we study the statistical and fluid mechanics of microswimmers in ways that were unimaginable only 50 years ago. With light we can image swimming bacteria in 3D, apply controllable force fields or sculpt their 3D environment. In addition to shaping the physical world outside cells we can use light to control the internal state of genetically modified bacteria. I will review our recent work with light-bacteria interactions, going from some fundamental problems in the fluid and statistical mechanics of microswimmers to the use of bacteria as propellers for micro-machines or as a "living" paint controlled by light.

We are interested in understanding how microbes impact the behavior of host animals. Animal nervous systems likely evolved in environments richly surrounded by microbes, yet the impact of bacteria on nervous system function has been relatively under-studied. A challenge has been to identify systems in which both host and microbe are amenable to genetic manipulation, and which enable high-throughput behavioral screening in response to defined and naturalistic conditions. To accomplish these goals, we use an animal host — the roundworm C. elegans, which feeds on bacteria — in combination with its natural gut microbiome to identify inter-organismal signals driving host-microbe interactions and decision-making. C. elegans has some of the most extensive molecular, neurobiological and genetic tools of any multicellular eukaryote, and, coupled with the ease of gnotobiotic culture in these worms, represents a highly attractive system in which to study microbial influence on host behavior. Using this system, we discovered that commensal bacterial metabolites directly modulate nervous system function of their host. Beneficial gut microbes of the genus Providencia produce the neuromodulator tyramine in the C. elegans intestine. Using a combination of behavioral analysis, neurogenetics, metabolomics and bacterial genetics we established that bacterially produced tyramine is converted to octopamine in C. elegans, which acts directly in sensory neurons to reduce odor aversion and increase sensory preference for Providencia. We think that this type of sensory modulation may increase association of C. elegans with these microbes, increasing availability of this nutrient-rich food source for the worm and its progeny, while facilitating dispersal of the bacteria.

Motility is a crucial virulence factor for many species of bacteria, but it is not fully understood how bacterial motility is practically involved in pathogenicity. This time I will give a talk on the association of motility with pathogenicity in the zoonotic spirochete bacterium Leptospira. Recently, we measured swimming force of individual leptospires using optical tweezers and found that they can generate ~30 times of the swimming force of E. coli. We also observed that leptospires increase the reversal frequency of swimming at the gel-liquid interface, resembling host dermis exposed to contaminated water (Abe et al., 2020, Sci Rep). These could be involved in percutaneous infection of the spirochete. We have shown that Leptospira not only swims in liquid but also moves over solid surfaces (Tahara et al., 2018, Sci Adv). We quantified the surface motility called “crawling” on cultured kidney tissues from various mammals, showing that pathogenic leptospires crawl over the tissue surfaces more persistently that non-pathogenic ones (Xu et al., 2020, Front Microbiol). I will discuss the spirochete motility related to pathogenicity from the biophysical viewpoint.