Cellular arrangement impacts metabolic activity and antibiotic tolerance in Pseudomonas aeruginosa biofilms.

Cells must access resources to survive, and the anatomy of multicellular structures influences this access. In diverse multicellular eukaryotes, resources are provided by internal conduits that allow substances to travel more readily through tissue than they would via diffusion. Microbes growing in multicellular structures, called biofilms, are also affected by differential access to resources and we hypothesized that this is influenced by the physical arrangement of the cells. In this study, we examined the microanatomy of biofilms formed by the pathogenic bacterium Pseudomonas aeruginosa and discovered that clonal cells form striations that are packed lengthwise across most of a mature biofilm's depth. We identified mutants, including those defective in pilus function and in O-antigen attachment, that show alterations to this lengthwise packing phenotype. Consistent with the notion that cellular arrangement affects access to resources within the biofilm, we found that while the wild type shows even distribution of tested substrates across depth, the mutants show accumulation of substrates at the biofilm boundaries. Furthermore, we found that altered cellular arrangement within biofilms affects the localization of metabolic activity, the survival of resident cells, and the susceptibility of subpopulations to antibiotic treatment. Our observations provide insight into cellular features that determine biofilm microanatomy, with consequences for physiological differentiation and drug sensitivity.

Tardigrades exhibit robust interlimb coordination across walking speeds and terrains.

Tardigrades must negotiate heterogeneous, fluctuating environments and accordingly utilize locomotive strategies capable of dealing with variable terrain. We analyze the kinematics and interleg coordination of freely walking tardigrades (species: Hypsibius exemplaris). We find that tardigrade walking replicates several key features of walking in insects despite disparities in size, skeleton, and habitat. To test the effect of environmental changes on tardigrade locomotor control circuits we measure kinematics and interleg coordination during walking on two substrates of different stiffnesses. We find that the phase offset between contralateral leg pairs is flexible, while ipsilateral coordination is preserved across environmental conditions. This mirrors similar results in insects and crustaceans. We propose that these functional similarities in walking coordination between tardigrades and arthropods is either due to a generalized locomotor control circuit common to panarthropods or to independent convergence onto an optimal strategy for robust multilegged control in small animals with simple circuitry. Our results highlight the value of tardigrades as a comparative system toward understanding the mechanisms-neural and/or mechanical-underlying coordination in panarthropod locomotion.

Flexible locomotion in complex environments: the influence of species, speed and sensory feedback on panarthropod inter-leg coordination.

Panarthropods (a clade containing arthropods, tardigrades and onychophorans) can adeptly move across a wide range of challenging terrains and their ability to do so given their relatively simple nervous systems makes them compelling study organisms. Studies of forward walking on flat terrain excitingly point to key features in inter-leg coordination patterns that seem to be 'universally' shared across panarthropods. However, when movement through more complex, naturalistic terrain is considered, variability in coordination patterns - from the intra-individual to inter-species level - becomes more apparent. This variability is likely to be due to the interplay between sensory feedback and local pattern-generating activity, and depends crucially on species, walking speed and behavioral goal. Here, I gather data from the literature of panarthropod walking coordination on both flat ground and across more complex terrain. This Review aims to emphasize the value of: (1) designing experiments with an eye towards studying organisms in natural environments; (2) thoughtfully integrating results from various experimental techniques, such as neurophysiological and biomechanical studies; and (3) ensuring that data is collected and made available from a wider range of species for future comparative analyses.

Catch bond drives stator mechanosensitivity in the bacterial flagellar motor.

The bacterial flagellar motor (BFM) is the rotary motor that rotates each bacterial flagellum, powering the swimming and swarming of many motile bacteria. The torque is provided by stator units, ion motive force-powered ion channels known to assemble and disassemble dynamically in the BFM. This turnover is mechanosensitive, with the number of engaged units dependent on the viscous load experienced by the motor through the flagellum. However, the molecular mechanism driving BFM mechanosensitivity is unknown. Here, we directly measure the kinetics of arrival and departure of the stator units in individual motors via analysis of high-resolution recordings of motor speed, while dynamically varying the load on the motor via external magnetic torque. The kinetic rates obtained, robust with respect to the details of the applied adsorption model, indicate that the lifetime of an assembled stator unit increases when a higher force is applied to its anchoring point in the cell wall. This provides strong evidence that a catch bond (a bond strengthened instead of weakened by force) drives mechanosensitivity of the flagellar motor complex. These results add the BFM to a short, but growing, list of systems demonstrating catch bonds, suggesting that this "molecular strategy" is a widespread mechanism to sense and respond to mechanical stress. We propose that force-enhanced stator adhesion allows the cell to adapt to a heterogeneous environmental viscosity and may ultimately play a role in surface-sensing during swarming and biofilm formation.

Mechanics of torque generation in the bacterial flagellar motor.

The bacterial flagellar motor (BFM) is responsible for driving bacterial locomotion and chemotaxis, fundamental processes in pathogenesis and biofilm formation. In the BFM, torque is generated at the interface between transmembrane proteins (stators) and a rotor. It is well established that the passage of ions down a transmembrane gradient through the stator complex provides the energy for torque generation. However, the physics involved in this energy conversion remain poorly understood. Here we propose a mechanically specific model for torque generation in the BFM. In particular, we identify roles for two fundamental forces involved in torque generation: electrostatic and steric. We propose that electrostatic forces serve to position the stator, whereas steric forces comprise the actual "power stroke." Specifically, we propose that ion-induced conformational changes about a proline "hinge" residue in a stator α-helix are directly responsible for generating the power stroke. Our model predictions fit well with recent experiments on a single-stator motor. The proposed model provides a mechanical explanation for several fundamental properties of the flagellar motor, including torque-speed and speed-ion motive force relationships, backstepping, variation in step sizes, and the effects of key mutations in the stator.

The Limiting Speed of the Bacterial Flagellar Motor.

Recent experiments on the bacterial flagellar motor have shown that the structure of this nanomachine, which drives locomotion in a wide range of bacterial species, is more dynamic than previously believed. Specifically, the number of active torque-generating complexes (stators) was shown to vary across applied loads. This finding brings under scrutiny the experimental evidence reporting that limiting (zero-torque) speed is independent of the number of active stators. In this study, we propose that, contrary to previous assumptions, the maximum speed of the motor increases as additional stators are recruited. This result arises from our assumption that stators disengage from the motor for a significant portion of their mechanochemical cycles at low loads. We show that this assumption is consistent with current experimental evidence in chimeric motors, as well as with the requirement that a processive motor driving a large load via an elastic linkage must have a high duty ratio.

diCal-IBD: demography-aware inference of identity-by-descent tracts in unrelated individuals.

UNLABELLED: We present a tool, diCal-IBD, for detecting identity-by-descent (IBD) tracts between pairs of genomic sequences. Our method builds on a recent demographic inference method based on the coalescent with recombination, and is able to incorporate demographic information as a prior. Simulation study shows that diCal-IBD has significantly higher recall and precision than that of existing single-nucleotide polymorphism-based IBD detection methods, while retaining reasonable accuracy for IBD tracts as small as 0.1 cM. AVAILABILITY: http://sourceforge.net/projects/dical-ibd.

Development of spatial coarse-to-fine processing in the visual pathway.

The sequential analysis of information in a coarse-to-fine manner is a fundamental mode of processing in the visual pathway. Spatial frequency (SF) tuning, arguably the most fundamental feature of spatial vision, provides particular intuition within the coarse-to-fine framework: low spatial frequencies convey global information about an image (e.g., general orientation), while high spatial frequencies carry more detailed information (e.g., edges). In this paper, we study the development of cortical spatial frequency tuning. As feedforward input from the lateral geniculate nucleus (LGN) has been shown to have significant influence on cortical coarse-to-fine processing, we present a firing-rate based thalamocortical model which includes both feedforward and feedback components. We analyze the relationship between various model parameters (including cortical feedback strength) and responses. We confirm the importance of the antagonistic relationship between the center and surround responses in thalamic relay cell receptive fields (RFs), and further characterize how specific structural LGN RF parameters affect cortical coarse-to-fine processing. Our results also indicate that the effect of cortical feedback on spatial frequency tuning is age-dependent: in particular, cortical feedback more strongly affects coarse-to-fine processing in kittens than in adults. We use our results to propose an experimentally testable hypothesis for the function of the extensive feedback in the corticothalamic circuit.

Kinematics and coordination of moth flies walking on smooth and rough surfaces.

The moth fly, Clogmia albipunctata, is a common synanthropic insect with a worldwide range that lives in nearly any area with moist, decaying organic matter. These habitats comprise both smooth, slippery substrates (e.g., bathroom drains) and heterogeneous, bumpy ground (e.g., soil in plant pots). By using terrain of varying levels of roughness, we focus specifically on how substrate roughness at the approximate size scale of the organism affects kinematics and coordination in adult moth flies. Finally, we compare and contrast our characterizations of locomotion in C. albipunctata with previous work of insect walking in naturalistic environments.

Evolutionarily diverse fungal zoospores show contrasting swimming patterns specific to ultrastructure.

Zoosporic fungi, also called chytrids, produce single-celled motile spores with flagellar swimming tails (zoospores).1,2 These fungi are key components of aquatic food webs, acting as pathogens, saprotrophs, and prey.3,4,5,6,7,8 Little is known about the swimming behavior of fungal zoospores, a crucial factor governing dispersal, biogeographical range, ecological function, and infection dynamics.6,9 Here, we track the swimming patterns of zoospores from 12 evolutionarily divergent species of zoosporic fungi from across seven orders of the Chytridiomycota and the Blastocladiomycota. We report two major swimming patterns that correlate with the cytoskeletal ultrastructure of these zoospores. Specifically, we show that species without major cytoplasmic tubulin components swim in a circular fashion, while species with prominent cytoplasmic tubulin structures swim in a pattern akin to a random walk (move-stop-redirect-move). We confirm cytoskeletal architecture by performing fluorescence confocal microscopy across all 12 species. We then treat representative species with variant swimming behaviors and cytoplasmic-cytoskeletal arrangements with tubulin-stabilizing (Taxol) and depolymerizing (nocodazole) pharmacological compounds. We observed that when treating the "random walk" species with nocodazole, their swimming behavior changed to a circular-swimming pattern. Confocal imaging of the nocodazole-treated zoospores demonstrates that these cells maintain flagellum tubulin structures but lack their characteristic cytoplasmic tubulin structures. Our data demonstrate that the capability of zoospores to perform "complex" random-walk movement is linked to the presence of prominent cytoplasmic tubulin structures and suggest a link between cytology, sensory systems, and swimming behavior in a diversity of zoosporic fungi.

Analysis of unstable behavior in a mathematical model for erythropoiesis.

We consider an age-structured model that describes the regulation of erythropoiesis through the negative feedback loop between erythropoietin and hemoglobin. This model is reduced to a system of two ordinary differential equations with two constant delays for which we show existence of a unique steady state. We determine all instances at which this steady state loses stability via a Hopf bifurcation through a theoretical bifurcation analysis establishing analytical expressions for the scenarios in which they arise. We show examples of supercritical Hopf bifurcations for parameter values estimated according to physiological values for humans found in the literature and present numerical simulations in agreement with the theoretical analysis. We provide a strategy for parameter estimation to match empirical measurements and predict dynamics in experimental settings, and compare existing data on hemoglobin oscillation in rabbits with predictions of our model.

Cell arrangement impacts metabolic activity and antibiotic tolerance in Pseudomonas aeruginosa biofilms.

Cells must access resources to survive, and the anatomy of multicellular structures influences this access. In diverse multicellular eukaryotes, resources are provided by internal conduits that allow substances to travel more readily through tissue than they would via diffusion. Microbes growing in multicellular structures, called biofilms, are also affected by differential access to resources and we hypothesized that this is influenced by the physical arrangement of the cells. In this study, we examined the microanatomy of biofilms formed by the pathogenic bacterium Pseudomonas aeruginosa and discovered that clonal cells form striations that are packed lengthwise across most of a mature biofilm's depth. We identified mutants, including those defective in pilus function and in O-antigen attachment, that show alterations to this lengthwise packing phenotype. Consistent with the notion that cellular arrangement affects access to resources within the biofilm, we found that while the wild type shows even distribution of tested substrates across depth, the mutants show accumulation of substrates at the biofilm boundaries. Furthermore, we found that altered cellular arrangement within biofilms affects the localization of metabolic activity, the survival of resident cells, and the susceptibility of subpopulations to antibiotic treatment. Our observations provide insight into cellular features that determine biofilm microanatomy, with consequences for physiological differentiation and drug sensitivity.

Universal Features in Panarthropod Inter-Limb Coordination during Forward Walking.

Terrestrial animals must often negotiate heterogeneous, varying environments. Accordingly, their locomotive strategies must adapt to a wide range of terrain, as well as to a range of speeds to accomplish different behavioral goals. Studies in Drosophila have found that inter-leg coordination patterns (ICPs) vary smoothly with walking speed, rather than switching between distinct gaits as in vertebrates (e.g., horses transitioning between trotting and galloping). Such a continuum of stepping patterns implies that separate neural controllers are not necessary for each observed ICP. Furthermore, the spectrum of Drosophila stepping patterns includes all canonical coordination patterns observed during forward walking in insects. This raises the exciting possibility that the controller in Drosophila is common to all insects, and perhaps more generally to panarthropod walkers. Here, we survey and collate data on leg kinematics and inter-leg coordination relationships during forward walking in a range of arthropod species, as well as include data from a recent behavioral investigation into the tardigrade Hypsibius exemplaris. Using this comparative dataset, we point to several functional and morphological features that are shared among panarthropods. The goal of the framework presented in this review is to emphasize the importance of comparative functional and morphological analyses in understanding the origins and diversification of walking in Panarthropoda. Introduction.

ATP synthase: Evolution, energetics, and membrane interactions.

The synthesis of ATP, life's "universal energy currency," is the most prevalent chemical reaction in biological systems and is responsible for fueling nearly all cellular processes, from nerve impulse propagation to DNA synthesis. ATP synthases, the family of enzymes that carry out this endless task, are nearly as ubiquitous as the energy-laden molecule they are responsible for making. The F-type ATP synthase (F-ATPase) is found in every domain of life and has facilitated the survival of organisms in a wide range of habitats, ranging from the deep-sea thermal vents to the human intestine. Accordingly, there has been a large amount of work dedicated toward understanding the structural and functional details of ATP synthases in a wide range of species. Less attention, however, has been paid toward integrating these advances in ATP synthase molecular biology within the context of its evolutionary history. In this review, we present an overview of several structural and functional features of the F-type ATPases that vary across taxa and are purported to be adaptive or otherwise evolutionarily significant: ion channel selectivity, rotor ring size and stoichiometry, ATPase dimeric structure and localization in the mitochondrial inner membrane, and interactions with membrane lipids. We emphasize the importance of studying these features within the context of the enzyme's particular lipid environment. Just as the interactions between an organism and its physical environment shape its evolutionary trajectory, ATPases are impacted by the membranes within which they reside. We argue that a comprehensive understanding of the structure, function, and evolution of membrane proteins-including ATP synthase-requires such an integrative approach.

Load-dependent adaptation near zero load in the bacterial flagellar motor.

The bacterial flagellar motor is an ion-powered transmembrane protein complex which drives swimming in many bacterial species. The motor consists of a cytoplasmic 'rotor' ring and a number of 'stator' units, which are bound to the cell wall of the bacterium. Recently, it has been shown that the number of functional torque-generating stator units in the motor depends on the external load, and suggested that mechanosensing in the flagellar motor is driven via a 'catch bond' mechanism in the motor's stator units. We present a method that allows us to measure-on a single motor-stator unit dynamics across a large range of external loads, including near the zero-torque limit. By attaching superparamagnetic beads to the flagellar hook, we can control the motor's speed via a rotating magnetic field. We manipulate the motor to four different speed levels in two different ion-motive force (IMF) conditions. This framework allows for a deeper exploration into the mechanism behind load-dependent remodelling by separating out motor properties, such as rotation speed and energy availability in the form of IMF, that affect the motor torque.

Biophysics at the coffee shop: lessons learned working with George Oster.

Over the past 50 years, the use of mathematical models, derived from physical reasoning, to describe molecular and cellular systems has evolved from an art of the few to a cornerstone of biological inquiry. George Oster stood out as a pioneer of this paradigm shift from descriptive to quantitative biology not only through his numerous research accomplishments, but also through the many students and postdocs he mentored over his long career. Those of us fortunate enough to have worked with George agree that his sharp intellect, physical intuition, and passion for scientific inquiry not only inspired us as scientists but also greatly influenced the way we conduct research. We would like to share a few important lessons we learned from George in honor of his memory and with the hope that they may inspire future generations of scientists.

Stability analysis in spatial modeling of cell signaling.

Advances in high-resolution microscopy and other techniques have emphasized the spatio-temporal nature of information transfer through signal transduction pathways. The compartmentalization of signaling molecules and the existence of microdomains are now widely acknowledged as key features in biochemical signaling. To complement experimental observations of spatio-temporal dynamics, mathematical modeling has emerged as a powerful tool. Using modeling, one can not only recapitulate experimentally observed dynamics of signaling molecules, but also gain an understanding of the underlying mechanisms in order to generate experimentally testable predictions. Reaction-diffusion systems are commonly used to this end; however, the analysis of coupled nonlinear systems of partial differential equations, generated by considering large reaction networks is often challenging. Here, we aim to provide an introductory tutorial for the application of reaction-diffusion models to the spatio-temporal dynamics of signaling pathways. In particular, we outline the steps for stability analysis of such models, with a focus on biochemical signal transduction. WIREs Syst Biol Med 2018, 10:e1395. doi: 10.1002/wsbm.1395 This article is categorized under: Biological Mechanisms > Cell Signaling Analytical and Computational Methods > Dynamical Methods Models of Systems Properties and Processes > Mechanistic Models.

Geckos Race Across the Water's Surface Using Multiple Mechanisms.

Acrobatic geckos can sprint at high speeds over challenging terrain [1], scamper up the smoothest surfaces [2], rapidly swing underneath leaves [3], and right themselves in midair by swinging only their tails [4, 5]. From our field observations, we can add racing on the water's surface to the gecko's list of agile feats. Locomotion at the air-water interface evolved in over a thousand species, including insects, fish, reptiles, and mammals [6]. To support their weight, some larger-legged vertebrates use forces generated by vigorous slapping of the fluid's surface followed by a stroke of their appendage [7-12], whereas smaller animals, like arthropods, rely on surface tension to walk on water [6, 13]. Intermediate-sized geckos (Hemidactylus platyurus) fall squarely between these two regimes. Here, we report the unique ability of geckos to exceed the speed limits of conventional surface swimming. Several mechanisms likely contribute in this intermediate regime. In contrast to bipedal basilisk lizards [7-10], geckos used a stereotypic trotting gait with all four limbs, creating air cavities during slapping to raise their head and anterior trunk above water. Adding surfactant to the water decreased velocity by half, confirming surface tension's role. The superhydrophobic skin could reduce drag during semi-planing. Geckos laterally undulated their bodies, including their submerged posterior trunk and tail, generating thrust for forward propulsion, much like water dragons [14] and alligators [15]. Geckos again remind us of the advantages of multi-functional morphologies providing the opportunity for multiple mechanisms for motion.

Spatial distribution of intracortical porosity varies across age and sex.

Cortical bone porosity is a major determinant of strength, stiffness, and fracture toughness of cortical tissue. The goal of this work was to investigate changes in spatial distribution and microstructure of cortical porosity associated with aging in men and women. The specific aims were to: 1) develop an automated technique for spatial analysis of cortical microstructure based on HR-pQCT data, and; 2) apply this technique to explore sex- and age-specific spatial distribution and microstructure of porosity within the cortex. We evaluated HR-pQCT images of the distal tibia from a cross-sectional cohort of 145 individuals, characterizing detectable pores as being in the endosteal, midcortical, or periosteal layers of the cortex. Metrics describing porosity, pore number, and pore size were quantified within each layer and compared across sexes, age groups, and cortical layers. The elderly cohort (65-78 years, n=22) displayed higher values than the young cohort (20-29 years, n=29) for all parameters both globally and within each layer. While all three layers displayed significant age-related porosity increases, the greatest difference in porosity between the young and elderly cohort was in the midcortical layer (+344%, p<0.001). Similarly, the midcortical layer reflected the greatest differences between young and elderly cohorts in both pore number (+243%, p<0.001) and size (+28%, p<0.001). Females displayed greater age-related changes in porosity and pore number than males. Females and males displayed comparable small to non-significant changes with age in pore size. In summary, considerable variability exists in the spatial distribution of detectable cortical porosity at the distal tibia, and this variability is dependent on age and sex. Intracortical pore distribution analysis may ultimately provide insight into both mechanisms of pore network expansion and biomechanical consequences of pore distribution.

Exploiting pallidal plasticity for stimulation in Parkinson's disease.

OBJECTIVE: Continuous application of high-frequency deep brain stimulation (DBS) often effectively reduces motor symptoms of Parkinson's disease patients. While there is a growing need for more effective and less traumatic stimulation, the exact mechanism of DBS is still unknown. Here, we present a methodology to exploit the plasticity of GABAergic synapses inside the external globus pallidus (GPe) for the optimization of DBS. APPROACH: Assuming the existence of spike-timing-dependent plasticity (STDP) at GABAergic GPe-GPe synapses, we simulate neural activity in a network model of the subthalamic nucleus and GPe. In particular, we test different DBS protocols in our model and quantify their influence on neural synchrony. MAIN RESULTS: In an exemplary set of biologically plausible model parameters, we show that STDP in the GPe has a direct influence on neural activity and especially the stability of firing patterns. STDP stabilizes both uncorrelated firing in the healthy state and correlated firing in the parkinsonian state. Alternative stimulation protocols such as coordinated reset stimulation can clearly profit from the stabilizing effect of STDP. These results are widely independent of the STDP learning rule. SIGNIFICANCE: Once the model settings, e.g., connection architectures, have been described experimentally, our model can be adjusted and directly applied in the development of novel stimulation protocols. More efficient stimulation leads to both minimization of side effects and savings in battery power.