Biology subjects

Thomas, S. A.

Publications and source records attributed to Thomas, S. A..

At least 19 recordsLinked to original sources

bioRxiv2026

Upper respiratory tract bacterial microbiome features associated with Streptococcus pneumoniae colonisation, environmental and demographic factors among older children in a rural setting in The Gambia

Abstract The upper respiratory tract (URT) microbiome plays a vital role in respiratory health, with older children (5-14 years old) serving as key reservoirs for the transmission of bacteria to younger individuals (< 5 years old). However, the URT microbiome in this age group remains poorly characterised, particularly in relation to colonisation by the pathobiont Streptococcus pneumoniae. To address this gap, we analysed URT samples collected over eight visits from 134 healthy children aged 5-14 years in The Gambia, using culture-dependent methods to detect S. pneumoniae and culture-independent 16S rRNA sequencing to characterise the microbiome in parallel. 16S rRNA sequencing identified Streptococcus and Prevotella as the dominant microbiome constituents. When integrating longitudinal sampling data, we observed temporal instability with frequent switches in microbiome profiles over time. URT samples colonised with S. pneumoniae were associated with increased abundance of Moraxella, Streptococcus and Haemophilus, whereas non-colonised URT samples were enriched with Granulicatella and Filifactor. S. pneumoniae colonisation was also associated with lower Shannon diversity compared with non-colonised samples. Age and season were the strongest determinants of microbiome composition; increasing age was associated with decreasing prevalence and abundance of taxa including Haemophilus and Moraxella, while gram-positive taxa were predominantly enriched in the dry season. These findings provide new insight into the URT microbiome of older children in The Gambia and highlight Granulicatella and Filifactor as potential contributors to colonisation resistance against S. pneumoniae.

microbiology↗

bioRxiv2026

A Novel Conditional Adra2a-Knockout Mouse Line Reveals Cell-specific Contributions to Specific Dimensions of Sedation

IntroductionHere, we create a conditional Adra2a line and use it to show that sedative, hypnotic, and hypothermic effects of 2-agonists are neuronally mediated via the 2A adrenergic receptor. MethodsWe generated mice with loxP sites flanking Adra2a using CRISPR/Cas9 gene targeting. This line was crossed with lines encoding Cre recombinase (Cre) under the control of the Vgat, Snap25, and Dbh promoters. Cell-specific knockout was confirmed using fluorescent in-situ hybridization demonstrating targeted reduction in Adra2a mRNA in the appropriate cell types. Mice were given intraperitoneal dexmedetomidine (0.3 or 1 mg/kg) or saline, and 20 minutes later righting reflex was assessed, followed by 3 rounds of rotarod testing, with fall time and end temperature recorded. Spontaneous activity was recorded using beam break for an hour after. Mice of each genotype were implanted with EEG leads and recorded while given 0.3 mg/kg IP dexmedetomidine. ResultsWe created a conditional knockout and demonstrated cell-type specific reduction of Adra2a mRNA in crossed lines with cell-specific Cre. The pan-neuronal Adra2a knockout showed resistance to all temperature, sedative, and hypnotic effect endpoints in response to the 2-agonist dexmedetomidine. Adrenergic knockout demonstrated resistance to 2-agonist hypnosis and moderate resistance to hypothermia and impaired coordination with forced movement. GABAergic knockout showed resistance only to impairment of spontaneous movement by 2-agonists. Spectral analysis of the EEG showed an increase in proportion of delta power with a sedative dose of dexmedetomidine in all lines except the pan-neuronal Adra2a knockout. DiscussionFuture studies will pursue both the specific subtype(s) and location of neuronal populations responsible for sedative, hypnotic, and hypothermic effects of 2-agonists.

neuroscience↗

bioRxiv2026

Bile acid dependent attenuation of toxin mediated disease is independent of colonization resistance against C. difficile

Clostridioides difficile infection (CDI) is a severe antibiotic associated disease and a major cause of morbidity and mortality worldwide. CDI is thought to arise from the loss of protective gut microbes that mediate functions such as secondary bile acid metabolism and nutrient competition, yet the relative contributions of these mechanisms remain unclear. To determine how these processes influence C. difficile growth, virulence, and disease, we performed in vitro and in vivo experiments using two Clostridia strains previously associated with colonization resistance against C. difficile. Neither organism prevented colonization or growth through nutrient competition alone. In contrast, secondary bile acid metabolism significantly reduced toxin-mediated disease in vivo in a strain dependent manner. These findings demonstrate that secondary bile acid modulation is an important component of CDI prevention independent of nutrient competition and suggest that attenuating virulence, in addition to limiting colonization, may represent a key strategy for next-generation CDI therapeutics.

microbiology↗

bioRxiv2025

Unique Unbiased Median Solution for Even Sample Sizes

Data in experimental biology are frequently marred by outliers and asymmetric distributions. The median, being a robust estimator of central tendency, is less sensitive to outliers than the mean. However, for ranked datasets with an even number of observations, the conventional median--calculated as the average of the two middle values--can introduce bias by implicitly assuming symmetry in the data distribution. This study aims to identify a median estimator that is unbiased. To derive the unbiased median estimator, we minimized the sum of residuals raised to a rational power approaching one. We compared the properties of the unbiased and conventional medians using Poisson-distributed datasets. Random samples were generated with the Mersenne Twister algorithm implemented in IgorPro software (WaveMetrics Inc., Oregon). For odd sample sizes, the unbiased median coincides with the conventional median (the middle value). For even sample sizes, the unbiased median is defined as the value that equalizes the product of distances to data points above and below it--a definition that differs from the conventional median in asymmetric distributions. Although both median estimators tend to underestimate the mean of Poisson-distributed data, the unbiased median is consistently closer to the expected value. Additionally, the unbiased median exhibits lower variance compared to the conventional median. Thus, for even sample sizes, the proposed unbiased median provides a central tendency measure that is unbiased, more accurate, and has reduced variance relative to the conventional median.

bioinformatics↗

bioRxiv2025

Clostridioides difficile Toxins Alter Host Metabolic Pathway and Bile Acid Homeostasis Gene Expression In Colonic Epithelium

A major risk factor for acquiring Clostridioides difficile is antibiotic usage that disrupts a healthy microbial gut community facilitating establishment of infection. Once established, C. difficile secretes endotoxins (TcdA and TcdB) that are internalized into host colonic epithelial cells where they disrupt gut barrier function and induce hyper inflammation resulting in severe diarrhea and possibly leading to death. We employed three different platforms to explore gene expression of cells in the gut when exposed to C. difficile or its toxins, TcdA and TcdB. An antibiotic treated mouse model of C. difficile infection (CDI) was used to identify differential gene expression with a NanoString Technologies mouse inflammatory gene panel consisting of 770 genes including a subset of bile acid (BA) homeostasis and nuclear receptor genes. In the cecal tissue of mice with CDI, significant down expression was observed for genes involved in PPAR signaling, cholesterol and glucose metabolism, while a significant increase in expression was observed for IL-17 related inflammatory genes. Similarly, Caco-2 cell culture and primary human colonocytes (hCE) exposed to toxins for 24 hours showed altered expression in several PPAR regulated and cholesterol metabolic genes similar to those found in mice. These cell culture experiments also revealed significant alterations in gene expression of the FXR BA regulatory pathway. Together these data suggest that exposure to C. difficile and its toxins may alter host cholesterol metabolic processes, including BA transport and synthesis.

microbiology↗

bioRxiv2025

Saturation kinetics and specificity of transporters for L-arginine and asymmetric dimethylarginine (ADMA) at the blood-brain and blood-CSF barriers.

Nitric oxide synthases (NOS) synthesize nitric oxide (NO) from L-arginine in endothelial and neuronal cells. Asymmetric dimethylarginine (ADMA) is a homologue of arginine and an endogenous inhibitor of NOS. As NO is a critical signalling molecule and influences physiological pathways in health and disease, the transfer of arginine and ADMA across the blood-CNS barriers is of interest. Our research group have previously demonstrated the presence of saturable transporters for [3H]-L-arginine and [3H]-ADMA at the blood-brain and blood-CSF barriers using in vitro and in situ methods. In this study, we determine the identity and kinetic characteristics of these transporters by means of the in situ brain/choroid plexus perfusion technique in anaesthetised mice. Results indicated that [3H]-arginine and [3H]-ADMA could be transported across blood-brain and blood-CSF barriers by the cationic amino acid transporter, system-y+. In contrast to the results obtained with arginine where transport was predominately by a single transport system (system-y+), ADMA delivery to the CNS was more complex and involved multiple transport systems (system y+, B0,+, y+L and b0,+) suggesting its concentration is tightly regulated. System y+ and system y+L transporters could be involved in the CNS to blood efflux of ADMA that we have previously observed. The half-saturation constant (Km) and maximal influx rate of the saturable component (Vmax) for [3H]-ADMA transport into the frontal cortex was 29.07{+/-}7.19 M and 0.307{+/-}0.017 nmol.min-1.g-1, respectively, and into the CSF was 30.59{+/-}25.41 M and 2.07{+/-}0.38 nmol.min-1.g-1, respectively. This information could help explain the arginine paradox providing evidence that ADMA interacts with transporters that can remove ADMA from cells. These removal mechanisms could be stimulated by excess arginine in the plasma resulting in increased NO production. It remains to be seen if arginine supplementation could be used to increase NO production and improve hypoperfusion observed in disease states such as Alzheimers and stroke.

neuroscience↗

bioRxiv2024

Central activation of catecholamine-independent lipolysis drives the end-stage catabolism of all adipose tissues

Several adipose depots, including constitutive bone marrow adipose tissue (cBMAT), resist conventional lipolytic cues, making them metabolically non-responsive. However, under starvation, wasting, or cachexia, the body can eventually catabolize these stable adipocytes through unknown mechanisms. To study this, we developed a mouse model of brain-evoked depletion of all fat, including cBMAT, independent of food intake. Genetic, surgical, and chemical approaches demonstrated that depletion of stable fat required adipose triglyceride lipase-dependent lipolysis but was independent of local nerves, the sympathetic nervous system, and catecholamines. Instead, concurrent hypoglycemia and hypoinsulinemia activated a potent catabolic state by suppressing lipid storage and increasing catecholamine-independent lipolysis via downregulation of cell-autonomous lipolytic inhibitors Acvr1c, G0s2, and Npr3. This was also sufficient to delipidate classical adipose depots. Overall, this work defines unique adaptations of stable adipocytes to resist lipolysis in healthy states while isolating a potent in vivo neurosystemic pathway by which the body can rapidly catabolize all adipose tissues.

physiology↗

bioRxiv2024

L-Arginine and asymmetric dimethylarginine (ADMA) transport across the mouse blood-brain and blood-CSF barriers: evidence of saturable transport at both interfaces and CNS to blood efflux.

L-Arginine is the physiological substrate for the nitric oxide synthase (NOS) family, which synthesises nitric oxide (NO) in endothelial and neuronal cells. NO synthesis can be inhibited by endogenous asymmetric dimethylarginine (ADMA). NO has explicit roles in cellular signalling and vasodilation. Impaired NO bioavailability represents the central feature of endothelial dysfunction associated with vascular diseases. Interestingly, dietary supplementation with L-arginine has been shown to alleviate endothelial dysfunctions caused by impaired NO synthesis. In this study the transport kinetics of [3H]-arginine and [3H]-ADMA into the central nervous system (CNS) were investigated using physicochemical assessment and the in situ brain/choroid plexus perfusion technique in anesthetized mice. Results indicated that L-arginine and ADMA are tripolar cationic amino acids and have a gross charge at pH 7.4 of 0.981. L-Arginine (0.00149{+/-}0.00016) has a lower lipophilicity than ADMA (0.00226{+/-}0.00006) as measured using octanol-saline partition coefficients. The in situ perfusion studies revealed that [3H]-arginine and [3H]-ADMA can cross the blood-brain barrier (BBB) and the blood-CSF barrier. [3H]-Arginine (11.6nM) and [3H]-ADMA (62.5nM) having unidirectional transfer constants (Kin) into the frontal cortex of 5.84{+/-}0.86 and 2.49{+/-}0.35 l.min-1.g-1, respectively, and into the CSF of 1.08{+/-}0.24 and 2.70{+/-}0.90 l.min-1.g-1, respectively. In addition, multiple-time uptake studies revealed the presence of CNS-to-blood efflux of ADMA. Self- and cross-inhibition studies indicated the presence of transporters at the BBB and the blood-CSF barriers for both amino acids, which were shared to some degree. Importantly, these results are the first to demonstrate: (i) saturable transport of [3H]-ADMA at the blood-CSF barrier (choroid plexus) and (ii) a significant CNS to blood efflux of [3H]-ADMA. Our results suggest that the arginine paradox, in other words the clinical observation that NO-deficient patients respond well to oral supplementation with L-arginine even though the plasma concentration is easily sufficient to saturate endothelial NOS, could be related to ADMA transport.

neuroscience↗

bioRxiv2024

Air pollution drives macrophage senescence through a phagolysosome-15-lipoxygenase pathway

Urban particulate matter (uPM) poses significant health risks, particularly to the respiratory system. Fine particles, such as PM2.5, can penetrate deep into the lungs and exacerbate a range of health problems, including emphysema, asthma, and lung cancer. PM exposure is also linked to extra-pulmonary disorders like heart and neurodegenerative diseases. Moreover, prolonged exposure to elevated PM levels can reduce overall life expectancy. Senescence is a dysfunctional cell state typically associated with age but can also be precipitated by environmental stressors. This study aimed to determine whether uPM could drive senescence in macrophages, an essential cell type involved in particulate phagocytosis-mediated clearance. While it is known that uPM exposure impairs immune function, this deficit is multi-faceted and incompletely understood, partly due to the use of particulates such as diesel exhaust particle (DEP) as a surrogate for true uPM. uPM was collected from several locations in the USA, including Baltimore, Houston, and Phoenix. Bone marrow-derived macrophages (BMDMs) were stimulated with uPM or reference particulates (e.g., DEP) to assess senescence-related parameters. We report that uPM-exposed BMDMs adopt a senescent phenotype characterized by increased IL-1 secretion, senescence- associated {beta}-galactosidase activity, and diminished proliferation. Exposure to allergens failed to elicit such a response, supporting a distinction between different types of environmental exposures. uPM-induced senescence was independent of key macrophage activation pathways, specifically inflammasome and scavenger receptor. However, inhibition of the phagolysosome pathway abrogated senescence markers, supporting this phenotypes attribution to uPM phagocytosis. These data suggest uPM exposure leads to macrophage senescence, which may contribute to immunopathology.

immunology↗

bioRxiv2023

Interaction of amisulpride with GLUT1 at the blood-brain barrier. Relevance to Alzheimer's disease

Blood-brain barrier (BBB) dysfunction may be involved in the increased sensitivity of Alzheimers disease (AD) patients to antipsychotics, including amisulpride. Studies indicate that antipsychotics interact with facilitated glucose transporters (GLUT), including GLUT1, and that GLUT1 BBB expression decreases in AD. We tested the hypotheses that amisulpride (charge: +1) interacts with GLUT1, and that BBB transport of amisulpride is compromised in AD. GLUT1 substrates and inhibitors, and GLUT-interacting antipsychotics were identified by literature review and their physicochemical characteristics summarised. Interactions between amisulpride, and GLUT1 were studied using in silico approaches and the human cerebral endothelial cell line, hCMEC/D3. Brain distribution of [3H]amisulpride was determined using in situ perfusion in wild type (WT) and 5xFamilial AD (5xFAD) mice. With transmission electron microscopy (TEM) we investigated brain capillary degeneration in WT and 5xFAD mice, and human samples. Western blots determined BBB transporter expression in mouse and human. Literature review revealed that, although D-glucose has no charge, charged molecules can interact with GLUT1. GLUT1 substrates are smaller (184.95{+/-}6.45g/mol) than inhibitors (325.50{+/-}14.40g/mol), and GLUT-interacting antipsychotics (369.38{+/-}16.04). Molecular docking showed beta-D-glucose (free energy binding: -15.39kcal/mol) and amisulpride (-29.04kcal/mol) interact with GLUT1. Amisulpride did not affect [14C]D-glucose accumulation in hCMEC/D3. 5xFAD mice showed increased brain [3H]amisulpride uptake, and no cerebrovascular space changes compared to WT. TEM revealed brain capillary degeneration in human AD. There was no significant effect of AD on mouse GLUT1 and P-gp BBB expression, and in human GLUT1 expression. In contrast, caudate P-glycoprotein expression was decreased in human AD capillaries versus controls. This study provides new details about the BBB transport of amisulpride, evidence that amisulpride interacts with GLUT1, and that BBB transporter expression is altered in AD. This suggests that antipsychotics exacerbate the cerebral hypometabolism in AD. Further research into the mechanism of amisulpride transport by GLUT1 is important for improving antipsychotics safety.

neuroscience↗

bioRxiv2023

The las and rhl Quorum Sensing Systems in Pseudomonas aeruginosa Form a Multi-Signal Reciprocal Network Which Can Tune Reactivity to Variations in Physical and Social Environments

Researchers often view the multi-signal quorum sensing systems of Pseudomonas aeruginosa as a hierarchy, topped by the las system which acts as a master regulator. By experimentally controlling the concentration of auto-inducer signals in a signal null strain (PAO1{Delta}lasI{Delta}rhlI), we show that the two primary quorum sensing systems--las and rhl--act reciprocally rather than hierarchically. Just as the las systems 3-oxo-C12-HSL can induce increased expression of rhlI, the rhl systems C4-HSL increases the expression level of lasI. We develop a mathematical model to quantify relationships both within and between the las and rhl quorum sensing systems and the downstream genes they influence. The results show that not only do the systems interact reciprocally, but they do so cooperatively and nonlinearly, with the combination of C4-HSL and 3-oxo-C12-HSL increasing expression level far more than the sum of their individual effects. We computationally assess how our parameterized model responds to variation in social (population density) and physical (mass transfer) environment and demonstrate that a reciprocal architecture is more responsive to density and more robust to mass transfer than a strict hierarchy.

microbiology↗

bioRxiv2022

The dynamic response of quorum-sensing to density is robust to signal supplementation and signal synthase knockouts

Quorum sensing (QS) is a widespread mechanism of environment sensing and behavioral coordination in bacteria. At its core, QS is based on the production, sensing and response to small signaling molecules. Previous work with Pseudomonas aeruginosa shows that QS can be used to achieve quantitative resolution and deliver a dosed response to the bacterias density environment, implying a sophisticated mechanism of control. To shed light on how the mechanistic signal components contribute to graded responses to density, we assess the impact of genetic (AHL signal synthase deletion) and/or signal supplementation (exogenous AHL and PQS addition) perturbations on lasB reaction-norms to changes in density. Our approach condenses data from 2,000 timeseries (over 74,000 individual observations) into a comprehensive view of QS-controlled gene expression across variation in genetic, environmental, and signal determinants of lasB expression. We first confirm that deleting either ({Delta}lasI, {Delta}rhlI) or both ({Delta}lasIrhlI) signal synthase genes attenuates QS response. In the {Delta}rhlI background we show persistent yet attenuated density-dependent lasB expression due to native 3-oxo-C12 signaling. We then test if density-independent quantities of signal (3-oxo-C12, C4, PQS or combined) added to the WT either flatten or increase the reaction norm and find that the WT response is robust to all tested concentrations of signal, alone or in combination. We then move to progressively supplementing the genetic knockouts and find that cognate signal supplementation ({Delta}lasI+3-oxo-C12, {Delta}rhlI+C4) is sufficient to restore lasB expression and as well as reactivity to density encoded by the intact signal synthase. We also find that dual supplementation of the double synthase knockout restores expression but does not flatten the reaction norm. Despite adding a density-independent amount of AHL, the double signal synthase can still quantitively sense density. Our results show that a positive reaction norm to density is robust to multiple combinations of gene deletion and density-independent signal supplementation and that while density-independent signal supplementation can increase mean expression, the WT QS still retains the ability to quantitatively respond to density. Our work develops a modular approach to query the robustness and mechanistic bases of the central environmental sensing phenotype of quorum sensing.

microbiology↗

bioRxiv2021

Objective method reveals minimum time required to quantify net cost of transport across slow to fast walking speeds

The U-shaped net cost of transport (COT) curve of walking has helped scientists understand the biomechanical basis that underlies energy minimization during walking. However, to produce an individuals net COT curve, data must be analyzed during periods of steady-rate metabolism. Traditionally, studies analyze the last few minutes of a 6-10 min trial, assuming that steady-rate metabolism has been achieved. Yet, it is possible that an individual achieves steady rates of metabolism much earlier. However, there is no consensus on how to objectively quantify steady-rate metabolism across a range of walking speeds. Therefore, we developed an objective method to determine the minimum time needed for humans to achieve steady rates of metabolism across slow to fast walking speeds. We hypothesized that a shorter time window could be used to produce a net COT curve that is comparable to the net COT curve created using traditional methods. We analyzed metabolic data from twenty-one subjects who completed several 7-min walking trials ranging from 0.50-2.00 m/s. We partitioned the metabolic data for each trial into moving 1-min, 2-min, and 3 min intervals and calculated their slopes. We statistically compared these slope values to values derived from the last 3-min of the 7-min trial, our gold standard comparison. We found that a minimum of 2 min is required to achieve steady-rate metabolism and that data from 2-4 min yields a net COT curve that is not statistically different from the one derived from experimental protocols that are generally accepted in the field. New and NoteworthyA simple slope method reveals that 4-min of walking at speeds between 0.50-1.75 m/s is the minimum time required to produce an accurate net cost of transport curve.

physiology↗

bioRxiv2021

Pseudomonas aeruginosa reaches collective decisions via transient segregation of quorum sensing activities across cells

Bacteria engage in a cell-to-cell communication process called quorum sensing (QS) to coordinate expression of cooperative exoproducts at the group level. While population-level QS-responses are well studied, we know little about commitments of single cells to QS. Here, we use flow cytometry to track the investment of Pseudomonas aeruginosa individuals into their intertwined Las and Rhl QS-systems. Using fluorescent reporters, we show that QS gene expression (signal synthase, receptor and exoproduct) was heterogenous and followed a gradual instead of a sharp temporal induction pattern. The simultaneous monitoring of two QS genes revealed that cells transiently segregate into low receptor (lasR) expressers that fully commit to QS, and high receptor expressers that delay QS commitment. Our mathematical model shows that such gene expression segregation could mechanistically be spurred by transcription factor limitation. In evolutionary terms, temporal segregation could serve as a QS-brake to allow for a bet-hedging strategy in unpredictable environments.

microbiology↗

bioRxiv2020

Training a neural network to learn other dimensionality reduction removes data size restrictions in bioinformatics and provides a new route to exploring data representations

High dimensionality omics and hyperspectral imaging datasets present difficult challenges for feature extraction and data mining due to huge numbers of features that cannot be simultaneously examined. The sample numbers and variables of these methods are constantly growing as new technologies are developed, and computational analysis needs to evolve to keep up with growing demand. Current state of the art algorithms can handle some routine datasets but struggle when datasets grow above a certain size. We present a training deep learning via neural networks on non-linear dimensionality reduction, in particular t-distributed stochastic neighbour embedding (t-SNE), to overcome prior limitations of these methods. One Sentence SummaryAnalysis of prohibitively large datasets by combining deep learning via neural networks with non-linear dimensionality reduction.

bioinformatics↗

bioRxiv2019

Temperature-dependence of metabolic rate in tropical and temperate aquatic insects: support for the Climate Variability Hypothesis in mayflies but not stoneflies.

A fundamental gap in climate change vulnerability research is an understanding of the relative thermal sensitivity of ectotherms. Aquatic insects are vital to stream ecosystem function and biodiversity but insufficiently studied with respect to their thermal physiology. With global temperatures rising at an unprecedented rate, it is imperative that we know how aquatic insects respond to increasing temperature and whether these responses vary among taxa, latitudes, and elevations. We evaluated the thermal sensitivity of standard metabolic rate in stream-dwelling baetid mayflies and perlid stoneflies across a ~2,000 m elevation gradient in the temperate Rocky Mountains in Colorado, U.S.A., and the tropical Andes in Napo, Ecuador. We used temperature-controlled water baths and microrespirometry to estimate changes in oxygen consumption. Tropical mayflies generally exhibited greater thermal sensitivity in metabolism compared to temperate mayflies; tropical mayfly metabolic rates increased more rapidly with temperature and the insects more frequently exhibited behavioral signs of thermal stress. By contrast, temperate and tropical stoneflies did not clearly differ. Varied responses to temperature among baetid mayflies and perlid stoneflies may reflect differences in evolutionary history or ecological roles as herbivores and predators, respectively. Our results show that there is physiological variation across elevations and species and that low elevation tropical mayflies may be especially imperiled by climate warming. Given such variation among species, broad generalizations about the vulnerability of tropical ectotherms should be made more cautiously.

ecology↗

bioRxiv2019

Bacterial quorum sensing allows graded responses to variations in density, on both individual and population scales

Quorum sensing (QS) is a mechanism of cell-cell communication that connects gene expression to environmental conditions (e.g. density) in many bacterial species, mediated by diffusible signal molecules. Current functional studies focus on a dichotomy of QS on/off (or, quorate / sub-quorate) states, overlooking the potential for intermediate, graded responses to shifts in the environment. Here, we track QS regulated protease (lasB) expression and show that Pseudomonas aeruginosa can deliver a graded behavioral response to fine-scale variation in population density, on both the population and single-cell scales. On the population scale, we see a graded response to variation in environmental population density. On the single-cell scale, we see significant bimodality at higher densities, with separate OFF and ON sub-populations that respond differentially to changes in density; static OFF cells and increasing intensity of expression among ON cells. Together these results indicate that QS can tune gene expression to graded environmental change, with no critical cell mass or quorum at which behavioral responses are activated on either the individual cell or population scale. In an infection context, our results indicate there is not a hard threshold separating sub-quorate stealth mode and a quorate attack mode.

microbiology↗

bioRxiv2019

In silico bacteria evolve robust cooperation via complex quorum-sensing strategies

Many species of bacteria collectively sense and respond to their social and physical environment via quorum sensing (QS), a communication system controlling extracellular cooperative traits. Despite detailed understanding of the mechanisms of signal production and response, there remains considerable debate over the functional role(s) of QS: in short, what is it for? Experimental studies have found support for diverse functional roles: density sensing, mass-transfer sensing, genotype sensing, etc. While consistent with theory, these results cannot separate whether these functions were drivers of QS adaption, or simply artifacts or spandrels of systems shaped by distinct ecological pressures. The challenge of separating spandrels from drivers of adaptation is particularly hard to address using extant bacterial species with poorly understood current ecologies (let alone their ecological histories). To understand the relationship between environmental challenges and trajectories of QS evolution, we used an agent-based simulation modeling approach. Given genetic mixing, our simulations produce behaviors that recapitulate features of diverse microbial QS systems, including coercive (high signal / low response) and generalized reciprocity (signal auto-regulation) strategists -- that separately and in combination contribute to QS-dependent resilience of QS-controlled cooperation in the face of diverse cheats. We contrast our in silico results with bacterial QS architectures that have evolved under largely unknown ecological contexts, highlighting the critical role of genetic constraints in shaping the shorter term (experimental evolution) dynamics of QS. More broadly, we see experimental evolution of digital organisms as a complementary tool in the search to understand the emergence of complex QS architectures and functions.\n\nAuthor summaryBacteria communicate and cooperate using complex cell-cell signaling systems known as quorum-sensing (QS). While the molecular mechanisms are often well understood, the reasons why bacteria use QS are less clear -- how has QS aided survival and growth? The answer to this question is dependent on the environment of adaptation, and unfortunately our current understanding of QS bacterial ecology is broadly lacking. To address this gap, we studied the evolution of digital organisms, individual-based computer simulations of bacterial populations evolving under defined environmental contexts. Our results pinpoint how simple environmental challenges (variable density and genetic mixing) can lead to the emergence of complex strategies that recapitulate features of bacterial QS, and open a path towards reverse-engineering the environmental drivers of QS.

microbiology↗