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bioRxiv2019

Microbiome Determinants and Physiological Effects of the Benzoate-Hippurate Microbial-Host Co-Metabolic Pathway

ObjectiveGut microbial products are involved in type 2 diabetes, obesity and insulin resistance. In particular, hippurate, a hepatic phase 2 conjugation product of microbial benzoate metabolism, has been associated with a healthy phenotype. This study aims to identify metagenomic determinants and test protective effects of hippurate. DesignWe profiled the urine metabolome by 1H Nuclear Magnetic Resonance (NMR) spectroscopy to derive associations with metagenomic sequences in 271 middle-aged Danish individuals to identify dietary patterns in which urine hippurate levels were associated with health benefits. We follow up with benzoate and hippurate infusion in mice to demonstrate causality on clinical phenotypes. ResultsIn-depth analysis identifies that the urine hippurate concentration is associated with microbial gene richness, microbial functional redundancy as well as functional modules for microbial benzoate biosynthetic pathways across several enterotypes. Through dietary stratification, we identify a subset of study participants consuming a diet rich in saturated fat in which urine hippurate, independently of gene richness, accounts for links with metabolic health that we previously associated with gene richness. We then demonstrate causality in vivo through chronic subcutaneous infusions of hippurate or benzoate (20 nmol/day) resulting in improved glycemic control in mice fed a high-fat diet. Hippurate improved insulin secretion through increased {beta}-cell mass and reduced liver inflammation and fibrosis, whereas benzoate treatment resulted in liver inflammation. ConclusionOur translational study shows that the benzoate-hippurate pathway brings a range of metabolic improvements in the context of high-fat diets, highlighting the potential of hippurate as a mediator of metabolic health.

systems biology↗

bioRxiv2020

ER stress in the dorsal root ganglia regulates BK channel physiology and contributes to pain hypersensitivity in a mouse model of multiple sclerosis

Neuropathic pain is a common symptom of multiple sclerosis (MS) and current treatment options are ineffective. In this study, we investigated whether endoplasmic reticulum (ER) stress in dorsal root ganglia (DRG) contributes to pain hypersensitivity in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. Inflammatory cells and increased levels of ER stress markers are evident in post-mortem DRGs from MS patients. Similarly, we observed ER stress in the DRG of mice with EAE and relieving ER stress with a chemical chaperone, 4-phenylbutyric acid (4-PBA), reduced pain hypersensitivity. In vitro, 4-PBA and the selective PERK inhibitor, AMG44, normalize cytosolic Ca2+ transients in putative DRG nociceptors. We went to assess disease-mediated changes in the functional properties of Ca2+-sensitive BK-type K+ channels in DRG neurons. We found that the conductance-voltage (GV) relationship of BK channels was shifted to a more positive voltage, together with a more depolarized resting membrane potential in EAE cells. Our results suggest that ER stress in sensory neurons of MS patients and mice with EAE is a source of pain and that ER stress modulators can effectively counteract this phenotype.

neuroscience↗

bioRxiv2020

A Robust Method for Collecting X-ray Diffraction Data from Protein Crystals across Physiological Temperatures

Traditional X-ray diffraction data collected at cryo-temperatures have delivered invaluable insights into the three-dimensional structures of proteins, providing the backbone of structure-function studies. While cryo-cooling mitigates radiation damage, cryo-temperatures can alter protein conformational ensembles and solvent structure. Further, conformational ensembles underlie protein function and energetics, and recent advances in room-temperature X-ray crystallography have delivered conformational heterogeneity information that is directly related to biological function. The next challenge is to develop a robust and broadly applicable method to collect single-crystal X-ray diffraction data at and above room temperatures and was addressed herein. This approach provides complete diffraction datasets with total collection times as short as ~5 sec from single protein crystals, dramatically increasing the amount of data that can be collected within allocated synchrotron beam time. Its applicability was demonstrated by collecting 1.09-1.54 [A] resolution data over a temperature range of 293-363 K for proteinase K, thaumatin, and lysozyme crystals. Our analyses indicate that the diffraction data is of high-quality and do not suffer from excessive dehydration or damage.

biophysics↗

bioRxiv2020

Dynamic Regulation of Growth and Physiology of Microbes under Complex Changing Conditions

As bacteria transition from exponential to stationary phase they change greatly in size, morphology, growth and expression-profiles. These responses also vary between individual cells, but it has proven difficult to track cell lineages along the growth curve to determine the progression of events or correlations between how individual cells enter and exit dormancy. We developed a platform for tracking >105 parallel cell lineages in dense and changing cultures, independently validating that the imaged cells closely track batch populations. Initial applications show that for both Escherichia coli and Bacillus subtilis, growth changes from an adder mode in exponential phase to mixed adders-timers entering stationary phase, and then a near-perfect sizer upon exit - creating broadly distributed cell sizes in stationary phase but rapidly returning to narrowly distributed sizes upon exit. Furthermore, cells that undergo more divisions entering stationary phase suffer reduced survival after long periods of dormancy but are the only cells observed that persist against antibiotics.

microbiology↗

bioRxiv2020

Bedscales: A non-contact adherence-independent multi-person sensor for longitudinal physiologic monitoring in the home bed

Home health monitoring technologies promise to improve care and reduce costs, yet they are limited by the need for adherence to self-monitoring, usage of an app, or application of a wearable. While implantable sensors overcome the adherence barrier, they are expensive and require invasive procedures. Here, we describe a non-invasive, non-contact, adherence-independent sensor, that when placed beneath the legs of a patients home bed, longitudinally monitors total body weight, detailed respiratory signals, and ballistocardiograms for months, without requiring any active patient participation. Accompanying algorithms demix weight and respiratory signals when the bed is shared by a partner or a pet. Validation studies during overnight clinical sleep studies exhibit quantitative equivalence to commercial sensors and allow discrimination of obstructive and central sleep apneas. In-home studies discriminate atrial fibrillation from normal sinus rhythm. To demonstrate real-world feasibility, we performed 3 months of continuous in-home monitoring in a patient with heart failure as he awaited and recovered from coronary artery bypass surgery. By overcoming the adherence barrier, Bedscales has the potential to create a multidimensional picture of chronic disease, learn signatures of impending hospitalization, and enable optimization of care in the home. DisclosuresDrs. King and Coleman and Nicholas Harrington are inventors on a patent application describing the Bedscales technology. Dr. Kevin R. King discloses consulting relationships with Bristol Myers Squibb and Astrazeneca, Medimmune and is founder of Nightingale Labs. Dr. Deepak L. Bhatt discloses the following relationships - Advisory Board: Cardax, Cereno Scientific, Elsevier Practice Update Cardiology, Medscape Cardiology, PhaseBio, Regado Biosciences; Board of Directors: Boston VA Research Institute, Society of Cardiovascular Patient Care, TobeSoft; Chair: American Heart Association Quality Oversight Committee; Data Monitoring Committees: Baim Institute for Clinical Research (formerly Harvard Clinical Research Institute, for the PORTICO trial, funded by St. Jude Medical, now Abbott), Cleveland Clinic (including for the ExCEED trial, funded by Edwards), Duke Clinical Research Institute, Mayo Clinic, Mount Sinai School of Medicine (for the ENVISAGE trial, funded by Daiichi Sankyo), Population Health Research Institute; Honoraria: American College of Cardiology (Senior Associate Editor, Clinical Trials and News, ACC.org; Vice-Chair, ACC Accreditation Committee), Baim Institute for Clinical Research (formerly Harvard Clinical Research Institute; RE-DUAL PCI clinical trial steering committee funded by Boehringer Ingelheim; AEGIS-II executive committee funded by CSL Behring), Belvoir Publications (Editor in Chief, Harvard Heart Letter), Duke Clinical Research Institute (clinical trial steering committees, including for the PRONOUNCE trial, funded by Ferring Pharmaceuticals), HMP Global (Editor in Chief, Journal of Invasive Cardiology), Journal of the American College of Cardiology (Guest Editor; Associate Editor), Medtelligence/ReachMD (CME steering committees), Population Health Research Institute (for the COMPASS operations committee, publications committee, steering committee, and USA national co-leader, funded by Bayer), Slack Publications (Chief Medical Editor, Cardiology Todays Intervention), Society of Cardiovascular Patient Care (Secretary/Treasurer), WebMD (CME steering committees); Other: Clinical Cardiology (Deputy Editor), NCDR-ACTION Registry Steering Committee (Chair), VA CART Research and Publications Committee (Chair); Research Funding: Abbott, Afimmune, Amarin, Amgen, AstraZeneca, Bayer, Boehringer Ingelheim, Bristol-Myers Squibb, Chiesi, CSL Behring, Eisai, Ethicon, Ferring Pharmaceuticals, Forest Laboratories, Fractyl, Idorsia, Ironwood, Ischemix, Lilly, Medtronic, PhaseBio, Pfizer, PLx Pharma, Regeneron, Roche, Sanofi Aventis, Synaptic, The Medicines Company; Royalties: Elsevier (Editor, Cardiovascular Intervention: A Companion to Braunwalds Heart Disease); Site Co-Investigator: Biotronik, Boston Scientific, CSI, St. Jude Medical (now Abbott), Svelte; Trustee: American College of Cardiology; Unfunded Research: FlowCo, Merck, Novo Nordisk, Takeda. Dr. Akshay S. Desai discloses the following relationships - Research grants to Brigham and Womens Hospital to support clinical trial activities from Alnylam, AstraZeneca, and Novartis; Consulting fees from Abbott, Alnylam, AstraZeneca, Biofourmis, Boehringer-Ingelheim, Boston Scientific, Merck, Novartis, Relypsa, Regeneron. Dr. Owens reports consulting fees from Novartis, and research grants to UCSD from Snoozeal, Nitto Denko, and Masimo.

bioengineering↗

bioRxiv2020

Tumor-infiltrating nerves create an electro-physiologically active microenvironment and contribute to treatment resistance

Patients with densely innervated tumors do poorly as compared to those with sparsely innervated disease. Why some tumors heavily recruit nerves while others do not, remains unknown as does the functional contribution of tumor-infiltrating nerves to cancer. Moreover, while patients receive chemotherapeutic treatment, whether these drugs affect nerve recruitment has not been tested. Using a murine model of ovarian cancer, we show that tumor-infiltrating sensory nerves potentiate tumor growth, decrease survival, and contribute to treatment resistance. Furthermore, matched patient samples show significantly increased tumor innervation following chemotherapy. In vitro analysis of tumor-released extracellular vesicles (sEVs) shows they harbor neurite outgrowth activity. These data suggest that chemotherapy may alter sEV cargo, endowing it with robust nerve recruiting capacity.

cancer biology↗

bioRxiv2020

Automatic deep learning driven label-free image guided patch clamp system for human and rodent in vitro slice physiology

Patch clamp recording of neurons is a labor-intensive and time-consuming procedure. We have developed a tool that fully automatically performs electrophysiological recordings in label-free tissue slices. The automation covers the detection of cells in label-free images, calibration of the micropipette movement, approach to the cell with the pipette, formation of the whole-cell configuration, and recording. The cell detection is based on deep learning. The model was trained on a new image database of neurons in unlabeled brain tissue slices. The pipette tip detection and approaching phase use image analysis techniques for precise movements. High-quality measurements were performed on hundreds of human and rodent neurons. We also demonstrate that further molecular and anatomical analysis can be performed on the recorded cells. The software has a diary module that automatically logs patch clamp events. Our tool can multiply the number of daily measurements to help brain research. ONE SENTENCE SUMMARYNovel deep learning and image analysis algorithms for automated patch clamp systems to reliably measure neurons in human and rodent brain slices.

neuroscience↗

bioRxiv2020

Robust and Gradient Thickness Porous Membranes for In Vitro Modeling of Physiological Barriers

Porous membranes are fundamental elements for tissue-chip barrier and co-culture models. However, the exaggerated thickness of commonly available membranes impedes an accurate in vitro reproduction of the biological multi-cellular continuum as it occurs in vivo. Existing techniques to fabricate membranes such as solvent cast, spin-coating, sputtering and PE-CVD result in uniform thickness films. To understand critical separation distances for various barrier and co-culture models, a gradient thickness membrane is needed. Here, we developed a robust method to generate ultrathin porous parylene C (UPP) membranes not just with precise thicknesses down to 300 nm, but with variable gradients in thicknesses, while at the same time having porosities up to 25%. We also show surface etching and increased roughness lead to improved cell attachment. Next, we examined the mechanical properties of UPP membranes with varying porosity and thickness and fit our data to previously published models, which can help determine practical upper limits of porosity and lower limits of thickness. Lastly, we validate a straightforward approach allowing the successful integration of the UPP membranes into a prototyped 3D-printed scaffold enabling in vitro barrier modeling and investigation of cell-cell interplay over variable distances using thickness gradients.

bioengineering↗

bioRxiv2020

NDI: A platform-independent data interface and database for neuroscience physiology and imaging experiments

Collaboration in neuroscience is impeded by the difficulty of sharing primary data, results, and software across labs. Here we introduce Neuroscience Data Interface (NDI), a platform-independent standard that allows an analyst to use and create software that functions independently from the format of the raw data or the manner in which the data is organized into files. The interface is rooted in a simple vocabulary that describes common apparatus and storage devices used in neuroscience experiments. Results of analyses - and analyses of analyses - are stored as documents in a scalable, queryable database that stores the relationships and history among the experiment elements and documents. The interface allows the development of an application ecosystem where applications can focus on calculation rather than data format or organization. This tool can be used by individual labs to exchange and analyze data, and it can serve to curate neuroscience data for searchable archives.

neuroscience↗

bioRxiv2020

ACE2 interaction networks in COVID-19: a physiological framework for prediction of outcome in patients with cardiovascular risk factors

(1) BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (coronavirus disease 2019; COVID-19) is associated with adverse outcomes in patients with cardiovascular disease (CVD). The aim of the study was to characterize the interaction between SARS-CoV-2 and Angiotensin-Converting Enzyme 2 (ACE2) functional networks with a focus on CVD.; (2) MethodsUsing the network medicine approach and publicly available datasets, we investigated ACE2 tissue expression and described ACE2 interaction networks which could be affected by SARS-CoV-2 infection in the heart, lungs and nervous system. We compared them with changes in ACE-2 networks following SARS-CoV-2 infection by analyzing public data of stem cell-derived cardiomyocytes (hiPSC-CMs). This analysis was performed using the NERI algorithm, which integrates protein-protein interaction with co-expression networks. We also performed miRNA-target predictions to identify which ones regulate ACE2-related networks and could play a role in the COVID19 outcome. Finally, we performed enrichment analysis for identifying the main COVID-19 risk groups. (3) ResultsWe found similar ACE2 expression confidence levels in respiratory and cardiovascular systems, supporting that heart tissue is a potential target of SARS-CoV-2. Analysis of ACE2 interaction networks in infected hiPSC-CMs identified multiple hub genes with corrupted signalling which can be responsible for cardiovascular symptoms. The most affected genes were EGFR, FN1, TP53, HSP90AA1, and APP, while the most affected interactions were associated with MAST2 and CALM1. Enrichment analysis revealed multiple diseases associated with the interaction networks of ACE2, especially cancerous diseases, obesity, hypertensive disease, Alzheimers disease, non-insulin-dependent diabetes mellitus, and congestive heart failure. Among affected ACE2-network components connected with SARS-Cov-2 interactome, we identified AGT, CAT, DPP4, CCL2, TFRC and CAV1, associated with cardiovascular risk factors. We described for the first time miRNAs which were common regulators of ACE2 networks and virus-related proteins in all analyzed datasets. The top miRNAs were miR-27a-3p, miR-26b-5p, miR-10b-5p, miR-302c-5p, hsa-miR-587, hsa-miR-1305, hsa-miR-200b-3p, hsa-miR-124-3p, and hsa-miR-16-5p.; (4) ConclusionOur study provides a complete mechanistic framework for investigating the ACE2 network which was validated by expression data. This framework predicted risk groups, including the established ones, thus providing reliable novel information regarding the complexity of signalling pathways affected by SARS-CoV-2. It also identified miR which could be used in personalized diagnosis in COVID-19.

genomics↗

bioRxiv2020

Phenological, Physiological and yield markers as efficient tools to identify drought tolerant rice genotypes in Eastern India

Rice production is severely threatened by drought stress in Eastern India. To develop drought tolerant varieties, selection of donors for breeding programme is crucial. Twenty one selected rice genotypes including both tolerant and sensitive to drought were grown under well-watered and drought stress conditions in dry seasons of two successive years of 2017 and 2018. Leaf water potential, relative water content displayed significant difference among the genotypes during vegetative screening. At reproductive stage drought screening, days to 50% flowering was delayed in all genotypes except N22 and Anjali (showed early flowering) however grain yield and other yield related traits decreased significantly compared to well watered condition. Correlation analysis of phenological and yield related traits with grain yield revealed that tiller numbers and panicle numbers are highly correlated with grain yield both under well-watered and water stress conditions and contributes maximum towards grain yield. The dendrogram grouped Mahamaya, Sahabhagidhan, Poornima, IBD 1, Hazaridhan, Samleshwari and Danteshwari into one cluster which performed better under water stress conditions and had grain yield more than 1.69 tha-1. Sahabhagidhan, Poornima, Vandana, and N22 displayed tolerance to drought both under vegetative and reproductive conditions which could be a good selection for the breeders to develop drought tolerant rice cultivars for eastern region of India.

plant biology↗

bioRxiv2020

Physiological responses of two accessions of tomato to three distance regimes in juvenile oil palm-tomato intercrop in rainforest zone of Nigeria

This study was conducted during the rainy season of 2017 within the alleys of juvenile oil palms that were 2-year-old at the oil palm plantation located in Ala, Akure-North Local Government, Ondo State. Tomato accessions (NGB 01665 and NG/AA/SEP/09/053) were intercropped at 1, 2 and 3 m from the juvenile oil palm within the alley in a randomized complete block design. The results showed that tomato (NGB 01665 and NG/AA/SEP/09/053) planted at 3 m from the juvenile oil palm performed better than those at other planting distances in terms of growth and yield attributes with number of fruit; fruit weight and yield per hectare of 39.50, 2265.8 g and 3.74 ton/ha respectively. However, the control (sole) recorded the best yield but was not significantly different from those planted at 3 m from the juvenile oil palm. Varietal advantage was recorded by tomato (NGB 01665) with higher number of fruits, fruit weight and yield per hectare (26.94, 1834 g and 3.158 ton/ha) respectively. Intercropping advantage was not recorded for any of the intercropping distance regimes during the trial (when the juvenile oil palms were 2-year-old), with LER values less than unity.

plant biology↗

bioRxiv2020

Leaf reflectance can surrogate foliar economics better than physiological traits across macrophyte species

Macrophytes are key players in aquatic ecosystems diversity, but knowledge on variability of their functional traits, among and within species, is still limited. Remote sensing is a high-throughput, feasible option for characterizing plant traits at different scales, provided that reliable spectroscopy models are calibrated with congruous empirical data. We sampled leaves from six floating and emergent macrophyte species common in temperate areas, covering different phenological stages, seasons, and environmental conditions, and measured leaf reflectance (400-2500 nm) and leaf traits (dealing with photophysiology, pigments and structure). We explored optimal spectral bands combinations and established non-parametric reflectance-based models for selected traits, eventually showing how airborne hyperspectral data can capture spatial-temporal macrophyte variability. Our key finding is that structural - leaf dry matter content, leaf mass per area - and biochemical - chlorophyll-a content and chlorophylls to carotenoids ratio - traits can be surrogated by leaf reflectance with relative error under 20% across macrophyte species, while performance of reflectance-based models for photophysiological traits depends on species. This finding shows the link between leaf reflectance and leaf economics (structure and biochemistry) for aquatic plants, thus supporting the use of remote sensing for enhancing the level of detail of macrophyte functional diversity analysis, to intra-site and intra-species scales.

ecology↗

bioRxiv2020

A Genomics Resource for Genetics, Physiology, and Breeding of West African Sorghum

Local landrace and breeding germplasm is a useful source of genetic diversity for regional and global crop improvement initiatives. Sorghum (Sorghum bicolor L. Moench) in West Africa has diversified across a mosaic of cultures and end-uses, and along steep precipitation and photoperiod gradients. To facilitate germplasm utilization, a West African sorghum association panel (WASAP) of 756 accessions from national breeding programs of Niger, Mali, Senegal, and Togo was assembled and characterized. Genotyping-by-sequencing was used to generate 159,101 high-quality biallelic SNPs, with 43% in intergenic regions and 13% in genic regions. High genetic diversity was observed within the WASAP ({pi} = 0.00045), only slightly less than in a global diversity panel ({pi} = 0.00055). Linkage disequilibrium decayed to background level (r2 < 0.1) by ~50 kb in the WASAP. Genome-wide diversity was structured both by botanical type, and by populations within botanical type, with eight ancestral populations identified. Most populations were distributed across multiple countries, suggesting several potential common gene pools across the national programs. Genome-wide association studies of days to flowering and plant height revealed eight and three significant quantitative trait loci (QTL), respectively, with major height QTL at canonical height loci Dw3 and SbHT7.1. Colocalization of two of eight major flowering time QTL with flowering genes previously described in US germplasm (Ma6 and SbCN8) suggests that photoperiodic flowering in WA sorghum is conditioned by both known and novel genes. This genomic resource provides a foundation for genomics-enabled breeding of climate-resilient varieties in West Africa.

genomics↗

bioRxiv2020

Compensation of physiological motion enables high-yield whole-cell recording in vivo

1.1.1Whole-cell patch-clamp recording in vivo is the gold-standard method for measuring subthreshold electrophysiology from single cells during behavioural tasks, sensory stimulations, and optogenetic manipulation. However, these recordings require a tight, gigaohm resistance, seal between a glass pipette electrodes aperture and a cells membrane. These seals are difficult to form, especially in vivo, in part because of a strong dependence on the distance between the pipette aperture and cell membrane. We elucidate and utilize this dependency to develop an autonomous method for placement and synchronization of pipettes tip aperture to the membrane of a nearby, moving neuron, which enables high-yield seal formation and subsequent recordings in the deep in the brain of the living mouse, in the thalamus. This synchronization procedure nearly doubles the reported gigaseal yield in the thalamus (>3 mm below the pial surface) from 26% (n=17/64) to 48% (n=32/66). Whole-cell recording yield improved from 10% (n = 9/88) to 24% (n=18/76) when motion compensation was used during the gigaseal formation. As an example of its application, we utilized this system to investigate the role of the sensory environment and ventral posterior medial region (VPM) projection synchrony on intracellular dynamics in the barrel cortex. This method results in substantially greater subcortical whole-cell recording yield than previously reported and thus makes pan-brain whole-cell electrophysiology practical in the living mouse brain.

neuroscience↗

bioRxiv2020

ABA analogue produced by Bacillus marisflavi modulates the physiological response of host-plant under drought stress

Present study aims to understand the molecular mechanism involved in beneficial rhizobacteria mediated alleviation of drought stress in host plant. Bacillus marisflavi CRDT-EB-1 isolated from the rhizosphere soil was found effective in inducing resistance against drought stress in mustard seedlings. Among the different bacterial derivatives tested, the culture filtrate was found to contain bioactive molecules. Solvent extract of bacterial culture filtrate yielded seven distinct bands/ fractions on thin layer chromatography (TLC). The fraction four (F4) with Rf value 0.35-0.40 was significant in reducing adverse effect of drought stress in host plants. Application of F4 resulted in delayed drooping point and higher drought tolerance index (3.34), induced stomatal closure (9.648 ), seed germination inhibition (12%), and reduced the GA3 induced -amylase activity in germinating barley seeds. On TLC, F4 turned colorless to orange color upon the spray of 2,4-dinitrophenylhydrazine reagent indicated the presence of aldehyde group. Supporting to this, the peaks between 9.8 to 10.0 ppm in 1H-NMR chromatogram confirmed the presence of aldehyde group. Upon LC-MS/MS analysis of crude extract of culture filtrate and F4 revealed the presence of compounds with the molecular mass 250.33 and 266.33. By analyzing these data, the identity of the bioactive compounds were predicted as xanthoxin and xanthoxic acid, which are well-known precursor of Abscisic acid (ABA) in plants. The present study concludes the capability of ABA analogue (xanthoxin like compounds) production by B. marisflavi CRDT-EB-1 and its involvement in inducing drought stress tolerance in the host plant.

microbiology↗

bioRxiv2020

A general approach for analysis of physiologically structured population models: the R package 'PSPManalysis'

SummaryHow environmental conditions affect the life history of individual organisms and how these effects translate into dynamics of population and communities on ecological and evolutionary time scales is a central question in many eco-evolutionary studies.Physiologically structured population models (PSPMs) offer a theoretical approach to address such questions as they are built upon a function-based model of the life history, which explicitly describes how life history depends on individual traits as well as on environmental factors. PSPMs furthermore explicitly account for population feedback on these environmental factors, which translates into density-dependent effects on the life history. PSPMs can thus capture life histories in quite some detail but lead to population-level formulations in terms of partial differential equations that are generally hard to analyse.Here I present a general methodology and a R software package for computing how the ecological steady states of PSPMs depend on model parameters and to detect bifurcation points in the computed curves where dynamics change drastically. The package makes specifying the population model unnecessary and only requires a relatively straightforward implementation of the life history functions as input. It furthermore allows for analysing the evolutionary dynamics and evolutionary singular states of the PSPMs based on Adaptive Dynamics theory.Given the central role of the individual life history in many studies, there is substantial scope for using the presented methodology in fields as diverse as ecology, ecotoxicology, conservation biology and evolutionary biology, where it has already been applied to problems like the evolution of cannibalism, niche shifts and metamorphosis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

ecology↗

bioRxiv2020

Genetic and Physiological Characterization of the Antibacterial Activity of Bacillus subtilis subsp. inaquosorum Strain T1 Effective Against pirABVp-Bearing Vibrio parahaemolyticus

Acute hepatopancreatic necrosis disease (AHPND) is caused by PirAB toxin-producing Vibrio parahaemolyticus and has devastated the global shrimp aquaculture industry. One approach for preventing growth of AHPND-producing Vibrio spp. is through the application of beneficial bacteria capable of inhibiting these pathogens. In this study we focus on the inhibitory activity of Bacillus subtilis subsp. inaquosorum strain T1, which hinders V. parahaemolyticus growth in co-culture experiments in a density-dependent manner; inhibition was also obtained using cell-free supernatants from T1 stationary phase cultures. Using a mariner-based transposon mutagenesis, 17 mutants were identified having complete or partial loss of inhibitory activity. Of those having total activity loss, 13 had insertions within a 42.6 kb DNA region comprising 15 genes whose deduced products were homologous to non-ribosomal polypeptide synthetases (NRPSs), polyketide synthases (PKSs) and related activities, which were mapped as one transcriptional unit. Mutants with partial activity contained insertions in spo0A and oppA, indicating stationary phase control. Expression of lacZ transcriptional fusions to NRPS and PKS genes was negligible during growth and at their highest during early stationary phase. Inactivation of sigH resulted in loss of inhibitor activity, indicating a role for {sigma}H in transcription. Disruption of abrB resulted in NRPS and PKS gene overexpression during growth as well as enhanced growth inhibition. This is the first study examining expression and control of an NRPS-PKS region unique to the inaquosorum subspecies of B. subtilis and an understanding of factors involved in T1 inhibitor production will enable its development for use as a potential tool against AHPND Vibrio pathogens in shrimp aquaculture. IMPORTANCEThe shrimp aquaculture industry has been impacted by the rise of acute hepatopancreatic necrosis disease (AHPND), resulting in significant financial losses annually. Caused by strains of the bacterial pathogen, Vibrio parahaemolyticus, treatment of AHPND involves the use of antibiotics, which leads to a rise in antibiotic resistant strains. An alternative approach is through the application of beneficial microorganisms having inhibitory activities against AHPND-generating pathogens. In this study, we examine the genetic basis for the ability of Bacillus subtilis strain T1 to inhibit growth of an AHPND Vibrio strain and show that activity is associated with genes having the potential for synthesizing antibacterial compounds. We found that expression of these genes is under stationary phase control and showed that inactivation of a global transition state regulator results in enhancement of inhibitory activity against the AHPND Vibrio. Our approach for understanding the factors involved in production B. subtilis strain T1 inhibitory activity may allow for development of this strain for use as a potential tool for the prevention of AHPND outbreaks.

microbiology↗