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At least 127 records · Page 7
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Changes in environmental and engineered conditions alter the plasma membrane lipidome of fractured shale bacteria

ABSTRACT Microorganisms that persist in fractured shale reservoirs cause several problems including secreting foul gases and forming biofilms. Current biocontrol measures often fail due to limited knowledge of their in situ activities. The plasma membrane protects the cell, mediates many of its critical functions, and responds to intracellular cues and ecological perturbations through physicochemical modifications. As such, it provides valuable insight into the physiological adaptation of microorganisms in disturbed environmental systems. Here, we (i) demonstrate how changes in salinity and hydraulic retention time (HRT) influence the plasma membrane intact polar lipid (IPL) chemistry of model bacterium, Halanaerobium congolense WG10, and mixed microbial consortia enriched from shale-produced fluids and (ii) elucidate adjustments in membrane IPL chemistry during biofilm growth relative to planktonic cells. We incubated H. congolense WG10 in chemostats under three salinities (7%, 13%, and 20% NaCl), operated under three HRTs (19.2, 24, and 48 h), and in drip flow biofilm reactors under the same salinity gradients. Also, mixed microbial consortia in produced fluids were enriched in triplicate chemostat vessels under three HRTs (19.2, 24, and 72 h) and biofilm reactors. Lipids were analyzed by ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Our results show that phosphatidylglycerols, cardiolipins, and phosphatidylethanolamines were predominantly enriched in planktonic H. congolense WG10 cells grown at hypersalinity (20%) compared to optimum (13%). In addition, several zwitterionic phosphatidylcholines and phosphatidylethanolamines were higher in abundance during biofilm growth. These observations suggest that microbial adaptation and biofilm formation in fractured shale are enabled by strategic plasma membrane IPL chemistry adjustments. IMPORTANCE Microorganisms inadvertently introduced into the shale reservoir during fracturing face multiple stressors including brine-level salinities and starvation. However, some anaerobic halotolerant bacteria adapt and persist for long periods of time. They produce hydrogen sulfide, which sours the reservoir and corrodes engineering infrastructure. In addition, they form biofilms on rock matrices, which decrease shale permeability and clog fracture networks. These reduce well productivity and increase extraction costs. Under stress, microbes remodel their plasma membrane to optimize its roles in protection and mediating cellular processes such as signaling, transport, and energy metabolism. Hence, by observing changes in the membrane lipidome of model shale bacteria, Halanaerobium congolense WG10, and mixed consortia enriched from produced fluids under varying subsurface conditions and growth modes, we provide insight that advances our knowledge of the fractured shale biosystem. We also offer data-driven recommendations for improving biocontrol efficacy and the efficiency of energy recovery from unconventional formations.

03 NATURAL GAS↗
DOE OSTI2022

Microbial colonization and persistence in deep fractured shales is guided by metabolic exchanges and viral predation

Background: Microbial colonization of subsurface shales following hydraulic fracturing offers the opportunity to study coupled biotic and abiotic factors that impact microbial persistence in engineered deep subsurface ecosystems. Shale formations underly much of the continental USA and display geographically distinct gradients in temperature and salinity. Complementing studies performed in eastern USA shales that contain brine-like fluids, here we coupled metagenomic and metabolomic approaches to develop the first genome-level insights into ecosystem colonization and microbial community interactions in a lower-salinity, but high-temperature western USA shale formation. Results: We collected materials used during the hydraulic fracturing process (i.e., chemicals, drill muds) paired with temporal sampling of water produced from three different hydraulically fractured wells in the STACK (Sooner Trend Anadarko Basin, Canadian and Kingfisher) shale play in OK, USA. Relative to other shale formations, our metagenomic and metabolomic analyses revealed an expanded taxonomic and metabolic diversity of microorganisms that colonize and persist in fractured shales. Importantly, temporal sampling across all three hydraulic fracturing wells traced the degradation of complex polymers from the hydraulic fracturing process to the production and consumption of organic acids that support sulfate- and thiosulfate-reducing bacteria. Furthermore, we identified 5587 viral genomes and linked many of these to the dominant, colonizing microorganisms, demonstrating the key role that viral predation plays in community dynamics within this closed, engineered system. Lastly, top-side audit sampling of different source materials enabled genome-resolved source tracking, revealing the likely sources of many key colonizing and persisting taxa in these ecosystems. Conclusions: These findings highlight the importance of resource utilization and resistance to viral predation as key traits that enable specific microbial taxa to persist across fractured shale ecosystems. We also demonstrate the importance of materials used in the hydraulic fracturing process as both a source of persisting shale microorganisms and organic substrates that likely aid in sustaining the microbial community. Moreover, we showed that different physicochemical conditions (i.e., salinity, temperature) can influence the composition and functional potential of persisting microbial communities in shale ecosystems. Together, these results expand our knowledge of microbial life in deep subsurface shales and have important ramifications for management and treatment of microbial biomass in hydraulically fractured wells.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Integration of Omics into a New Comprehensive Rate Law for Competitive Terminal Electron-Accepting Processes in Reactive Transport Models: Application to N, Fe, S, and Contaminant Transformations in Stream and Wetland Sediments

Surface waters represent important sources of alternate energy and drinking water in the United States, and characterizing the biogeochemical processes that affect surface water quality is relevant to the DOE-BER mission. Sediment biogeochemical processes regulate the release of carbon (C), nutrients, and contaminants to surface waters and thus influence water quality. Sediment biogeochemical processes are dynamic and affected by the deposition and remobilization of solid material and changes in environmental conditions driven by water discharge variations. Wetlands are important natural filters of surface waters which may either trap, metabolize, or mobilize nutrients and contaminants. Despite their importance, biogeochemical processes regulating nutrient and contaminant release and C transformation in stream and wetland sediments cannot be predicted accurately by current mathematical models. These reactive transport models largely rely on detectable changes in geochemical conditions to activate metabolic processes, do not accurately account for the competition between microbial processes, and poorly constrain effects of hydrological perturbations on biogeochemical processes. In this BER-SBR exploratory project, metagenomic and geochemical signatures were combined to identify microbially-mediated redox processes in anaerobic stream and wetland sediments from the Savannah River Site (SRS, ANL SFA) and East Fork Poplar Creek (EFPC, ORNL SFA) that play important roles in C, uranium (U), and mercury (Hg) transformations. In addition, sediment incubations were conducted to examine the competition between anaerobic respiration processes Finally, new rate laws were developed for reactive transport models that rely on complementary metagenomic and geochemical signatures to identify the underlying anaerobic microbial processes in stream and wetland sediments, describe the competition between the dominant metabolic processes involved in nutrient release and U and Hg mobilization, and more accurately quantify carbon transformation and the response of microbial processes to changes in redox conditions associated with hydrological forcing. These rate laws were optimized in batch reactors with SRS wetland sediments, where iron and sulfate reduction dominate. Anaerobic carbon remineralization processes followed the expected thermodynamic sequence of microbial respiration with depth in both sediments, except that geochemical signals indicated that sulfate reduction was inactive in EFPC sediments and moderate in SRS wetland sediments. Estimates indicated that microbial iron reduction contributed to at least half of the production of reduced iron in these sediments. Incubations demonstrated that nitrate reduction, denitrification, and dissimilatory nitrate reduction to ammonium were active in the natural EFPC sediment and activated upon nitrate amendment in these nitrate-rich sediments. In turn, these processes were outcompeted by the addition of either iron oxides or sulfate as alternative terminal electron acceptors. Although geochemical products of sulfate reduction were not detected in the incubations, the abundance of sulfate reduction genes increased with depth in the sediment and was equally more pronounced in treatments amended with either iron oxides or sulfate. Simultaneously, anaerobic sulfide oxidizing bacteria coupling sulfide oxidation to DNRA (and not conventional denitrification) were apparently enriched over time, regardless of the treatments. These findings indicate that sulfate reduction is important in freshwater stream sediments and probably catalyzed by a cryptic sulfur cycle involving nitrogen species, in which the sulfur products from sulfate reduction are immediately removed by side reactions and not detectable by geochemical measurements alone. Similar experiments in SRS sediments, however, demonstrated little interaction between nitrogen and sulfur cycling microorganisms. Sulfate reduction was impacted by the addition of more thermodynamically favorable electron acceptors, suggesting either that iron-reducing microorganisms outcompeted sulfate-reducing microorganisms for organic substrate, depleting the stock of electron donor available for sulfate reduction, or that the cryptic sulfur cycle was shunted by the precipitation of FeS generated as a result of the abiotic reduction of iron oxides by dissolved sulfide. A diagnostic modeling exercise was conducted to further investigate the competition between terminal electron accepting processes. As conventional kinetic models typically do not account for cryptic cycles and used inaccurate formulations to describe competition between microbial communities, new metabolic rate laws were developed that explicitly express the electron acceptor-specific enzyme of each energetically favorable metabolic process based on gene abundance detected in the incubations. The model was tested with the sediment slurry incubation data to determine whether substrate competition could explain the decrease in sulfate reduction observed in the presence of iron oxide competitor. The model was able to reproduce geochemical concentrations really well in each treatment once the model was calibrated with the unamended control, suggesting that microbial competition was indeed driven by thermodynamic considerations. Overall, carbon remineralization processes and rates will be reproduced much more realistically with the new metabolic rate laws.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Dynamic, Adaptive, Systems and Materials: Complex, Simple and Emergent Behaviors

This program has been funded by DoE/BES for twenty years. It has moved into and out of various subjects as it has developed, but it has retained its focus on complexity and complex systems. The project has evolved in the following way: Self-Assembly and Biomimetic Self-Assembly: All self-assembling systems depend upon a minimum of two types of interaction: a repulsion and an attraction. For the familiar molecular systems, attractive interactions are typically hydrogen bonds and electrostatic interactions. Repulsive interactions include steric effects, hydrophobic effects (in biological systems), and charge-charge repulsion. We have expanded this repertoire to include surface interactions, magnetic interactions, and others. I list these systems in the order in which we have explored them: i) A key emphasis in current work is in understanding how the movement of ions in a magnetic field (the Lorentz effect) interacts with catalytic systems. We have demonstrated that an acceleration in rate of reduction of CO 2 to CO can be accomplished by applying an external magnetic field. This acceleration is largely due to the application of the Lorentz effect on mass transport at the catalyst’s surface. ii) We have also extensively explored the influence of electrostatics, as exhibited in self-assembling systems, by tribocharging. iii) Another key system involves surface tension effects; examples include interactions between heavy particles floating at a liquid-air interface, and interacting by changes in surface area; interactions of bubbles and bubble rafts, behaviors of bubble trains in microfluidic networks, and behaviors of microorganisms in constraining environments. iv) This work has intentionally de-emphasized biological systems; but it does include some work on protein-ligand interactions and interactions among microorganisms. v) We have also explored applications of some of these effects, these explorations include bubble rafts as diffraction gratings, exploration of the structures that can be obtained by tribocharging and uses of these structures in exploring nucleation and melting of crystals. vi) Although not a major focus of this work, several other topics have emerged and offer opportunities for future work. These include the behavior of bubble trains and bubble rafts in microfluidic systems. A particularly interesting example is the formation of bubble trains that repeat in the alteration of large and small bubbles according to rules we do not presently understand, but are uniquely large-period oscillating systems. These systems offer a new route into understanding the instabilities of the type represented by oscillations. vii) We have also begun exploratory projects on magnetic levitation (especially to determine molecular density), and information storage (in molecules). Magnetic Levitation: Self-assembling and biomimetic systems require both attraction and repulsion. We have used electrostatics (tribocharging), interfacial free-energies (surface tension and related forces) and others. Potential uses include reconfigurable diffraction gratings and liquid lenses; exploration of mechanisms in tribocharging; tunneling in EGaIn junctions; and bubble trains (especially in micro-fluidic systems). Examples of systems representing these topics is included in the following papers: Complexity: Disks rotating at a water-air interface; Benard-Marangoni effects; Vortex-Crystals from spinning magnetic disks (Marangoni effects); EGaIn Electrode to study quantum tunneling; Self-Assembly of electrostatically-charged metallic spheres (electrets); Dynamically reconfigurable lens; Using computational designs of ligands for enzymes; Electrostatic self-assembly by tribocharging; Monodisperse bubble trains in microchannel systems; Inverted dripping faucet; Flames; Printing of micro-organisms to regenerate the “ink” of printing device; Using micro-organisms to move loads (“microoxen”); Motion of bacterial swarms near surfaces; Making monodisperse particles in microfluidic systems; Coding/decoding of information stored in droplet trains in microfluidic networks; Magnetic levitation; and Information storage. i) Tribocharging. The change in focus of this work on electrets from the fundamentals of charging to applications of these materials in studying self-assembly using electrostatic interactions. ii) Bubbles in Microchannels. The realization that systems of bubbles in microchannels represented a major opportunity to study complexity in a very tractable system, and the development of a semi-quantitative theory of this subject. iii) Flames. The growth of “flames” remains an exploratory subject for the research, although their currently relatively little active work involving it ongoing. iv) Systems with Microorganisms. The removal of work in biological systems from this project. Based on work supported in this program, we now have a significant project on the development of microfluidic tools for studying C. elegans (a nematode), but this work was not appropriate for a program focused on complexity, and we developed separate support for it. (It is, however, an example of successful seeding of a new area by BES.) The work on electrets has gone through a period in which a part of the program was the subject of a MURI; the focus of this work was to develop materials that did not charge electrostatically on friction or contact. The MURI is now over, and the work on dynamic self-assembly (supported by BES) is the major focus. “Flames” has also enjoyed synergistic support, in terms of a project supported by DARPA on flame suppression (in the absence of extinguishing agents, using acoustic and electrostatic interactions). This work was helpful in understanding some of the basics of flames, but is entirely distinct from the BES focus in complexity. A growing interest is in the Lorentz effect. The Lorentz effect is the force exerted on charged particles (electrons, ions, charged molecules) when they move through a perpendicular magnetic field. The Lorentz effect is almost ubiquitous in modern technology: examples of applications include electric motors, dynamos, cathode ray tubes, many batteries, and most systems that control electrical currents with magnetic forces. We have begun to explore the Lorentz effect in electrochemical systems and heterogeneous catalytic systems involving charged organic species and inorganic ions. This work is still at an early stage, but initial studies that Lorentz effects can be large when ions move through magnetic fields, or magnetic fields move in the presence of ions.

36 MATERIALS SCIENCE↗
DOE OSTI2023

Metaoptics Enabled Multifunctional Imaging

Despite the fact that most microorganisms occupy two- and three-dimensional space in heterogeneous arrangements and in proximity to other microorganisms of different species, much of our current knowledge about their metabolic processes is derived from homogenizing, then analyzing, the chemical content of cultures containing a single microbial species. This project addressed this fundamental dichotomy by developing a set of optical imaging principles combining multiple new technologies. The imaging strategies developed in this project combine metaoptics structures with active in situ nanoscale control of the chemical environment and applied them to a microbial system, Myxococcus xanthus, with particular relevance to the DoE mission. The combination of metaoptical architectures and nanoscale control over the molecular environment enables: (a) precise control over the electromagnetic (EM) field at length scales smaller than the wavelength of light; (b) control of the interaction of the EM field with critical molecular systems in DOE relevant microbes; (c) control over the chemical environment – especially the presence and quantity of reactive oxygen species (ROS) that can affect redox homeostasis; and (d) the ability to ask new kinds of questions not accessible to ‘omics’ approaches or standard methods of biological imaging. These capabilities are applicable to detailed studies of metabolic pathways in microbes and to lignocellulosic biomass deconstruction. To accomplish these objectives, we pursued two over-arching technical goals: (1) the development of new metaoptics-enabled approaches to imaging and spectroscopic characterization; and (2) the development of tools to control the chemical environment of a microbial sample with nanometer-scale precision. Goal 1 was addressed through the design, fabrication, and characterization of new metasurfaces capable of super-resolution imaging through extreme confinement of the optical field. Goal 2 was addressed by controlling the redox potential on the nanoscale in microbial communities and characterizing their effect on intrinsic bacterial fluorophores which act as molecular sentinels and through characterization of soluble factors secreted by Myxococcus xanthus by confocal Raman imaging. The optical imaging/sensing approaches developed here make it possible to use these powerful new imaging and sensing modalities in metabolic studies by making it possible to visualize and track the spatial and temporal expression patterns of natural or engineered pathways in microorganisms.

09 BIOMASS FUELS↗
DOE OSTI2024

Engineering Methanogenic Microbiomes to Redirect Flux to Biomass

In this study, we present a method for acquiring and characterizing novel microbial consortia that regulates methanogens and methanotrophs through selective cultivation and metagenomic analysis of indigenous microorganisms in the environment. In addition, we present the work performed as part of this project to model the pathways that act as limiting factors in microbial methane metabolism based on a carbon cycle model. In this report, we describe the methods for selective cultivation of methane-metabolism-related microorganisms from environmental samples, the method for monitoring their methane consumption performance, and the method and results for verifying their functions using quantitative PCR and metagenomics techniques. The microbial consortia containing methanotrophs were obtained through selective cultivation and molecular biological verification, and their methane consumption performance was evaluated. In addition, the potential of the existence of bacteriophages interacting with methane metabolism-related microorganisms was identified through metagenomic sequencing.

09 BIOMASS FUELS↗
DOE OSTI2025

Acetate as a Platform for Carbon-Negative Production of Renewable Fuels and Chemicals (Final Technical Report)

This project was an industrial-academic collaboration between experts at the University of Wisconsin-Madison, the University of Kentucky, and LanzaTech, a world leader in the use of gas fermentation to sustainably produce fuels and chemicals. The project developed technologies to create an integrated process for converting carbon dioxide and renewable hydrogen into molecules that can be blended with liquid transportation fuels or used in an array of chemical applications. The project was motivated by the Program Objectives of eliminating carbon dioxide release in the production of chemicals by integrating the unique and efficient capabilities of two microorganisms into a single process. The first microbe, an acetogen, produces acetate from carbon dioxide and hydrogen while the second microbe upgrades acetate from acetogen fermentation permeates to higher-value chemical products. The carbon dioxide released in the upgrading process is recycled internally to produce more acetate. As such, the process can be designed to operate with zero carbon dioxide release and net positive carbon dioxide capture. The process has the potential to provide an alternative paradigm to the current bioeconomy – one in which acetate is the primary energy carrier instead of sugars. Our process by-passes photosynthesis and the barriers created by biomass as primary chemical feedstock. As such, the process can be scaled to meet existing sources of carbon dioxide emissions and located anywhere renewable hydrogen can be provided. Our work developed microorganisms with optimized metabolism for producing acetate and other microorganisms with improved conversion of acetate to dodecanol and dodecyl-acetate. We developed synthetic biology tools for a promising non-model bacterium that could enhance metabolic engineering efforts to convert acetate to chemical products. We conducted protein engineering studies to improve the activity of key enzymes involved in our metabolic pathways. We conducted a full technoeconomic analysis that set technical targets for each strain to meet economic goals. We identified key technical barriers in our process and proposed strategies to overcome them.

09 BIOMASS FUELS↗
DOE OSTI2025

Molecular mechanisms of mycorrhizal‐decomposer interactions and impacts on terrestrial biogeochemistry (Final Technical Report)

Soil microorganisms make nutrients available for plants and control a large portion of greenhouse gas emissions from terrestrial ecosystems by respiring CO 2 to the atmosphere, yet our understanding of how they do this is not clear enough to predict the rate of carbon (C) and nutrient flow through soils and the ecosystems they support. A common and abundant type of soil microorganism that associates with the roots of live plants, known as ectomycorrhizal fungi (EMF), is often able to control the amount of nutrients and C that are released from soil (e.g., as CO 2 respired to the atmosphere) during decomposition by microorganisms.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2025

EVT 16s Data and Large Supplementary Files

Soil microorganisms often interact to carry out decomposition of complex organic carbon and nitrogen compounds, such as chitin, but the high diversity and complexity of the soil microbiome and habitat has posed a challenge to elucidating such interactions between soil microorganisms. Here, we seek to address this challenge through analysis of a model soil consortium (MSC-2) of eight soil bacterial species. Our aim was to elucidate specific roles of the member species during chitin metabolism. Samples were collected from MSC-2 incubated in chitin-enriched soil over three months. Multi-omics was used to understand how the community composition, transcripts, proteins and chitin decomposition shifted over time. The data clearly and consistently revealed a temporal shift during chitin decomposition with defined contributions by individual species. A Streptomyces genus member (sp001905665) was a key player in early steps of chitin decomposition, with other MSC-2 members being central in carrying out later steps. These results illustrate how multi-omics applied to a defined consortium untangles interactions between soil microorganisms.

McClure, Ryan [Pacific Northwest National Laborato↗
DOE OSTI2020

FeGenie: a comprehensive tool for the identification of iron genes and iron gene neighborhoods in genomes and metagenome assemblies

Iron is a micronutrient for nearly all life on Earth. It can be used as an electron donor and electron acceptor by iron-oxidizing and iron-reducing microorganisms and is used in a variety of biological processes, including photosynthesis and respiration. While it is the fourth most abundant metal in the Earth’s crust, iron is often limiting for growth in oxic environments because it is readily oxidized and precipitated. Much of our understanding of how microorganisms compete for and utilize iron is based on laboratory experiments. However, the advent of next-generation sequencing and surge in publicly available sequence data has made it possible to probe the structure and function of microbial communities in the environment. To bridge the gap between our understanding of iron acquisition, iron redox cycling, iron storage, and magnetosome formation in model microorganisms and the plethora of sequence data available from environmental studies, we have created a comprehensive database of hidden Markov models (HMMs) based on genes related to iron acquisition, storage, and reduction/oxidation in Bacteria and Archaea. Along with this database, we present FeGenie, a bioinformatics tool that accepts genome and metagenome assemblies as input and uses our comprehensive HMM database to annotate provided datasets with respect to iron-related genes and gene neighborhood. An important contribution of this tool is the efficient identification of genes involved in iron oxidation and dissimilatory iron reduction, which have been largely overlooked by standard annotation pipelines. We validated FeGenie against a selected set of 28 isolate genomes and showcase its utility in exploring iron genes present in 27 metagenomes, 4 isolate genomes from human oral biofilms, and 17 genomes from candidate organisms, including members of the candidate phyla radiation. We show that FeGenie accurately identifies iron genes in isolates. Furthermore, analysis of metagenomes using FeGenie demonstrates that the iron gene repertoire and abundance of each environment is correlated with iron richness. While this tool will not replace the reliability of culture-dependent analyses of microbial physiology, it provides reliable predictions derived from the most up-to-date genetic markers. FeGenie’s database will be maintained and continually updated as new genes are discovered.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Microbial Phosphorus Mobilization Strategies Across a Natural Nutrient Limitation Gradient and Evidence for Linkage With Iron Solubilization Traits

Microorganisms have evolved several mechanisms to mobilize and mineralize occluded and insoluble phosphorus (P), thereby promoting plant growth in terrestrial ecosystems. However, the linkages between microbial P-solubilization traits and the preponderance of insoluble P in natural ecosystems are not well known. We tested the P solubilization traits of hundreds of culturable bacteria representative of the rhizosphere from a natural gradient where P concentration and bioavailability decline as soil becomes progressively more weathered. Aluminum, iron phosphate and organic P (phytate) were expected to dominate in more weathered soils. A defined cultivation medium with these chemical forms of P was used for isolation. A combination of soil chemical, spectroscopic analyses and 16S rRNA gene sequencing were used to understand the in situ ability for solubilization of these predominant forms of P. Locations with more occluded and organic P harbored the greatest abundance of P-mobilizing microorganisms, especially Burkholderiaceae ( Caballeronia and Paraburkholderia spp.). Nearly all bacteria utilized aluminum phosphate, however fewer could subsist on iron phosphate (FePO 4 ) or phytate. Microorganisms isolated from phytic acid were also most effective at solubilizing FePO 4 , suggesting that phytate solubilization may be linked to the ability to solubilize Fe. Significantly, we observed Fe to be co-located with P in organic patches in soil. Siderophore addition in lab experiments reinstated phytase mediated P-solubilization from Fe-phytate complexes. Taken together, these results indicate that metal-organic-P complex formation may limit enzymatic P solubilization from phytate in soil. Additionally, the linked traits of phytase and siderophore production were mostly restricted to specific clades within the Burkholderiaceae . We propose that Fe complexation of organic P (e.g., phytate) represents a major constraint on P turnover and availability in acidic soils, as only a limited subset of bacteria appear to possess the traits required to access this persistent pool of soil P.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Construction of nano slow-release systems for antibacterial active substances and its applications: A comprehensive review

At present, nano-carrier materials with antibacterial activity are of great significance. Due to the widespread resistance of many pathogenic microorganisms, it has seriously threatened human health. The natural antimicrobial substances extracted from fruits and vegetables can significantly improve their stability combined with nano-carrier materials. The resistance of pathogenic microorganisms will be substantially reduced, greatly enhancing the effect of active antimicrobial substances. Nanotechnology has excellent research prospects in the food industry, antibacterial preservation, food additives, food packaging, and other fields. This paper introduces nano-carrier materials and preparation techniques for loading and encapsulating active antibacterial substances in detail by constructing a nano-release system for active antibacterial substances. The antibacterial effect can be achieved by protecting them from adverse external conditions and destroying the membrane of pathogenic microorganisms. The mechanism of the slow release of the bacteriostatic active substance is also described. The mechanism of carrier loading and release is mainly through non-covalent forces between the bacteriostatic active substance and the carrier material, such as hydrogen bonding, π-π stacking, van der Waals forces, electrostatic interactions, etc., as well as the loading and adsorption of the bacteriostatic active substance by the chemical assembly. Finally, its wide application in food and medicine is introduced. It is hoped to provide a theoretical basis and technical support for the efficient utilization and product development of bacteriostatic active substances.

Cao, Jiayong↗
DOE OSTI2022

Synthetic Soil Aggregates: Bioprinted Habitats for High-Throughput Microbial Metaphenomics

The dynamics of microbial processes are difficult to study in natural soil, owing to the small spatial scales on which microorganisms operate and to the opacity and chemical complexity of the soil habitat. To circumvent these challenges, we have created a 3D-bioprinted habitat that mimics aspects of natural soil aggregates while providing a chemically defined and translucent alternative culturing method for soil microorganisms. Our Synthetic Soil Aggregates (SSAs) retain the porosity, permeability, and patchy resource distribution of natural soil aggregates—parameters that are expected to influence emergent microbial community interactions. We demonstrate the printability and viability of several different microorganisms within SSAs and show how the SSAs can be integrated into a multi-omics workflow for single SSA resolution genomics, metabolomics, proteomics, lipidomics, and biogeochemical assays. We study the impact of the structured habitat on the distribution of a model co-culture microbial community and find that it is significantly different from the spatial organization of the same community in liquid culture, indicating a potential for SSAs to reproduce naturally occurring emergent community phenotypes. The SSAs have the potential as a tool to help researchers quantify microbial scale processes in situ and achieve high-resolution data from the interplay between environmental properties and microbial ecology.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Extracellular electron transfer increases fermentation in lactic acid bacteria via a hybrid metabolism

Energy conservation in microorganisms is classically categorized into respiration and fermentation; however, recent work shows some species can use mixed or alternative bioenergetic strategies. We explored the use of extracellular electron transfer for energy conservation in diverse lactic acid bacteria (LAB), microorganisms that mainly rely on fermentative metabolism and are important in food fermentations. The LAB Lactiplantibacillus plantarum uses extracellular electron transfer to increase its NAD + /NADH ratio, generate more ATP through substrate-level phosphorylation, and accumulate biomass more rapidly. This novel, hybrid metabolism is dependent on a type-II NADH dehydrogenase (Ndh2) and conditionally requires a flavin-binding extracellular lipoprotein (PplA) under laboratory conditions. It confers increased fermentation product yield, metabolic flux, and environmental acidification in laboratory media and during kale juice fermentation. The discovery of a single pathway that simultaneously blends features of fermentation and respiration in a primarily fermentative microorganism expands our knowledge of energy conservation and provides immediate biotechnology applications.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Signatures of prescribed fire in the microbial communities of Cornus florida are largely undetectable five months post-fire

Prescribed burn is a management tool that influences the physical structure and composition of forest plant communities and their associated microorganisms. Plant-associated microorganisms aid in host plant disease tolerance and increase nutrient availability. The effects of prescribed burn on microorganisms associated with native ecologically and economically important tree species, such as Cornus florida L. (flowering dogwood), are not well understood, particularly in aboveground plant tissues (e.g., leaf, stem, and bark tissues). The objective of this study was to use 16S rRNA gene and ITS2 region sequencing to evaluate changes in bacterial and fungal communities of five different flowering dogwood-associated niches (soil, roots, bark, stem, and leaves) five months following a prescribed burn treatment. The alpha- and beta-diversity of root bacterial/archaeal communities differed significantly between prescribed burn and unburned control-treated trees. In these bacterial/archaeal root communities, we also detected a significantly higher relative abundance of sequences identified as Acidothermaceae, a family of thermophilic bacteria. No significant differences were detected between prescribed burn-treated and unburned control trees in bulk soils or bark, stem, or leaf tissues. The findings of our study suggest that prescribed burn does not significantly alter the aboveground plant-associated microbial communities of flowering dogwood trees five months following the prescribed burn application. Further studies are required to better understand the short- and long-term effects of prescribed burns on the microbial communities of forest trees.

16S rRNA↗
DOE OSTI2022

DOE new players Carbon cycle (2016-2021)

The overarching scientific goals of our multidisciplinary grant was to further expand understanding about the key microorganisms (players), metabolic strategies (processes), and interspecies relationships (interactions) involved in the formation and oxidation of methane in the environment. This research applied novel environmental metagenomics, transcriptomics, and proteomic techniques, state-of-the-art analytical imaging, stable isotope geochemistry, and reaction-transport modeling to address these goals and develop an ‘ecosystems level’ understanding of the factors which regulate microbial methane cycling in anoxic sedimentary ecosystems. For decades, it was believed that the obligate step in methanogenesis catalyzed by methyl coenzyme M reductase (Mcr) was limited to a specific branch of the archaeal Domain, formerly known as the Euryarchaeota. Less than a decade ago co-I Tyson’s team published a surprising metagenomic-based discovery of divergent Mcr genes in a novel uncultured phylum (Bathyarchaeota), catalyzing a major shift in thinking about the diversity of microorganisms that encode the potential for methane (or higher alkane) metabolism in anoxic environments (Evans et al., 2015). In our work here, we further expand on the groups of archaea harboring the genomic potential for methane or hydrocarbon metabolism using environmental metagenomics and new gene targeted bioinformatics techniques. We additionally advanced understanding about the terminal electron acceptors and metabolic potential supporting the anaerobic oxidation of methane (AOM) in terrestrial ecosystems, specifically focused on new lineages of ANME archaea capable of respiring manganese oxides with methane presumably using large extracellular multi-heme cytochrome complexes. New details about specific syntrophic mechanisms underlying the exchange of electrons during sulfate-coupled methane oxidation between ANME-2 archaea and their sulfate-reducing bacterial partners were also elucidated as part of this funded project. Through a series of experimental ‘omics and single cell stable isotope probing studies with incubated environmental sediment samples and a cultured model electrogenic microorganism combined with model-based predictions. Combined, this work provides strong support for the hypothesis of direct interspecies electron transfer (DIET) is the dominant syntrophic mechanism controlling the anaerobic oxidation of methane with sulfate over other proposed mechanisms and additionally illustrates important spatial constraints and the underlying physico-chemical factors influencing AOM syntrophic consortia structure for DIET and extracellular metal respiration. This collaborative multi-institutional project successfully advanced several of our milestone goals including the identification of new microbial players containing methyl coenzyme M reductases hypothesized to be central to methane or hydrocarbon cycling in anoxic environments and enhancing fundamental knowledge about the role extracellular electron transfer plays in the ecophysiology of methanotrophic archaea respiring metal oxides and in the physical and metabolic structuring of syntrophic interactions in methane-rich sedimentary ecosystems.

03 NATURAL GAS↗
DOE OSTI2022

Biogeochemical Characteristics of Earth's Volcanic Permafrost: An Analog of Extraterrestrial Environments

This article describes a study of frozen volcanic deposits collected from volcanoes Tolbachik and Bezymianny on the Kamchatka Peninsula, Russia, and Deception Island volcano, Antarctica. In addition, we studied suprasnow ash layers deposited after the 2007 eruptions of volcanoes Shiveluch and Bezymianny on Kamchatka. The main objectives were to characterize the presence and survivability of thermophilic microorganisms in perennially frozen volcanic deposits. As opposed to permafrost from the polar regions, viable thermophiles were detected in volcanic permafrost by cultivation, microscopy, and sequencing. In the permafrost of Tolbachik volcano, we observed methane formation by both psychrophilic and thermophilic methanogenic archaea, while at 37°C, methane production was noticeably lower. Thermophilic bacteria isolated from volcanic permafrost from the Deception Island were 99.93% related to Geobacillus stearothermophilus. Furthermore, our data showed biological sulfur reduction to sulfide at 85°C and even at 130°C, where hyperthermophilic archaea of the genus Thermoproteus were registered. Sequences of hyperthermophilic bacteria of the genus Caldicellulosiruptor were discovered in clone libraries from fresh volcanic ash deposited on snow. Microorganisms found in volcanic terrestrial permafrost may serve as a model for the alien inhabitants of Mars, a cryogenic planet with numerous volcanoes. Thermophiles and hyperthermophiles and their metabolic processes represent a guideline for the future exploration missions on Mars.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Microbial Community Structure and Ecological Networks during Simulation of Diatom Sinking

Microbial-mediated utilization of particulate organic matter (POM) during its downward transport from the surface to the deep ocean constitutes a critical component of the global ocean carbon cycle. However, it remains unclear as to how high hydrostatic pressure (HHP) and low temperature (LT) with the sinking particles affects community structure and network interactions of the particle-attached microorganisms (PAM) and those free-living microorganisms (FLM) in the surrounding water. In this study, we investigated microbial succession and network interactions in experiments simulating POM sinking in the ocean. Diatom-derived 13C- and 12C-labeled POM were used to incubate surface water microbial communities from the East China Sea (ECS) under pressure (temperature) of 0.1 (25 °C), 20 (4 °C), and 40 (4 °C) MPa (megapascal). Our results show that the diversity and species richness of the PAM and FLM communities decreased significantly with HHP and LT. Microbial community analysis indicated an increase in the relative abundance of Bacteroidetes at high pressure (40 MPa), mostly at the expense of Gammaproteobacteria, Alphaproteobacteria, and Gracilibacteria at atmospheric pressure. Hydrostatic pressure and temperature affected lifestyle preferences between particle-attached (PA) and free-living (FL) microbes. Ecological network analysis showed that HHP and LT enhanced microbial network interactions and resulted in higher vulnerability to networks of the PAM communities and more resilience of those of the FLM communities. Most interestingly, the PAM communities occupied most of the module hubs of the networks, whereas the FLM communities mainly served as connectors of the modules, suggesting their different ecological roles of the two groups of microbes. These results provided novel insights into how HHP and LT affected microbial community dynamics, ecological networks during POM sinking, and the implications for carbon cycling in the ocean.

59 BASIC BIOLOGICAL SCIENCES↗