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All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Editorial: Cell Free Biocatalysis for the Production of Bioproducts

Biological approaches show promise for the sustainable and cost competitive production of commodity fuels and chemicals. A lot of efforts have been devoted to engineering microbial chassis to produce select fuels and chemicals. These approaches have had some level of success exemplified, of course, by the production of ethanol at high yield and titer but also recent successes with commodity chemicals such as 1,4-butanediol, 1,3-propanediol, and even to some extent with sustainable aviation fuel intermediates such as farnesene. However, many of these processes still suffer from low yields, usually below the standard needed to drive a sustainable and atom efficient bioeconomy. Additionally, for many other chemicals and fuel precursors, titers and yields are only a fraction of what they are for ethanol. Indeed, there exist many limiting factors associated with the use of microbial biocatalysts. Some of these factors include the diversion of carbon from products to sustain growth and biomass formation, the production of byproducts or regulatory mechanisms, the toxicity of end products or intermediates, and the difficulty of separating products from growth media.

09 BIOMASS FUELS↗
DOE OSTI2022

Spectroscopy-based isotopic (δ 13 C) analysis for high spatial resolution of carbon exchange in the rhizosphere

The rhizosphere is a highly dynamic zone bridging plant roots with needed nutrient resources in soil. While the rhizosphere may be small, it has a disproportionally large impact on plant success and biomass production. A suite of rhizosphere-hosted microbial and geochemical interactions facilitate nutrient acquisition by plant roots, and, in turn, the roots stimulate these processes by supplying organic carbon into the rhizosphere. The small physical dimensions of the rhizosphere, however, can constrain efforts to elucidate key carbon exchange processes and their spatial extent and localization. We present a method for spatially resolved δ 13 C analysis of rhizosphere samples by coupling laser ablation (LA) sampling with isotopic analysis using capillary absorption spectroscopy (CAS) which differs from conventional mass spectrometer (MS) approaches. The CAS system has high sensitivity (requires fewer nanomoles of CO 2 per analysis) than comparable MS systems, which enables reduced sample size requirements to thereby improve spatial resolution (from 25 μm to as low as a projected 5 μm spatial resolution). We demonstrate the utility of CAS using rhizosphere samples from switchgrass plants exposed to 13 CO 2 . As a result, this technique will provide a capability for tracking the extent and spatial distribution of root exudate into the rhizosphere at highly detailed spatial scales.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1971

Microbiologic assay of space hardware.

Review of the procedures used in the microbiological examination of space hardware. The general procedure for enumerating aerobic and anaerobic microorganisms and spores is outlined. Culture media and temperature-time cycles used for incubation are reviewed, along with assay systems designed for the enumeration of aerobic and anaerobic spores. The special problems which are discussed are involved in the precise and accurate enumeration of microorganisms on surfaces and in the neutralization of viable organisms buried inside solid materials that could be released to a planet's surface if the solid should be fractured. Special attention is given to sampling procedures including also the indirect techniques of surface assays of space hardware such as those using detachable or fallout strips. Some data on comparative levels of microbial contamination on lunar and planetary spacecraft are presented.

Favero, M. S.↗
NASA NTRSDate not supplied

BioNutrients-2: Improvements to the BioNutrients-1 Nutrient Production System

The BioNutrients (BN) project aims to develop an on-demand biological production system capable of rapid and safe delivery of multiple nutrients in single-use production packs for long-duration missions. The general concept of this system is to develop edible engineered microorganisms that produce nutrients for human consumption following long-term storage, revival, and growth. BN-1 is a five-year flight project currently on the International Space Station (ISS). It is testing the long-term storage of various microorganisms for the biomanufacturing of space-relevant compounds and validating the performance of the first generation of production packs. The BN-2 project expands the BN-1 flight project scope by introducing novel products, broadening the range of microbial food sources, and improving upon production pack hardware. We are optimizing the biomanufacturing platform for in situ production of yogurt and kefir on the ISS. BN-2 also will test engineered microorganisms, including a probiotic strain, to produce carotenoids or follistatin. To improve the feasibility of this technology for mission use, we transitioned from a hard-shell pack to a second-generation fluorinated ethylene propylene (FEP) bag. The single-use FEP bioreactor facilitates growth of target organisms in a contaminant free environment, while reducing mass and volume compared to the BN-1 hardware. These bioreactors will be employed for production of additional space-relevant products in the BN-2 project. On-orbit testing of FEP production packs will enable optimization of the platform for eventual flight use by the crew.

Natalie Ball↗
NASA NTRS2012

Genotypic & Phenotypic Diversity of Microbial Isolates from the Mars Exploration Rovers

Mars-bound rovers such as the Mars Exploration Rover (MER) endure strict planetary protection implementation campaigns to assess bioburden. The objective of this study is to identify cultivable microorganisms isolated by the NASA Standard Assay from spacecraft during pre-launch and evaluate their potential to survive conditions on the Martian surface. Of approximately 350 isolates collected from the MER spacecraft archive, 171 microorganisms were reconstituted for characterization via 16S rRNA fingerprinting. Alignment of 16S sequences revealed high levels of sequence similarity to spore-forming species, overwhelmingly of the genera Bacillus (73.7%) and Paenibacillus (14.0%). Samples underwent phenotype characterization employing multiple carbon sources and ion concentrations in an automated microarray format using the Omnilog system. Working and stock cultures were prepared to address the immediate needs for day-to-day culture utilization and long-term preservation, respectively. Results from this study produced details about the microbes that contaminate surfaces of spacecraft, as well as a preliminary evaluation of a rapid biochemical ID method that also provides a phenotypic assessment of contaminants. The overall outcome of this study will benefit emerging cleaning and sterilization technologies for preventing forward contamination that could negatively impact future life detection or sample return missions.

Mars Exploration Rovers↗
DOE OSTI2021

Soil and Water Chemistry and Trace Metal Extractability and Speciation in Wetland Soils from Illinois and South Carolina and Stream Sediments from Tennessee

Dataset revised on October 15, 2021. This revision adds sulfur and iron X-ray absorption near-edge structure spectra for the wetland soils and stream sediments from the field areas. It also renames the sample locations in a way that is more intuitive to readers of the companion paper that is under review. Finally, the data filenames and organization have been updated in their labeling to parallel the data sources in the associated paper. The abstract text and methods were also revised to reflect the data that was added to the dataset.Trace metals are essential for microbially-mediated biogeochemical processes occurring in anoxic wetland soils and stream bed sediments, such as denitrification, methanogenesis, and mercury methylation. Low availability of these elements may potentially inhibit key components of anaerobic carbon and nitrogen cycling and contaminant transformation. The solid-phase speciation of trace metals likely plays an important role in controlling their bioavailability. Metal speciation is well studied in contaminated soils and sediments as well as those naturally elevated in trace metals. However, less is known regarding the chemical forms of trace metals in systems having concentrations similar to geological background levels, the very settings where metal limitations may be most prevalent. We have investigated trace metal concentrations, extractability, and solid-phase speciation in three freshwater subsurface aquatic systems: marsh wetland soils, riparian wetland soils, and the sediments of a streambed.Data are provided for marsh wetland soils at Argonne National Laboratory, riparian wetland soils in the Tims Branch watershed at Savannah River National Laboratory, and stream bed sediments from East Fork Poplar Creek near Oak Ridge National Laboratory. Soil and sediment elemental abundances, mineralogy, and extractable nutrients as well as dissolved major elements, anions, trace metals, and nutrients in the overlying surface waters are provided. In addition, the results of sequential chemical extraction for the trace metals cobalt, nickel, copper, and zinc from the soils and sediment are reported as well as X-ray absorption near-edge structure (XANES) spectra in these materials are reported. To aid interpretation of these data, XANES spectra of sulfur in the soils and sediments as well as both XANES and extended X-ray absorption fine structure (EXAFS) spectra of iron in these materials are reported. The data package also includes the XANES spectra of reference standards and a potential interferent in the measurements. All data are provided in text-based CSV format with header sections indicating the data contained in each file and the corresponding units. Note that "u" is used in place of Greek lower case mu to indicate the micro prefix on units.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2020

Modeling Functional Organic Chemistry in Arctic Rivers: An Idealized Siberian System

Rivers of the Arctic will become ever more important for the global climate, since they carry a majority of continental dissolved organic carbon flux into the rapidly changing polar ocean. Aqueous organics comprise a wide array of functional groups, several of which are likely to impact coastal and open water biophysical properties. Light attenuation, interfacial films, aerosol formation, gas release and momentum exchange can all be cited. We performed Lagrangian kinetic modeling for the evolution of riverine organic chemistry as the molecules in question make their way from the highlands to Arctic outlets. Classes as diverse as the proteins, sugars, lipids, re-condensates, humics, bio-tracers and small volatiles are all included. Our reduced framework constitutes an idealized northward flow driving a major hydrological discharge rate and primarily representing the Russian Lena. Mountainous, high solute and tundra sources are all simulated, and they meet up at several points between soil and delta process reactors. Turnover rates are parameterized beginning with extrapolated coastal values imposed along a limited tributary network, with connections between different terrestrial sub-ecologies. Temporal variation of our total dissolved matter most closely resembles the observations when we focus on the restricted removal and low initial carbon loads, suggesting relatively slow transformation along the water course. Thus, channel combinations and mixing must play a dominant role. Nevertheless, microbial and photochemical losses help determine the final concentrations for most species. Chemical evolution is distinct for the various functionalities, with special contributions from pre- and post-reactivity in soil and delta waters. Several functions are combined linearly to represent the collective chromophoric dissolved matter, characterized here by its absorption. Tributaries carry the signature of lignin phenols to segregate tundra versus taiga sources, and special attention is paid to the early then marine behaviors of low molecular weight volatiles. Heteropolycondensates comprise the largest percentage of reactive carbon in our simulations due to recombination/accumulation, and they tend to be preeminent at the mouth. Outlet concentrations of individual structures such as amino acids and absorbers lie above threshold values for biophysical influence, on the monolayer and light attenuation. The extent of coastal spreading is examined through targeted regional box modeling, relying on salinity and color for calibration. In some cases, plumes reach the scale of peripheral arctic seas, and amplification is expected during upcoming decades. Conclusions are mapped from the Lena to other boreal discharges, and future research questions are outlined regarding the bonding type versus mass release as permafrost degrades. Dynamic aqueous organic coupling is recommended for polar system models, from headwaters to coastal diluent.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2020

Direct Injection of Biomineralizing Agents to Restore Injectivity and Wellbore Integrity

Summary In this manuscript, we describe the second of two field demonstrations of microbially induced calcium carbonate precipitation (MICP) performed in a failed waterflood injection well in Indiana. In 2012, fracture-related flow pathways developed in the wellbore cement, causing injection water to bypass the oil-bearing formation and enter a high-permeability sandstone thief zone, thereby substantially decreasing injection pressure. In the first field demonstration, our study team characterized the well's mode of failure and successfully applied MICP to decrease flow through the defective cement. However, because the MICP treatment was conducted using a bailer delivery system, the degree of permeability reduction achievable was not adequate to fully restore the historic injection pressure of 1,400 psi at 1 gal/min. For the second field demonstration (reported herein), a direct injection system was developed that substantially increased the injection volume of MICP-promoting fluids. Two strategies were implemented to produce more ureolytic microbes: resuspending concentrated frozen cells immediately before injection and scaling up the bioreactor growth capacity. Multiple pulses of microbes and urea-calcium media were pumped into a string of 1-in.-diameter tubing separated by brine spacers and injected continuously at a flow rate of 3.4 to 1.4 gal/min. During the third day of injection, an injection pressure of 1,384 psi at a flow rate of 1.4 gal/min was achieved, and the experiment was terminated. This study demonstrates that MICP can be successfully used in large-volume applications where the time frame for the delivery of reactants is limited. This finding has significant relevance for commercialization of the MICP biotechnology in the oil and gas industry.

Engineering↗
NASA NTRS1994

Use of lunar regolith as a substrate for plant growth

Regenerative Life Support Systems (RLSS) will be required to regenerate air, water, and wastes, and to produce food for human consumption during long-duration missions to the Moon and Mars. It may be possible to supplement some of the materials needed for a lunar RLSS from resources on the Moon. Natural materials at the lunar surface may be used for a variety of lunar RLSS needs, including (1) soils or solid-support substrates for plant growth, (2) sources for extraction of essential, plant-growth nutrients, (3) substrates for microbial populations in the degradation of wastes, (4) sources of O2 and H2, which may be used to manufacture water, (5) feed stock materials for the synthesis of useful minerals (e.g., molecular sieves), and (6) shielding materials surrounding the outpost structure to protect humans, plants, and microorganisms from harmful radiation. Use of indigenous lunar regolith as a terrestrial-like soil for plant growth could offer a solid support substrate, buffering capacity, nutrient source/storage/retention capabilities, and should be relatively easy to maintain. The lunar regolith could, with a suitable microbial population, play a role in waste renovation; much like terrestrial waste application directly on soils. Issues associated with potentially toxic elements, pH, nutrient availability, air and fluid movement parameters, and cation exchange capacity of lunar regolith need to be addressed before lunar materials can be used effectively as soils for plant growth.

Ming, D. W.↗
NASA NTRS1999

Post-Viking microbiology: new approaches, new data, new insights

In the 20 years since the Viking experiments, major advances have been made in the areas of microbial systematics, microbial metabolism, microbial survival capacity, and the definition of environments on earth, suggesting that life is more versatile and tenacious than was previously appreciated. Almost all niches on earth which have available energy, and which are compatible with the chemistry of carbon-carbon bonds, are known to be inhabited by bacteria. The oldest known bacteria on earth apparently evolved soon after the formation of the planet, and are heat loving, hydrogen and/or sulfur metabolizing forms. Among the two microbial domains (kingdoms) is a great deal of metabolic diversity, with members of these forms being able to grow on almost any known energy source, organic or inorganic, and to utilize an impressive array of electron acceptors for anaerobic respiration. Both hydrothermal environments and the deep subsurface environments have been shown to support large populations of bacteria, growing on energy supplied by geothermal energy, thus isolating these ecosystems from the rest of the global biogeochemical cycles. This knowledge, coupled with new insights into the history of the solar system, allow one to speculate on possible evolution and survival of life forms on Mars.

Review↗
DOE OSTI2021

Warm-season net CO 2 uptake outweighs cold-season emissions over Alaskan North Slope tundra under current and RCP8.5 climate

Abstract Arctic warming has increased vegetation growth and soil respiration during recent decades. The rate of Arctic warming will likely amplify over the 21st century. Previous studies have revealed that the most severe Arctic warming occurred during the cold season (September to May). The cold-season warming has posited significant CO 2 emissions to the atmosphere via respiration, possibly offsetting warm-season (June to August) net CO 2 uptake. However, prevailing Earth system land models poorly represent cold-season CO 2 emissions, making estimates of Arctic tundra annual CO 2 budgets highly uncertain. Here, we demonstrate that an improved version of the energy exascale Earth system model (E3SM) land model (ELMv1-ECA) captures the large amount of cold-season CO 2 emissions over Alaskan Arctic tundra as reported by two independent, observationally-constrained datasets. We found that the recent seven-decades warming trend of cold-season soil temperature is three times that of the warm-season. The climate sensitivity of warm-season net CO 2 uptake, however, is threefold higher than for the cold-season net CO 2 loss, mainly due to stronger plant resilience than microbial resilience to hydroclimatic extremes. Consequently, the modeled warm-season net CO 2 uptake has a larger positive trend (0.74 ± 0.14 gC m −2 yr −1 ) than that of cold-season CO 2 emissions (0.64 ± 0.11 gC m −2 yr −1 ) from 1950 to 2017, supported by enhanced plant nutrient uptake and increased light- and water-use efficiency. With continued warming and elevated CO 2 concentrations under the representative concentration pathway (RCP) 8.5 scenario, the increasing rate of warm-season net CO 2 uptake is more than twice the rate of cold-season emissions (1.33 ± 0.32 gC m −2 yr −1 vs 0.50 ± 0.12 gC m −2 yr −1 ), making the modeled Alaskan Arctic tundra ecosystem a net CO 2 sink by 2100. However, other geomorphological and ecological disturbances (e.g. abrupt permafrost thaw, thermokarst development, landscape-scale hydrological changes, wildfire, and insects) that are not considered here might alter our conclusion.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2024

Integrating Chemical Catalysis and Biological Conversion of Carbon Intermediates for Deriving Value-Added Products from Carbon Dioxide

Carbon dioxide valorization represents an appealing approach to reducing greenhouse gases in the atmosphere. While electrocatalysis is an effective tool to reduce CO 2 into small carbon compounds, it becomes increasingly challenging to efficiently produce compounds with more carbon atoms. In contrast, while biological systems struggle to utilize CO 2 , they can readily upcycle other small carbon compounds. This project explores the use of a two-stage process that electrocatalytically converts CO 2 into methanol, formate, or acetate which is subsequently utilized by Methylotuvimicrobium alcaliphilum 20Z to produce medium chain length polyhydroxyalkanoate. A techno-economic analysis and life cycle assessment evaluates the commercial viability of the process as well as its carbon emissions. We show here an enhanced CO 2 -to-methanol electroconversion step coupled with the use of a microbial culture adapted to the process conditions to be the optimal configuration for economic potential.

09 BIOMASS FUELS↗
NASA NTRS1992

Long-term lunar stations: Some ecological considerations

A major factor for long-term success of a lunar station is the ability to keep an agroecosystem functioning at a desirable, stable steady-state with ecological stability and reliability. Design for a long-lived extraterrestrial manned station must take into account interactions among its subsystems to insure that overall functionality is enhanced (or at least not compromised). Physical isolation of feed production, human living areas, recycling, and other systems may be straightforward, however, microbiological isolation will be very difficult. While it is possible to eliminate plant-associated microbiological communities by growing the plants asepticallly, it is not practical to keep plants germ-free on a large scale if humans are working with them. Ecological theory strongly suggests that some kinds of communities or organisms effectively increase the stability of ecosystems and will protect the plants from potential pathogens. A carefully designed and maintained (lunar-derived) soil can provide a variety of habitats for effective microbial buffers while adding structure to the agroecosystem. A soil can also increase ecosystem reliability through buffering otherwise large element and compound fluctuations (of nutrients, wastes, etc.) as well as buffering temperature level and atmosphere composition. We are doing experiments in ecological dynamics and attempting to extend the relevant theories.

Maguire, Bassett, Jr.↗
DOE OSTI2025

Agricultural Management Legacy Effects on Switchgrass Growth and Soil Carbon Gains

Switchgrass ( Panicum virgatum L.) is a native North American grass currently considered a high-potential bioenergy feedstock crop. However, previous reports questioned its effectiveness in generating soil organic carbon (SOC) gains, with resultant uncertainty regarding the monoculture switchgrass's impact on the environmental sustainability of bioenergy agriculture. We hypothesize that the inconsistencies in past SOC accrual results might be due, in part, to differences in prior land management among the systems subsequently planted to switchgrass. To test this hypothesis, we measured SOC and other soil properties, root biomass, and switchgrass growth in an experimental site with a 30-year history of contrasting tillage and N-fertilization treatments, 7 years after switchgrass establishment. We determined switchgrass' monthly gross primary production (GPP) for six consecutive years and conducted deep soil sampling. Nitrogen fertilization expectedly stimulated switchgrass growth; however, a tendency for better plant growth was also observed under unfertilized settings in the former no-till soil. In topsoil, SOC significantly increased from 2007 to 2023 in fertilized treatments of both tillage histories, with the greatest increase observed in fertilized no-till. Fertilized no-till also had the highest particulate organic matter content in the topsoil, with no differences among the treatments observed in deeper soil layers. However, regardless of fertilization, the tillage history had a strong effect on stratification with depth of SOC, total N, and microbial biomass C. Results suggested that historic and ongoing N fertilization had a substantial impact on switchgrass growth and soil characteristics, while tillage legacy had a much weaker, but still discernible, effect.

bioenergy crop production system↗
NASA NTRS1973

Apollo experience report: Potable water system

A description of the design and function of the Apollo potable water system is presented. The command module potable water is supplied as a byproduct of the fuel cells. The cells, located in the service module, function primarily to supply electrical energy to the spacecraft. The source of the lunar module potable water is three tanks, which are filled before lift-off. The technique of supplying the water in each of these cases and the problems associated with materials compatibility are described. The chemical and microbiological quality of the water is reviewed, as are efforts to maintain the water in a microbially safe condition for drinking and food mixing.

Sauer, R. L.↗
DOE OSTI2026

Optimizing Cryo-Focused Pyrolysis GC/MS for Tracing Soil Organic Matter Across Diverse Ecosystems

The cycling of organic matter in terrestrial soils and sediments is central to a range of biogeochemical processes that regulate nutrient cycling, crop productivity, trace gas emissions, and contaminant transport. Pyrolysis-gas chromatography/mass spectrometry (py-GC/MS) is a powerful tool for characterizing bulk soil organic matter (SOM) at the molecular level. In this study, we used a cryo-focused py-GC/MS system to analyze soil samples from seven diverse ecosystems: vernal pool, prairie pothole, temperate forest, tropical forest, tundra, wildfire-affected boreal forest, and grassland. We addressed a key bottleneck in molecular-level SOM characterization by developing an automated data analysis pipeline to optimize py-GC/MS and complementary evolved gas analysis/mass spectrometry (EGA/MS) methods, incorporating advanced tools for peak deconvolution, developing a custom compound class library, and implementing fragmentation spectrum-based molecular networking for the first time. This improved workflow was applied to soil samples from all seven ecosystems, including multiple depths and density fractions. Our findings demonstrate that ecosystem type plays a dominant role in shaping compositional differences in SOM. We also identified trends in the source of SOM compounds (e.g., microbial vs plantderived) across soil depth and density fractions, which are critical for understanding persistence and turnover of SOM. Our molecular networking analysis indicated that although many compounds are widespread across ecosystems, others are restricted to specific environments, such as wetlands. This underscores the utility of molecular-level data in elucidating the complexity of SOM composition and the environmental drivers that shape it. Such molecular-level insights can deepen our knowledge of biogeochemical SOM cycles.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Microbial contribution to post-fire tundra ecosystem recovery over the 21st century

Abstract Tundra ecosystems have experienced an increased frequency of fire, and this trend is predicted to continue throughout the 21 st Century. Post-fire recovery is underpinned by complex interactions between microbial functional groups that drive nutrient cycling. Here we use a mechanistic model to demonstrate an acceleration of the nitrogen cycle post-fire driven by changes in niche space and microbial competitive dynamics. We show that over the first 5-years post-fire, fast-growing bacterial heterotrophs colonize regions of the soil previously occupied by slower-growing saprotrophic fungi. The bacterial heterotrophs mineralize organic matter, releasing nutrients into the soil. This pathway outweighs new sources of nitrogen and facilitates the recovery of plant productivity. We broadly show here that while consideration of distinct microbial metabolisms related to carbon and nutrient cycling remains rare in terrestrial ecosystem models, they are important when considering the rate of ecosystem recovery post-disturbance and the feedback to soil nutrient cycles on centennial timescales.

54 ENVIRONMENTAL SCIENCES↗