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All sourcesNASA NTRSDOE OSTI
DOE OSTI2020

Partners for life: building microbial consortia for the future

We report new technologies have allowed researchers to better design, build, and analyze complex consortia. These developments are fueling a wider implementation of consortium-based bioprocessing by leveraging synthetic biology, delivering on the field’s multitudinous promises of higher efficiencies, superior resiliency, augmented capabilities, and modular bioprocessing. Here we chronicle current progress by presenting a range of screening, computational, and biomolecular tools enabling robust population control, efficient division of labor, and programmatic spatial organization; furthermore, we detail corresponding advancements in areas including machine learning, biocontainment, and standardization. Additionally, we show applications in myriad sectors, including medicine, energy and waste sustainability, chemical production, agriculture, and biosensors. Concluding remarks outline areas of growth that will promote the utilization of complex community structures across the biotechnology spectrum.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

A Systems Approach to Increasing Carbon Flux to Seed Oil for Biofuels and Bioproducts Production in Camelina sativa (Final Report)

To combat climate change and alleviate the dependency of the United States on fossil fuels, the transition to biofuel crops has long been proposed as a crucial part of the long-term solution. Camelina sativa has emerged as one of the leading commercially viable options for biofuel and bioproduct production for the U.S. Camelina has the advantages of low agronomic inputs and natural resistance to diverse biotic and abiotic stresses relative to other oilseed crops, and Camelina oil-based blends have been tested and approved as liquid transportation fuels. A major limitation in the widespread adoption of Camelina as a viable industrial oilseed crop is its modest oil yields. This project directly investigated possible paths towards increasing oil, by employing tissue-specific and whole plant systems approaches to identify major regulatory bottlenecks. We developed a new high-throughput method for the identification of multi-gene transformants in polyploid species like Camelina sativa and used this to determine that, contrary to what mathematical models suggested, introduction of the microbial Entner-Doudoroff (ED) pathway into Camelina sativa did not result in significant seed oil increases. We developed flux maps providing accurate and statistically robust models of central carbon metabolism in Camelina, including of cultured Camelina embryos. This knowledge was used in the capacitation of a number of junior researchers in gaining knowledge on approaches used for quantitative flux map analyses. We identified several novel Camelina transcription factor genes regulating fatty acid biosynthesis that showed variable effects on seed oil accumulation in transgenic plants. Among a number of community resources, we also developed a knowledge base web resource (CamRegBase) to integrate Camelina gene regulatory information. Together, research outcomes from this project have contributed to a much better understanding of the regulation and bottlenecks to engineer seed oil production in Camelina.

09 BIOMASS FUELS↗
DOE OSTI2023

Conservation of beneficial microbes between the rhizosphere and the cyanosphere

Summary Biocrusts are phototroph‐driven communities inhabiting arid soil surfaces. Like plants, most photoautotrophs (largely cyanobacteria) in biocrusts are thought to exchange fixed carbon for essential nutrients like nitrogen with cyanosphere bacteria. Here, we aim to compare beneficial interactions in rhizosphere and cyanosphere environments, including finding growth‐promoting strains for hosts from both environments. To examine this, we performed a retrospective analysis of 16S rRNA gene sequencing datasets, host–microbe co‐culture experiments between biocrust communities/biocrust isolates and a model grass ( Brachypodium distachyon ) or a dominant biocrust cyanobacterium ( Microcoleus vaginatus ), and metabolomic analysis. All 18 microbial phyla in the cyanosphere were also present in the rhizosphere, with additional 17 phyla uniquely found in the rhizosphere. The biocrust microbes promoted the growth of the model grass, and three biocrust isolates ( Bosea sp._L1B56, Pseudarthrobacter sp. _ L1D14 and Pseudarthrobacter picheli_ L1D33) significantly promoted the growth of both hosts. Moreover, pantothenic acid was produced by Pseudarthrobacter sp._L1D14 when grown on B . distachyon exudates, and supplementation of plant growth medium with this metabolite increased B . distachyon biomass by over 60%. These findings suggest that cyanobacteria and other diverse photoautotrophic hosts can be a source for new plant growth‐promoting microbes and metabolites.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Rhizosphere Microbiome Diversity Potentially Supports Robust Nature of Field Pennycress ( Thlaspi arvense L.) in Dryland Cropping Systems of Eastern Washington

ABSTRACT Field pennycress ( Thlaspi arvense L.) is an annual in the Brassicaceae family and is currently being developed as an oilseed intermediate crop suitable for renewable biodiesel and jet fuel. It displays many desirable characteristics for this role including cold tolerance, a rapid life cycle, and a seed fatty acid profile conducive to bioenergy generation. These traits make field pennycress favorable for winter oilseed cultivation in the inland Pacific Northwest (iPNW). Simultaneously, intermediate crops are an increasingly recognized component of both agronomic sustainability and soil health management. Intermediate crops enhance soil microbial diversity, which benefits both soil and plant health. To understand the impact of field pennycress on soil microbial diversity, two natural accessions and seven experimental accessions were grown at three sites in Eastern Washington. Aboveground biomass and rhizosphere soil were then collected. Soil genomic DNA was extracted from rhizosphere samples and used to generate an amplicon library for bacterial (16S) and fungal (ITS) rRNA sequences. The resulting libraries were analyzed in QIIME2, which revealed that not only did the fad2 deficient line from the Spring32‐10 background have significantly increased aboveground biomass production compared to other pennycress genotypes, but also displayed significantly higher β‐diversity in the rhizosphere community specifically at the site experiencing the driest conditions. ANCOM analysis showed that multiple sequences similar to beneficial plant and soil health enhancing organisms such as Trichoderma spirale , Pseudomonas spp., and Methylobacterium goesingense were found to be enriched in the microbiome of the fad2 Spring32‐10 background also at that site. To add additional context to rhizosphere community data, root exudates from two pennycress genotypes were captured in magenta boxes and analyzed using HPLC. Future work will expand our understanding of the mechanisms by which field pennycress creates diversity in the rhizosphere, thus expanding our ability to cultivate this crop in the iPNW.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

Building a genome-based understanding of bacterial pH preferences

The environmental preferences of many microbes remain undetermined. This is the case for bacterial pH preferences, which can be difficult to predict a priori despite the importance of pH as a factor structuring bacterial communities in many systems. We compiled data on bacterial distributions from five datasets spanning pH gradients in soil and freshwater systems (1470 samples), quantified the pH preferences of bacterial taxa across these datasets, and compiled genomic data from representative bacterial taxa. While taxonomic and phylogenetic information were generally poor predictors of bacterial pH preferences, we identified genes consistently associated with pH preference across environments. We then developed and validated a machine learning model to estimate bacterial pH preferences from genomic information alone, a model that could aid in the selection of microbial inoculants, improve species distribution models, or help design effective cultivation strategies. More generally, we demonstrate the value of combining biogeographic and genomic data to infer and predict the environmental preferences of diverse bacterial taxa.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Nutrients cause consolidation of soil carbon flux to small proportion of bacterial community

Nutrient amendment diminished bacterial functional diversity, consolidating carbon flow through fewer bacterial taxa. Here, we show strong differences in the bacterial taxa responsible for respiration from four ecosystems, indicating the potential for taxon-specific control over soil carbon cycling. Trends in functional diversity, defined as the richness of bacteria contributing to carbon flux and their equitability of carbon use, paralleled trends in taxonomic diversity although functional diversity was lower overall. Among genera common to all ecosystems, Bradyrhizobium, the Acidobacteria genus RB41, and Streptomyces together composed 45–57% of carbon flow through bacterial productivity and respiration. Bacteria that utilized the most carbon amendment (glucose) were also those that utilized the most native soil carbon, suggesting that the behavior of key soil taxa may influence carbon balance. Mapping carbon flow through different microbial taxa as demonstrated here is crucial in developing taxon-sensitive soil carbon models that may reduce the uncertainty in climate change projections.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Tropical Forest Soil Microbiome Modulates Leaf Heat Tolerance More Strongly Under Warming Than Ambient Conditions

ABSTRACT It is unclear how plants respond to increasing temperatures. Leaf heat tolerance (LHT) is often at its upper limit in tropical forests, suggesting that climate change might negatively impact these forests. We hypothesized that intraspecific variation in LHT might be associated with changes in the soil microbiome, which might also respond to climate. We hypothesized that warming would increase LHT through changes in the soil microbiome: we combined an in situ tropical warming experiment with a shade house experiment in Puerto Rico. The shade house experiment consisted of growing seedlings of Guarea guidonia , a dominant forest species, under different soil microbiome treatments (reduced arbuscular mycorrhizal fungi, reduced plant pathogens, reduced microbes, and unaltered) and soil inoculum from the field experiment. Heat tolerance was determined using chlorophyll fluorescence ( F V /F m ) on individual seedlings in the field and on groups of seedlings (per pot) in the shade house. We sequenced soil fungal DNA to analyze the impacts of the treatments on the soil microbiome. In the field, seedlings from ambient temperature plots showed higher F V /F m values under high temperatures (0.648 at 46°C and 0.067 at 52°C) than seedlings from the warming plots (0.535 at 46°C and 0.031 at 52°C). In the shade house, the soil microbiome treatments significantly influenced the fungal community composition and LHT ( T crit and F V /F m ). Reduction in fungal pathogen abundance and diversity altered F V /F m before T 50 for seedlings grown with soil inoculum from the warming plots but after T 50 for seedlings grown with soil inoculum from the ambient plots. Our findings emphasize that the soil microbiome plays an important role in modulating the impacts of climate change on plants. Understanding and harnessing this relationship might be vital for mitigating the effects of warming on forests, emphasizing the need for further research on microbial responses to climate change.

Hernandes Villani, Gabriela [Department of Plant B↗
DOE OSTI2026

A Catabolic Powerhouse for Biorefineries: Characterization and Engineering Erwinia spp. Strain LJJL01 to Produce Bioproducts

Nonmodel microbial hosts can efficiently biotransform unconventional organic feedstocks into advanced bioproducts, leveraging their superior metabolic capabilities and resilience to process-relevant physicochemical conditions. In this study, we domesticated Erwinia spp. strain. LJJL01 (Er LJJL01) as a potent microbial chassis to advance the emerging biorefinery strategy for the valorization of lignin-rich biomass and plastic, thereby enabling the circular economy. The strain exhibits remarkable chemical tolerance and can biofunnel various substrates, including sugars, acids, polyols, and aromatics, in minimal salt media to produce native fine chemicals such as acetoin, 2,3-butanediol, and lactic acid. As a proof of concept, engineering this strain with advanced genetic tools enables the production of tailored high-value chemicals, such as cis,cis-muconate from plastic-derived terephthalate and polyhydroxybutyrate from lignocellulosic hydrolysate. We established Er LJJL01 as a potent microbial chassis for the green synthesis of bioproducts from unconventional organic feedstocks.

09 BIOMASS FUELS↗
NASA NTRS2008

Lunar Regolith Biomining: Workshop Report

On May 5th and 6th, 2007, NASA Ames Research Center hosted a workshop entitled 'Lunar Regolith Biomining'. The workshop addressed the feasibility of biologically-based mining of the lunar regolith along with identification of views and concepts for moving this topic forward to NASA. Workshop presentations provided background in topics of interest that served as the foundation for discussion in the subsequent breakout sessions. The first topical area included the history, status, and issues with biomining on Earth to familiarize all attendees with current activities. These presentations related the primary considerations in existing biomining, e.g., microbes of choice, pH of reactions, time and temperature, specific mining applications and locations, and benefits and/or limitations of biomining. The second area reviewed existing research efforts addressing biomining of planetary surfaces (Mars, Moon), including microbial considerations, and chemical necessities in biomining and biofuel production. The last element pertained to other non-biological considerations and influences in biomining efforts on the lunar surface such as radiation fluxes and effects, and the application of small satellite experiments to learn more about the lunar and Martian surfaces. Following the presentations, the workshop attendees divided into three breakout sessions to discuss areas of interest in greater detail and to define next steps in determining the feasibility of lunar regolith biomining. Topics for each of the three breakout sessions included: 1) bio-communities of choice, target product(s), and suggested ground studies; 2) physical/environmental issues and ground studies; and 3) the development of reference experiments for the Astrobiology Small payloads Workshop. The results of the breakout sessions are summarized and a list of participants is included.

Dalton, Bonnie P.↗
DOE OSTI2026

Fostering Peat Moss Feedbacks to Accelerate Peatland Restoration

Extensive knowledge exists on plant-species traits and functions, but we understand less about how population- or community-level emergent traits influence ecosystem functioning. This knowledge gap is important for ecosystems like peatlands, arid drylands, salt marshes, seagrass meadows and mangroves, where emergent traits of plant communities can create plant-environment feedbacks that amplify or dampen ecosystem processes. Recent insights from restoration ecology suggest that these feedbacks can critically influence restoration success. Despite growing recognition of emergent trait-driven feedbacks in other ecosystems, they remain underexplored in peatland restoration, world’s most carbon-dense ecosystem. Here, we review emergent self-amplifying and self-dampening feedbacks with net positive effects for peat moss-dominated systems. We show how these feedbacks can promote key physical, chemical, and biological processes that enhance peat moss growth, increase water retention, and reduce microbial decomposition of organic matter. Understanding and fostering these feedbacks offers a promising framework to accelerate peatland restoration across diverse degradation states.

Sphagnum↗
DOE OSTI2023

The algal microbiome protects Desmodesmus intermedius from high light and temperature stress

The mutualistic impacts of bacterial communities on algal growth and biomass accumulation are well documented, but it is unknown how temperature and light stress influence these mutualistic interactions. Here, we generated a bacteria-free (axenic) culture of the green alga Desmodesmus intermedius C046 and compared its growth and yield in the laboratory to its native xenic counterpart - featuring a microbiome that has been maintained with its algal host for at least 5 years – under unstressed and high light, high temperature stress conditions. We then added exogenous microbiomes from high light marine environments to the axenic culture to quantify the contribution of these newly generated microbial consortia in improving D. intermedius growth, yield, and resilience to high light and heat. The native microbiome increased growth and biomass accumulation, under both unstressed and stressed conditions. Three of the five newly derived microbiomes had increased growth compared to the axenic and negligible, decreased, or increased growth impacts compared to the xenic culture. Stress conditions led to decreased algal exudation, larger algal cells when grown alone, but smaller algal cells with an associated microbiome. In conclusion, our results suggest that the algal microbiome plays a role in microalgal response to heat and light stress, and efforts for screening new algal strains for high biomass productivity under those stress conditions should include a strategy for microbiome optimization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Metagenome-assembled genomes from Wind River Basin floodplain sediments Riverton, Wyoming site (May to September 2017)

Microorganisms play a key role in cycling nutrients and contaminants in the terrestrial environment depending on their genetic potential. Here we present metagenome-assembled genomes (MAGs) for the bacterial and archaeal community in floodplain sediment samples taken roughly every month in the period May 18 to September 13 in 2017 at a location (Pit2) close to DOE Legacy Management well 855 at the Riverton, Wyoming floodplain site in the Wind River Basin (WRB). The groundwater at this site exhibits persistent U, Mo, and sulfate plumes and is one of the field sites in focus for the SLAC Groundwater Quality SFA program. Cores were taken with a hand-auger and separated into 5-20 cm segments based on soil horizonation down to 150 cm depth below surface. Each segment was subsampled for microbial analyses. Corresponding 16S rRNA gene amplicon data is available at the NCBI Single Read Archive (SRA) Database BioProject ID PRJNA626616, and soil geochemistry data at doi:10.15485/1631972. 40 metagenomes were sequenced through JGI and can be found under Gold sequencing project: Gs0142591. Metagenomes were assembled, binned, and refined using metawrap to generate MAGs (>50% complete and < 10% contamination based on checkM scores). This dataset includes a zip file of 6993 MAG fasta files and a csv file with quality, taxonomic classification (GTDB RS220), and metagenome accessions for MAGs generated from the Wind River Basin (WRB). This dataset also includes a file-level metadata (flmd.csv) file that lists each file contained in the dataset with associated metadata and a data dictionary (dd.csv) file that contains column/row headers used throughout the files along with a definition, units, and data type.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Correlative SIP-FISH-Raman-SEM-NanoSIMS links identity, morphology, biochemistry, and physiology of environmental microbes

Microscopic and spectroscopic techniques are commonly applied to study microbial cells but are typically used on separate samples, resulting in population-level datasets that are integrated across different cells with little spatial resolution. To address this shortcoming, we developed a workflow that correlates several microscopic and spectroscopic techniques to generate an in depth analysis of individual cells. By combining stable isotope probing (SIP), fluorescence in situ hybridization (FISH), scanning electron microscopy (SEM), confocal Raman microspectroscopy (Raman), and nano-scale secondary ion mass spectrometry (NanoSIMS), we illustrate how individual cells can be thoroughly interrogated to obtain information about their taxonomic identity, structure, physiology, and metabolic activity. Analysis of an artificial community demonstrated that our correlative approach was able to resolve the activity of single cells using heavy water SIP in conjunction with Raman and/or NanoSIMS and establish their taxonomy and morphology using FISH and SEM. We then applied this workflow to a sample of yet uncultured multicellular magnetotactic bacteria. In addition to establishing their identity and activity, backscatter electron microscopy (BSE), NanoSIMS, and energy-dispersive X-ray spectroscopy (EDS) were employed to characterize the magnetosomes within the cells. By integrating these techniques, we demonstrate a cohesive approach to thoroughly study environmental microbes at single cell resolution.

Schaible, George↗
DOE OSTI2022

Hyphae move matter and microbes to mineral microsites: Integrating the hyphosphere into conceptual models of soil organic matter stabilization

Associations between soil minerals and microbially derived organic matter (often referred to as mineral-associated organic matter or MAOM) form a large pool of slowly cycling carbon (C). Here, the rhizosphere, soil immediately adjacent to roots, is thought to control the spatial extent of MAOM formation because it is the dominant entry point of new C inputs to soil. However, emphasis on the rhizosphere implicitly assumes that microbial redistribution of C into bulk (non-rhizosphere) soils is minimal. We question this assumption, arguing that because of extensive fungal exploration and rapid hyphal turnover, fungal redistribution of soil C from the rhizosphere to bulk soil minerals is common, and encourages MAOM formation. First, we summarize published estimates of fungal hyphal length density and turnover rates and demonstrate that fungal C inputs are high throughout the rhizosphere–bulk soil continuum. Second, because colonization of hyphal surfaces is a common dispersal mechanism for soil bacteria, we argue that hyphal exploration allows for the non-random colonization of mineral surfaces by hyphae-associated taxa. Third, these bacterial communities and their fungal hosts determine the chemical form of organic matter deposited on colonized mineral surfaces. Collectively, our analysis demonstrates that omission of the hyphosphere from conceptual models of soil C flow overlooks key mechanisms for MAOM formation in bulk soils. Moving forward, there is a clear need for spatially explicit, quantitative research characterizing the environmental drivers of hyphal exploration and hyphosphere community composition across systems, as these are important controls over the rate and organic chemistry of C deposited on minerals.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

Characterization of a novel polyextremotolerant fungus, Exophiala viscosa , with insights into its melanin regulation and ecological niche

Black yeasts are polyextremotolerant fungi that contain high amounts of melanin in their cell wall and maintain a primary yeast form. These fungi grow in xeric, nutrient depletes environments which implies that they require highly flexible metabolisms and have been suggested to contain the ability to form lichen-like mutualisms with nearby algae and bacteria. However, the exact ecological niche and interactions between these fungi and their surrounding community are not well understood. We have isolated 2 novel black yeasts from the genus Exophiala that were recovered from dryland biological soil crusts. Despite notable differences in colony and cellular morphology, both fungi appear to be members of the same species, which has been named Exophiala viscosa (i.e. E. viscosa JF 03-3 Goopy and E. viscosa JF 03-4F Slimy). A combination of whole genome sequencing, phenotypic experiments, and melanin regulation experiments have been performed on these isolates to fully characterize these fungi and help decipher their fundamental niche within the biological soil crust consortium. Our results reveal that E. viscosa is capable of utilizing a wide variety of carbon and nitrogen sources potentially derived from symbiotic microbes, can withstand many forms of abiotic stresses, and excretes melanin which can potentially provide ultraviolet resistance to the biological soil crust community. Besides the identification of a novel species within the genus Exophiala, our study also provides new insight into the regulation of melanin production in polyextremotolerant fungi.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Phase II Final Technical Report: Dynamic Gamma-ray Imaging for In Vivo Tracking of Microelement Transport Across Plant-Microbial Systems

The Department of Energy Office of Biological and Environmental Research (DOE BER) Mesoscale to Molecules Bioimaging Technology Program aims to develop new imaging and measurement technology to enable in situ and dynamic imaging across a range of spatial and temporal scales. Various imaging modalities are required to span the complete spatiotemporal landscape for bioenergy and environmental bioimaging needs. The high-resolution gamma-ray spectroscopy, imaging, and sensitivity of new high-purity germanium (HPGe) instruments provide a unique opportunity to complement and enhance these research goals. The HPGe-based Gamma-ray Imager for Plant Research (GIPR) developed here provides non-invasive, in vivo measurements to dynamically track the uptake and distribution of multiple gamma-emitting radioactive elements simultaneously as they move from the soil microbiome into living plants. The hand portable GIPR utilizes commercially available radioisotopes to provide spatial and temporal imaging of plant microelement exchange for the broader scientific community.

Kiser, Matthew↗
DOE OSTI2022

Success Through Synergy (STS): Increasing Cultivation Yield and Stability with Rationally Designed Consortia

Consortia of functionally diverse microbial assemblages hold potential to enhance the production of microalgae for biofuels and bioproducts. In laboratory experiments, increases in algal yield have been achieved through the amendment of algal cultures with growth-promoting bacteria, as well as the rational design of algal-algal consortia (i.e., combining strains with complementary traits). Despite this evidence, commercial systems typically rely on monocultures because of the technical challenges that multi-taxa cultures impose. In this project, we aimed to generate algae-bacteria and algae-algae consortia in the lab and test the performance of these consortia against monoculture counterparts in the field. We also aimed to generate toolkits that could be used by the scientific community and algal farmers to advance the cultivation of consortia.

60 APPLIED LIFE SCIENCES↗
NASA NTRSDate not supplied

Final Report of the COSPAR Meeting Series on Knowledge Gaps in Planetary Protection for Crewed Missions to Mars

The international scientific community has, through a Committee on Space Research (COSPAR) meeting series, participated in a seven-year review of knowledge gaps in planetary protection for crewed missions to Mars. This meeting series first identified a prioritized list of knowledge gaps that, if addressed, would permit development of an implementable planetary protection policy for the initial crewed Mars mission. Next, a consideration of the available venues (ground, LEO, deep space, planetary surfaces) yielded a roadmap of potential future investigations needed to close the knowledge gaps. Finally, the meeting series evaluated topic-specific measurements that are essential to establishing the figures of merit needed for hardware engineering design requirements. The three main topic threads considered within the meeting series were: microbial and human health monitoring; natural transport of terrestrial biological contamination on Mars, and; technologies and operations for contamination control (of spacecraft systems). Participants at the meeting series included engineers, scientists, managers and various subject matter experts from space agencies, academic and research institutions, commercial companies, and other government departments. This paper provides a detailed summary review of the findings of the meeting series, including descriptions of the basis for each of the figures of merit (where they were identified), and considerations of how planetary protection implementation and policy for crewed missions may progress from the current state of the art.

J Andy Spry↗