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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.↗

NASA NTRS2003

Preliminary Characterization of a Microbial Community of Rock Varnish from Death Valley, California

Rock varnish (also referred to as desert varnish in the literature because it is particularly noticeable in desert environments) is a dark, thin (typically 50-500 m thick), layered veneer composed of clay minerals cemented together by oxides and hydroxides of manganese and iron. Some scientists suggest that varnish may provide a historical record of environmental processes such as global warming and long-term climate change. However, despite more than 30 years of study using modern microanalytical and microbial culturing techniques, the nucleation and growth mechanisms of rock varnish remain a mystery. Rock varnish is of interest to the Mars science community because a varnish-like sheen has been reported on the rocks at the Viking Lander sites. It therefore important for us to understand the formation mechanisms of terrestrial varnish abiotic, biotic, or a combination of the two -- as this understanding may give us clues concerning the chemical and physical processes occurring on the surface of Mars. It is strongly believed by some in the biogeochemistry community that microbes have a role in forming rock varnish, and iron- and manganese-oxidation by microbes isolated from varnish has been extensively investigated. Only two of these studies have investigated the microbial genetics of varnish. These studies examined the morphological, physiological and molecular characteristics of microbes that had previously been cultured from various rock varnishes and identified the cultivars using 16S rDNA sequencing techniques. However, it is well known that most of organisms existing in nature are refractory to cultivation, so many important organisms would have been missed. The currently described work investigates the genetics of rock varnish microbial community from a site in the Whipple Mtns., south of Death Valley, CA, near Parker, Arizona. We employed both cultural and molecular techniques to characterize the microorganisms found within the varnish and surrounding soil with the objectives of (a) identifying microorganisms potentially involved in varnish formation, and (b) discovering microorganisms that simply use the varnish as an extreme habitat.

Kuhlman, K. R.↗

DOE OSTI2024

MISIP: a data standard for the reuse and reproducibility of any stable isotope probing-derived nucleic acid sequence and experiment

DNA/RNA-stable isotope probing (SIP) is a powerful tool to link in situ microbial activity to sequencing data. Every SIP dataset captures distinct information about microbial community metabolism, process rates, and population dynamics, offering valuable insights for a wide range of research questions. Data reuse maximizes the information derived from the labor and resource-intensive SIP approaches. Yet, a review of publicly available SIP sequencing metadata showed that critical information necessary for reproducibility and reuse was often missing. Here, we outline the Minimum Information for any Stable Isotope Probing Sequence (MISIP) according to the Minimum Information for any (x) Sequence (MIxS) framework and include examples of MISIP reporting for common SIP experiments. Our objectives are to expand the capacity of MIxS to accommodate SIP-specific metadata and guide SIP users in metadata collection when planning and reporting an experiment. The MISIP standard requires 5 metadata fields—isotope, isotopolog, isotopolog label, labeling approach, and gradient position—and recommends several fields that represent best practices in acquiring and reporting SIP sequencing data (e.g., gradient density and nucleic acid amount). The standard is intended to be used in concert with other MIxS checklists to comprehensively describe the origin of sequence data, such as for marker genes (MISIP-MIMARKS) or metagenomes (MISIP-MIMS), in combination with metadata required by an environmental extension (e.g., soil). The adoption of the proposed data standard will improve the reuse of any sequence derived from a SIP experiment and, by extension, deepen understanding of in situ biogeochemical processes and microbial ecology.

Simpson, Abigayle↗

DOE OSTI2025

Reduced soil diazotroph diversity decreases nitrogen fixation rates, but depends on land management

Soil diazotrophs convert atmospheric nitrogen into plant-available ammonium through free-living nitrogen fixation (FLNF). This sustainable nitrogen source can reduce our dependence on synthetic fertilizer inputs in conventional agricultural systems. However, we know little about the effect of diazotroph diversity on FLNF, especially given that FLNF is intermediate within the broad-narrow functional spectrum. Here, we determined how management-mediated shifts in diazotroph diversity would impact their ecosystem function (FLNF) by quantifying diazotroph diversity across a long-term management gradient during and after the growing season. In addition to field observations, we leveraged the same management gradient to manipulate diversity in soil microcosms via chloroform fumigation exposure. In the field, diazotroph diversity was significantly higher after the growing season, and the biologically-based annual cropping system harbored the highest diazotroph diversity. However, perennial cropping systems maintained the highest FLNF despite lower diazotroph diversity, and both soil moisture and temperature were stronger predictors of FLNF. Based on these results, integrating diverse perennial crops into agricultural landscape could result in greater N from FLNF, particularly at the end of the growing season. When we reduced biodiversity in a manipulation experiment, the diversity-FLNF association was stronger than in the field experiment, suggesting that FLNF communities are not as functionally redundant as taxonomically ‘broad’ ecosystem functions. Here, the strength of diversity-FLNF correlation varied by previous land management. Diazotroph diversity better predicted FLNF in annual and forest soil microcosms, and microbial biomass carbon better predicted FLNF in perennial soil microcosms. Taken together, our results show that while diazotroph diversity influences FLNF, especially under extreme environmental disturbances, abiotic factors like soil moisture and temperature are stronger constraints on FLNF in the field.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2026

Characterization of Multiple Trichloroethene, cis-Dichloroethene and 1,1-Dichloroethene Degrading Propanotrophic Communities

Aerobic cometabolism offers a viable strategy for the remediation of chlorinated solvent plumes at oxic sites where anaerobic approaches are limited. In this study, propane-enriched mixed cultures (derived from agricultural soils and an impacted site sediment) which previously degraded 1,4-dioxane, were evaluated for their capacity to also degrade trichloroethene (TCE), cis-1,2-dichloroethene (cDCE), and 1,1-dichloroethene (1,1-DCE) over successive transfers. Sustained biodegradation of TCE and cDCE was observed across multiple enrichments, and cultures enriched on one compound generally degraded the other. In contrast, 1,1-DCE biodegradation was restricted to a subset of cultures and removal times increased over transfers. Further, 1,1-DCE removal was absent at elevated concentrations, both trends consistent with inhibitory or toxic effects. Whole genome sequencing analyses revealed pronounced substrate-dependent selection of microbial communities, with cDCE-degrading cultures being dominated by Mycobacterium and Mycolicibacterium, whereas TCE-degrading cultures were dominated by Rhodococcus. Rhodococcus metagenome-assembled genomes (MAGs) in the TCE degrading cultures classified as R. opacus or R. wratislaviensis. 1,1-DCE degrading cultures were dominated by Pseudonocardia, although the associated MAGs contained a truncated propane monooxygenase alpha subunit. Functional gene analysis identified both group 5 (prmABCD) and putative group 6 propane monooxygenases. The following KBase narratives contain the quality controlled reads, MAGs (fasta assemblies) and the prokka annotations for each assembly TCE Site 1A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254918) TCE Soil 2A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254919) TCE Soil T3 A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254920) TCE Soil T4 A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254921) cDCE Site 1A 1B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254915) cDCE Soils T2 A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254927) cDCE Soil T3 A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254928) cDCE Soil 4A and B Propanotrophic MAGs (https://narrative.kbase.us/narrative/254942) 1,1-DCE T2 T3 Propanotrophic MAGs (https://narrative.kbase.us/narrative/254903)

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS2006

Comets, Asteroids, and the Origin of the Biosphere

During the past few decades, the role of comets in the delivery of water, organics, and prebiotic chemicals to the Biosphere of Earth during the Hadean (4.5-3.8 Ga) period of heavy bombardment has become more widely accepted. However comets are still largely regarded as frigid, pristine bodies of protosolar nebula material that are entirely devoid of liquid water and consequently unsuitable for life in any form. Complex organic compounds have been observed comets and on the water rich asteroid 1998 KY26, which has color and radar reflectivity similar to the carbonaceous meteorites. Near infrared observations have indicated the presence of crystalline water ice and ammonia hydrate on the large Kuiper Belt object (50000) Quaoar with resurfacing that may indicate cryovolcanic outgassing and the Cassini spacecraft has detected water-ice geysers on Saturn s moon Enceladus. Spacecraft observations of the chemical compositions and characteristics of the nuclei of several comets (Halley, Borrelly, Wild 2, and Tempel 1) have now firmly established that comets contain a suite of complex organic chemicals; water is the predominant volatile; and that extremely high temperatures (approx.350-400 K) can be reached on the surface of the very black (albedo-0.03) nuclei when the comets are with 1.5 AU from the Sun. Impact craters and pinnacles observed on comet Wild 2 suggest a thick crust and episodic outbursts and jets observed on the nuclei of several comets are interpreted as indications that localized regimes of liquid water and water vapor can periodically exist beneath the crust of some comets. The Deep Impact observations indicate that the temperature on the nucleus of of comet Tempel 1 at 1.5 AU varied from 330K on the sunlit side to a minimum of 280+/-8 K. It is interesting that even the coldest region of the comet surface was slightly above the ice/liquid water phase transition temperature. These results suggest that pools and films of liquid water can exist in a wide range of temperatures in cavities and voids at different depths just beneath the crust of a comet. The possibility that liquid water may exist over a wide range of temperatures on comets significantly enhances the possibility that these bodies may harbor niches suitable for microbial communities and ecosystems. Such niches would by ideal for the growth of psychrophilic, mesophilic, and possibly even thermophilic chemolithotrophs and photoautotrophs such as the motile filamentous cyanobacteria (e.g., Calothrix, Oscillatoria, Phormidium, and Spirulina) that can grow in geothermal springs and geysers at temperatures ranging from 320K to 345K and in cold polar desert soils. This paper reviews the observational data in support of the hypothesis that liquid water can exist in permafrost-like active regions just beneath the surface of comets when near perihelion and provides additional arguments in support of the hypothesis that comets, carbonaceous meteorites, and asteroids may have played a significant role in the origin and evolution of the Biosphere and in the distribution of microbial life throughout the Solar System.

Hoover, Richard B.↗

DOE OSTI2025

Manganese concentration influences nitrogen cycling in agricultural soil

Manganese (Mn) can modulate nitrogen (N) transformations in soil, yet its role in agroecosystems remains understudied. We conducted a 51-day microcosm incubation with agricultural soils differing in long-term N history (N 0 , no added N; N 1 , added 225 kg N ha −1 for a duration of 27 years) and amended with soluble Mn at 0 (M 0 ), 50 (M 1 ), or 250 (M 2 ) mg kg −1 and Glyceria striata (Lam.) residue. In N 1 soils, Mn additions (both M 1 and M 2 ) lowered total mineral N by 25 % relative to N 1 M 0 at day 51 and reduced 51-day cumulative N₂O by 32 % (N 1 M 1 ) and 42 % (N 1 M 2 ) vs. N 1 M 0 , whereas effects in N 0 were negligible. Mn also depressed ammonia-oxidizing bacterial amoA gene transcripts at day 15 in N 1 M 2 vs. N 1 M 0 (2.5 fold change). This reduction was likely due to increased N loss via complete denitrification to N₂ through microbial pathways such as nitrate/nitrite-dependent manganese oxidation (NDMO), where bacteria directly used the added Mn 2+ to reduce nitrate (NO 3 − ) and nitrite (NO 2 − ) to N 2 or as Mn dependent-ammonia oxidation (Mnammox) where bacteria oxidized ammonium (NH 4 + ) to N 2 , using Mn oxides as electron acceptors that formed from the oxidation of the added Mn. Other contributing mechanisms may include Mn-induced N immobilization, toxicity, and changes in the microbial community. These mechanistic results indicate that background Mn availability and redox dynamics can shape nitrification–denitrification pathways under N-rich conditions. In conclusion, we highlight how native Mn pools and redox state may help explain observed variability in N losses and greenhouse gas production across agricultural soils.

Decomposition↗

DOE OSTI2024

Abiotic Stress Reorganizes Rhizosphere and Endosphere Network Structure of Sorghum bicolor

Sorghum bicolor is a promising bioenergy feedstock with high biomass production and unusual tolerance for stresses, such as water and nutrient limitation. Although the membership of the sorghum microbiome in response to stress has been explored, relatively little is known about how microbe–microbe networks change under water- or nutrient-limited conditions. This is important because network changes can indicate impacts on the functionality and stability of microbial communities. We performed network-based analysis on the core bacterial and archaeal community of an agronomically promising high biomass bioenergy genotype, Grassl, grown under nitrogen and water stress. Stress caused relatively minor changes in bacterial abundances within soil, rhizosphere, and endosphere communities but led to significant changes in bacterial network structure and modularity. We found a complete reorganization of network roles in all plant compartments, as well as an increase in the modularity and proportion of positive associations, which potentially could represent coexistence and cooperation in the sorghum bacterial/archaeal community under stress. Although stressors are often believed to be destabilizing, we found stressed networks were as or more stable than non-stressed networks, likely due to their redundancy and compartmentalization. Together, these findings support the idea that both sorghum and its bacterial/archaeal community can be resilient to future environmental stressors.

09 BIOMASS FUELS↗

NASA NTRS2006

Comets, Asteroids, Meteorites, and the Origin of the Biosphere

During the past few decades, the delivery of water, organics, and prebiotic chemicals to the Biosphere of Earth during the Hadean (4.5-3.8 Ga) period of heavy bombardment by comets and asteroids has become more widely accepted. Comets are still largely regarded as frigid, pristine bodies of protosolar nebula material that are devoid of liquid water and therefore unsuitable for life. Complex organic compounds have been observed in comets and on the water-rich asteroid 1998 KY26 and near IR observations have indicated the presence of crystalline water ice and ammonia hydrate on the large Kuiper Belt object (50000) Quaoar that has resurfacing suggesting cryovolcanic outgassing. Spacecraft observations of the chemical compositions and characteristics of the nuclei of several comets (Halley, Borrelly, Wild 2, and Tempel 1) have shown that comets contain complex organic chemicals; that water is the predominant volatile; and that extremely high temperatures (approx. 350-400 K) can be reached on the surfae of the very black (albedo approx. 0.03) nuclei of comets when they approach the Sun. Impact craters and pinnacles observed on comet Wild 2 suggest a thick crust. Episodic outbursts and jets from the nuclei of several comets indicate that localized regimes of liquid water and water vapor can periodically exist beneath the comet crust. The Deep Impact mission found the temperature of the nucleus of comet Tempel 1 at 1.5 AU varied from a minimum of 280 plus or minus 8 K the 330K (57 C) on the sunlit side. In this paper it is argued that that pools and films of liquid water exist (within a wide range of temperatures) in cavities and voids just beneath the hot, black crust. The possibility of liquid water existing over a wide range of temperatures significantly enhances the possibility that comets might contain niches suitable for the growth of microbial communities and ecosystems. These regimes would be ideal for the growth of psychrophilic, mesophilic, and thermophilic photoautotrophs and chemolithotrophs such as the motile filamentous cyanobacteria (e.g., Calothrix, Oscillatoria, Phormidium, and Spirulina) that grow in geothermal springs and geysers of Earth at temperatures ranging fiom 320K to 345K and are also found growing in cold polar desert soils. The mineralized remains of morphotypes of all of these cyanobacteria have also been found in the Orgueil CI1 and the Murchison CN2 carbonaceous meteorites that may derive from cometary parent bodies. Observational results that support the hypothesis that liquid water can in active regions just beneath the surface of comets and that comets, carbonaceous meteorites, and asteroids may have played a significant role in the origin and evolution of the Biosphere and in the distribution of microbial life throughout the Solar System.

Hoover, Richard B.↗

DOE OSTI2024

Genetic modification of the shikimate pathway to reduce lignin content in switchgrass ( Panicum virgatum L.) significantly impacts plant microbiomes

Switchgrass (Panicum virgatum L.) is considered a sustainable biofuel feedstock, given its fast-impact growth, low input requirements, and high biomass yields. Improvements in bioenergy conversion efficiency of switchgrass could be made by reducing its lignin content. Engineered switchgrass that expresses a bacterial 3-dehydroshikimate dehydratase (QsuB) has reduced lignin content and improved biomass saccharification due to the rerouting of the shikimate pathway towards the simple aromatic protocatechuate at the expense of lignin biosynthesis. However, the impacts of this QsuB trait on switchgrass microbiome structure and function remain unclear. To address this, wild-type and QsuB-engineered switchgrass were grown in switchgrass field soils, and samples were collected from inflorescences, leaves, roots, rhizospheres, and bulk soils for microbiome analysis. We investigated how QsuB expression influenced switchgrass-associated fungal and bacterial communities using high-throughput Illumina MiSeq amplicon sequencing of ITS and 16S rDNA. Compared to wild-type, QsuB-engineered switchgrass hosted different microbial communities in roots, rhizosphere, and leaves. Specifically, QsuB-engineered plants had a lower relative abundance of arbuscular mycorrhizal fungi (AMF). Additionally, QsuB-engineered plants had fewer Actinobacteriota in root and rhizosphere samples. These findings may indicate that changes in the plant metabolism impact both AMF and Actinobacteriota similarly or potential interactions between AMF and the bacterial community. This study enhances understanding of plant-microbiome interactions by providing baseline microbial data for developing beneficial bioengineering strategies and by assessing nontarget impacts of engineered plant traits on the plant microbiome.

09 BIOMASS FUELS↗

DOE OSTI2024

Necromass responses to warming: A faster microbial turnover in favor of soil carbon stabilisation

Microbial byproducts and residues (hereafter ‘necromass’) potentially play the most critical role in soil organic carbon (SOC) sequestration. However, little is known about the influence of climate warming on necromass accumulation in the agroecosystem and the underlying mechanisms associated with microbial life strategies. Here, in order to address these knowledge gaps, we used amino sugars as biomarkers of microbial necromass, and investigated their variation through an 8-year trial in an agroecosystem with two warming levels (+1.6 and + 3.2 °C) compared to ambient temperature. The results showed that the lower warming level had no impact on total microbial necromass carbon. Conversely, warming the soil 3.2 °C above ambient increased total microbial necromass by 17 % and its contribution to SOC by 21.3 %, mainly by increasing fungal necromass (+19.8 %), whereas +3.2 °C warming had no impact on bacterial necromass. At the phylum level, compared with the ambient control, +3.2 °C warming induced an increase in the abundance of Proteobacteria and a decrease in both Acidobacteria and Actinobacteria, whereas in the fungal community, Ascomycota increased and Mortierellomycota decreased. This indicates that r-strategists outcompete K-strategists in warmer climates, which led to increased microbial necromass production and accumulation, as supported by the positive correlation between r-strategists and microbial necromass. Stronger microbial competition for resources also resulted in a higher biomass turnover rate, greater cell death, and greater production of microbial necromass. This was supported by the lower bacterial and fungal network complexity and trophic links under warming conditions. In addition, the necromass generated from accelerated microbial turnover further offsets warming-induced deceases in microbial biomass. Consequently, bulk SOC did not change, despite microbial necromass having a much greater response to warming than the soil C pool. Therefore, future climate warming may influence the composition and persistence of SOC during microbial degradation.

54 ENVIRONMENTAL SCIENCES↗

NASA NTRS2004

Microbial structural diversity estimated by dilution-extinction of phenotypic traits and T-RFLP analysis along a land-use intensification gradient

The present work tested whether the relationship between functional traits and inoculum density reflected structural diversity in bacterial communities from a land-use intensification gradient applying a mathematical model. Terminal restriction fragment length polymorphism (T-RFLP) analysis was also performed to provide an independent assessment of species richness. Successive 10-fold dilutions of a soil suspension were inoculated onto Biolog GN(R) microplates. Soil bacterial density was determined by total cell and plate counts. The relationship between phenotypic traits and inoculum density fit the model, allowing the estimation of maximal phenotypic potential (Rmax) and inoculum density (KI) at which Rmax will be half-reduced. Though Rmax decreased with time elapsed since clearing of native vegetation, KI remained high in two of the disturbed sites. The genetic pool of bacterial community did not experience a significant reduction, but the active fraction responding in the Biolog assay was adversely affected, suggesting a reduction in the functional potential. c2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

NASA Center KSC↗

DOE OSTI2026

Data for Genetic Variation in Zea mays Influences Microbial Nitrification and DeNitrification in Conventional Agroecosystems

Nitrogenous fertilizers provide a short-lived benefit to crops in agroecosystems, but stimulate nitrification and denitrification, processes that result in nitrate pollution, N2O production, and reduced soil fertility. Recent advances in plant microbiome science suggest that genetic variation in plants can modulate the composition and activity of rhizosphere N-cycling microorganisms. Here we attempted to determine whether genetic variation exists in Zea mays for the ability to influence the rhizosphere nitrifier and denitrifier microbiome under “real-world” conventional agricultural conditions. To capture an extensive amount of genetic diversity within maize we grew and sampled the rhizosphere microbiome of a diversity panel of germplasm that included ex-PVP inbreds ( Z. mays ssp. mays ), ex-PVP hybrids ( Z. mays ssp. may s), and teosinte ( Z. mays ssp. mexicana and Z. mays ssp. parviglumis ). From these samples, we characterized the microbiome, a suite of microbial genes involved in nitrification and denitrification and carried out N-cycling potential assays. Here we are showing that populations/genotypes of a single species can vary in their ecological interaction with denitrifers and nitrifers. Some hybrid and teosinte genotypes supported microbial communities with lower potential nitrification and potential denitrification activity in the rhizosphere, while inbred genotypes stimulated/did not inhibit these N-cycling activities. These potential differences translated to functional differences in N2O fluxes, with teosinte plots producing less GHG than maize plots. Taken together, these results suggest that Zea genetic variation can lead to changes in N-cycling processes that result in N leaching and N2O production, and thereby are selectable targets for crop improvement. Understanding the underlying genetic variation contributing to belowground microbiome N-cycling into our conventional agricultural system could be useful for sustainability.

Nitrogen↗

NASA NTRS1995

A 'crytic' microbial mat: A new model ecosystem for extant life on Mars

If life were present on Mars today, it would face potentially lethal environmental conditions such as a lack of water, frigid temperatures, ultraviolet radiation, and soil oxidants. In addition, the Viking missions did not detect near-surface organic carbon available for assimilation. Autotrophic organisms that lived under a protective layer of sand or gravel would be able to circumvent the ultraviolet radiation and lack of fixed carbon. Two terrestrial photosynthetic near-surface microbial communities have been identified, one in the inter- and supertidal of Laguna Ojo de Liebere (Baja California Sur, Mexico) and one in the acidic gravel near several small geysers in Yellowstone National Park (Wyoming, U.S.A.). Both communities have been studied with respect to their ability to fix carbon under different conditions, including elevated levels of inorganic carbon. Although these sand communities have not been exposed to the entire suite of Martian environmental conditions simultaneously, such communities can provide a useful model ecosystem for a potential extant Martian biota.

Rothschild, L. J.↗

DOE OSTI2025

Lipids represent a dynamic, yet stable pool of microbially-derived soil carbon

There is an emerging consensus that microorganisms are a primary source of persistent, slow-cycling soil organic matter (SOM), however which microbial residues contribute to SOM and what controls their accumulation remains unresolved. Lipids are a commonly overlooked biomolecular pool that could contribute significantly to stable SOM. While current estimates for phospholipid degradation in soils are rapid, lipids are structurally heterogeneous molecules that could turnover at different rates. Through a year-long soil incubation and compound-specific, stable isotope probing (SIP)-lipidomics, we were able to rigorously track the persistence of lipid compounds in silty and sandy switchgrass bioenergy crop soils. Here, we assessed the influence of lipid structure on soil lipid accrual and degradation as moderated by soil texture, since mineral association is presumed to be the primary mechanism for lipid persistence. After rapid incorporation of 13 C glucose into microbial biomass, we found 13 C label was retained broadly across chemically diverse lipid classes, even after one year. 13 C-labeled lipid profiles varied significantly with soil texture; however, we found no difference between sandy and silty soils in lipid retention, suggesting soil texture may only play a minor role in modulating lipid persistence. Only two lipid subclasses were found to be persistent (i.e., retention of 13 C label without significant degradation or production): phosphatidylinositol lipids and hydroxyceramide lipids, both of which are negatively charged, possibly facilitating stabilization by mineral complexation. However, several other subclasses displayed substantial ongoing production. In particular, the accumulation of triacylglycerol lipids across soil textures suggests that storage lipids may be an important component of SOC, highlighting a potential target for management strategies to promote C retention by lipid accrual. Overall, the retention for over a year of 13 C label in microbial intact lipid biomarkers reveals the importance of efficient biomass production and turnover when considering microbial C contributions to SOM.

compound-specific↗

DOE OSTI2025

Synthetic microbial communities: Bridging research and application in second-generation bioenergy feedstock microbiomes

The sustainable production of purpose-grown bioenergy feedstocks is essential in transitioning away from fossil fuels. Synthetic communities (SynComs) are consortia of microorganisms that can be used as biological interventions to support objectives like plant growth and stress tolerance. This review examines the state of knowledge regarding microbiomes and SynComs of second-generation bioenergy feedstocks, focusing on the rhizosphere. We first provide an overview of second-generation feedstocks, including switchgrass (Panicum virgatum), miscanthus (Miscanthus × giganteus), sorghum (Sorghum spp.), sugarcane (Saccharum spp.), and poplar (Populus spp.), and summarize our current understanding of their plant-soil-microbiome ecology. We next discuss considerations in the objectives, design, and evaluation of SynComs to enhance feedstock production, and then critically review the literature around their use. Our literature analysis revealed that SynCom performance varied substantially between controlled pilot experiments and field trials, possibly due to system complexity that could not be fully considered in their design and pilot evaluation. We identified a gap in the use of SynComs to support the unique sustainability objectives of biofuel feedstock agriculture, presenting an opportunity to leverage these additional microbial traits in SynCom designs. Finally, we emphasize the importance of targeted research to identify the ecological principles that govern the assembly, activation, and persistence of microbes in the feedstock rhizosphere, thereby enhancing our capacity to manage microbiomes under diverse environmental conditions and ensure their functionality. Beyond biofuels, SynComs are a promising microbiome management strategy for crop production; however, an ecologically informed design and evaluation of SynComs are advised.

SynCom↗

DOE OSTI2025

Dissection of Carbon and Nitrogen Cycling in Post-Fire Soil Environments using a Genome- Informed Experimental Community (Final Technical Report)

Wildfires are a natural part of many forest ecosystems, with globally important carbon (C) storage and nutrient cycling consequences, and they are increasing in frequency and severity in Western North America. Forest fires affect soil C stocks in complex ways; some C is released into the atmosphere through combustion, while a large percentage of the C is added to the soil in the form of pyrogenic organic matter. Worldwide, it is estimated that 16% of soil organic matter is pyrogenic, while locally, this number may be as high as 80%. Understanding how wildfires affect soil organic matter cycling requires understanding how microbes respond to pyrogenic organic matter and other post-fire soil conditions. However, our understanding of microbial interactions within post-fire soil was in its infancy at the time of our proposal. Outstanding questions included: Which microbes are capable of degrading pyrogenic organic matter? What are the relevant genes and metabolites associated with this degradation? What are the key interactions among post-fire microbes? Key highlights of outcomes supported by this grant included training eleven early-career scientists and two early-career PIs, publication of twelve peer-reviewed papers, cross-lab collaborations that empowered complex scientific approaches, the development of an open-source automated gas sampler to drive novel insights in C cycling, enhanced understanding of post-fire microbial community dynamics, and novel genetic and molecular insights into microbial responses to fire.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2025

Tracing priming effects in palsa peat carbon dynamics using a stable isotope-assisted metabolomics approach

Introduction: Peatlands store up to a third of global soil carbon, and in high latitudes their litter inputs are increasing and changing in composition under climate change. Although litter significantly influences peatland carbon and nutrient dynamics by changing the overall lability of peatland organic matter, the physicochemical mechanisms of this impact—and thus its full scope—remain poorly understood. Methods: We applied multimodal metabolomics (UPLC-HRMS, 1 H NMR) paired with 13 C Stable Isotope-Assisted Metabolomics (SIAM) to track litter carbon and its potential priming effects on both existing soil organic matter and carbon gas emissions. Through this approach, we achieved molecule-specific tracking of carbon transformations at unprecedented detail. Results: Our analysis revealed several key findings about carbon dynamics in palsa peat. Microbes responded rapidly to litter addition, producing a short-term increase in CO 2 emissions, fueled nearly exclusively by transformations of litter carbon. Litter inputs significantly contributed to the organic nitrogen pool through amino acids and peptide derivatives, which served as readily accessible nutrient sources for microbial communities. We traced the fate of plant-derived polyphenols including flavonoids like rutin, finding evidence of their degradation through heterocyclic C-ring fission, while accumulation of some polyphenols suggested their role in limiting overall decomposition. The SIAM approach detected subtle molecular changes indicating minimal and transient priming activity that was undetectable through conventional gas measurements alone. This transient response was characterized by brief microbial stimulation followed by rapid return to baseline metabolism. Pre-existing peat organic matter remained relatively stable; significant priming of its consumption was not observed, nor was its structural alteration. Discussion: This suggests that while litter inputs temporarily increase CO 2 emissions, they don’t sustain long-term acceleration of stored carbon decomposition or substantially decrease peat’s carbon store capacity. Our findings demonstrate how technological advancements in analytical tools can provide a more detailed view of carbon cycling processes in complex soil systems.

54 ENVIRONMENTAL SCIENCES↗