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NASA NTRSDate not supplied

Method Development for In Situ Microbiome Profiling of the Water Recovery System’s Wastewater Tank Onboard the International Space Station

A distinctive microbial community has inhabited the International Space Station (ISS) Water Recovery System (WRS) for over 14 years and has experienced the stressors associated with the microgravity environment. The WRS generates potable water for the crew from urine distillate, humidity condensate, Sabatier product water, and the occasional off-loading of ground-supplied water (1). The reservoir for these products, the wastewater tank, does not have a means of microbial control. Current in situ microbial monitoring of the WRS is limited to quarterly culture-based assessments of the potable water product using a microbial capture device and coliform detection bag. Additional analysis of the wastewater and condensate sources are collected into Teflon bags for analysis following return to the ground. The time between sample collection and the return to Earth, as well as the lack of preservation, results in a skewed depiction of the microbiome. Routinely observed from these returned wastewater samples are high counts (105 – 106 colony forming units per mL) and two prevailing genera, Ralstonia and Cupriavidus, as well as a high abundance of unidentified organisms (Table 1). The wastewater tank likely contains a more diverse microbiome, as a higher diversity of bacteria and fungi has been noted upstream and downstream of the tank.

Sarah Stahl-Rommel↗

NASA NTRSDate not supplied

Nanopore Sequencing in Space: The Advancement of In Situ Microbiome Analysis for the International Space Station and Beyond

Introduction: Routine assessment of the International Space Station (ISS) microbiome has been performed through in-flight culture and Earth-based analysis. Through utilization of the ISS, significant progress toward in situ microbial identification has been achieved. In 2016, the molecular space age began when DNA-based, Earth-prepared samples were amplified within miniPCR (Genes in Space-1), and subsequent samples were sequenced with the MinION (Biomolecule Sequencer). The following year, these platforms were used collectively to yield the first off-Earth identification of unknown bacteria collected and cultured onboard the ISS (Genes in Space-3). Further advancement occurred in 2018 when a culture-independent, direct swab-to-sequencer method revealed a more thorough depiction of the ISS surface microbiome (BEST). As NASA looks towards Artemis and extended exploration missions, it is critical to continue to harness the ISS to expand in situ nanopore sequencing-based analysis in support of microbial-related crew health, planetary protection, and space research initiatives.

Christian G. Mena↗

NASA NTRS2024

Substrate Matters: Ionic Silver Alters Lettuce Growth, Nutrient Uptake, and Root Microbiome in a Hydroponics System

Ionic silver (Ag+) is being investigated as a residual biocide for use in NASA spacecraft potable water systems on future crewed missions. This water will be used to irrigate future spaceflight crop production systems. We have evaluated the impact of three concentrations (31 ppb, 125 ppb, and 500 ppb) of ionic silver biocide solutions on lettuce in an arcillite (calcinated clay particle substrate) and hydroponic (substrate-less) growth setup after 28 days. Lettuce plant growth was reduced in the hydroponic samples treated with 31 ppb silver and severely stunted for samples treated at 125 ppb and 500 ppb silver. No growth defects were observed in arcillite-grown lettuce. Silver was detectable in the hydroponic-grown lettuce leaves at each concentration but was not detected in the arcillite-grown lettuce leaves. Specifically, when 125 ppb silver water was applied to a hydroponics tray, Ag+ was detected at an average amount of 7 μg/g (dry weight) in lettuce leaves. The increase in Ag+ corresponded with a decrease in several essential elements in the lettuce tissue (Ca, K, P, S). In the arcillite growth setup, silver did not impact the plant root zone microbiome in terms of alpha diversity and relative abundance between treatments and control. However, with increasing silver concentration, the alpha diversity increased in lettuce root samples and in the water from the hydroponics tray samples. The genera in the hydroponic root and water samples were similar across the silver concentrations but displayed different relative abundances. This suggests that ionic silver was acting as a selective pressure for the microbes that colonize the hydroponic water. The surviving microbes likely utilized exudates from the stunted plant roots as a carbon source. Analysis of the root-associated microbiomes in response to silver showed enrichment of metagenomic pathways associated with alternate carbon source utilization, fatty-acid synthesis, and the ppGpp (guanosine 3′-diphosphate 5′-diphosphate) stringent response global regulatory system that operates under conditions of environmental stress. Nutrient solutions containing Ag+ in concentrations greater than 31 ppb in hydroponic systems lacking cation-exchange capacity can severely impact crop production due to stunting of plant growth.

lettuce↗

DOE OSTI2026

Soil microbial ecology and microbiome-metabolite linkages improve understanding of ecosystem states along terrestrial-aquatic interfaces

Coastal soils are dynamic systems where unique microbial niches are shaped by the intensity and duration of flooding between the terrestrial and aquatic boundaries of the terrestrial-aquatic interface (TAI). We aimed to understand the soil microbial community (16S rRNA gene) along the TAIs of a freshwater versus estuarine region and how it relates to organic matter (OM, via Fourier Transform Ion Cyclotron Resonance Mass Spectrometry). We studied the TAI gradients along a transect from upland (forested), transition (stressed forest), to wetland at three sites in each of the Lake Erie (freshwater) and Chesapeake Bay (estuarine) regions. Microbial communities differed significantly by region, transect position, and site. Contrary to expectations, given their dynamic hydrologies, transitions represented midpoints in microbial richness and diversity. We identified a core microbiome conserved across all transect positions within a region, highlighting potential microbial functions most resilient to environmental change. Indicator taxa unique to each transect position defined specific niches shaped by soil biogeochemistry. Co-expression networks of feature-level β-nearest-taxon indices revealed positive relationships in bacterial and OM feature contributions to community assembly. Our study provides critical insights into microbial communities at the forefront of hydrological changes in coastal areas that connect the land to lakes and oceans and remain vulnerable to changing weather patterns.

coastal ecosystems↗

DOE OSTI2025

Tropical intertidal microbiome response to the 2024 Marine Honour oil spill

Marine fuel oil (MFO) spills in tropical coastal environments are under-characterized despite increasing risk from maritime activities. Microbial and geochemical responses to the June 2024 Marine Honour MFO spill on Singapore's intertidal sediments were analyzed in real time over 185 days. Using metagenomics and hydrocarbon profiling, microbial community shifts and hydrocarbon degradation were quantified across visibly oiled (high-impact) and clean (low-impact) sites. Microbiomes at all sites adapted rapidly to the spill through increased diversity and abundance of genes encoding alkane and aromatic compound degradation, detoxification, and biosurfactant production. The dominant hydrocarbon-degrading bacteria differed markedly from those reported in other crude oil spills and in regions with different climates. Oil deposition intensity strongly influenced microbial succession and hydrocarbon-degrading gene profiles, and this reflected early toxicity constraints in heavily oiled areas. The persistence of hydrocarbon degradation genes beyond hydrocarbon detection in sediments suggested long-term functional priming may occur. The study provides novel genome-resolved insight into the microbial response to MFO pollution, advances understanding of marine environmental biodegradation, and provides urgently needed baseline data for oil spill response strategies in Southeast Asia and beyond.

Coastal pollution↗

DOE OSTI2025

Polyphenol rewiring of the microbiome reduces methane emissions

Methane mitigation is regarded as a critical strategy to combat the scale of global warming. Currently, ~40% of methane emissions originate from microbial sources, which is causing strategies to suppress methanogens—either through direct toxic effects or by diverting their substrates and energy—to gain traction. Problematically, current microbial methane mitigation knowledge lacks detailed microbiome-centered insights, limiting translation across conditions and ecosystems. Here we utilize genome-resolved metatranscriptomes and metabolomes to assess the impact of a proposed methane inhibitor, catechin, on greenhouse gas emissions for high-methane-emitting peatlands. In microcosms, catechin drastically reduced methane emissions by 72%–84% compared to controls. Longitudinal sampling allowed for reconstruction of a catechin degradation pathway involving Actinomycetota and Clostridium, which break down catechin into smaller phenolic compounds within the first 21 days, followed by degradation of phenolic compounds by Pseudomonas_E from Days 21 to 35. These genomes co-expressed hydrogen-uptake genes, suggesting hydrogenases may act as a hydrogen sink during catechin degradation and consequently reduce hydrogen availability to methanogens. In support of this idea, there was decreased gene expression by hydrogenotrophic and hydrogen-dependent methylotrophic methanogens under catechin treatment. There was also reduced gene expression from genomes inferred to be functioning syntrophically with hydrogen-utilizing methanogens. We propose that catechin metabolic redirection effectively starves hydrogen-utilizing methanogens, offering a potent avenue for curbing methane emissions across diverse environments including ruminants, landfills, and constructed or managed wetlands.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2025

Patulin Inhibition of Specific Apple Microbiome Members Uncovers Hanseniaspora uvarum as a Potential Biocontrol Agent

Penicillium expansum is a major postharvest pathogen of apples, causing loss in fruits through tissue damage, as well as in apple products due to contamination with the mycotoxin patulin. During infections, patulin is a cultivar-dependent virulence factor that facilitates apple lesion development. Patulin also has characterized antimicrobial activity and is important for inhibiting other competitive phytopathogens, but the role of this inhibitory activity has not been investigated in the context of the apple microbiome. Here, in our current study, we isolated 68 apple microbiota and characterized their susceptibility to P. expansum extracts. We found gram-negative bacteria and basidiomycete yeast to demonstrate largely patulin-specific growth inhibition compared with gram-positive and ascomycete isolates. From co-cultures, we identified a Hanseniaspora and Gluconobacter pairing that reduced P. expansum biomass and found that H. uvarum alone is sufficient to reduce apple disease progression in vivo. We investigated possible mechanisms of H. uvarum biocontrol activity and found modest inhibition on apple puree plates, as well as a trend toward lower patulin levels at the wound site. Active biocontrol activity required live yeast, which was also effective in controlling Botrytis cinerea apple infections. Lastly, we explored the breadth of H. uvarum biocontrol activity with over 30 H. uvarum isolates and found consistent inhibition of P. expansum apple disease.

Hanseniaspora uvarum↗

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 OSTI2025

Intestinal microbiome profile of the brown rock sea cucumber ( Holothuria glaberrima ) using ITS and 16S rDNA amplicons from direct mechanical, enzymatic, and chemical metagenomic extraction

Using direct mechanical, enzymatic, and chemical extraction methods, the intestinal microbiome of the marine invertebrate Holothuria glaberrima was obtained. ITS and 16S rDNA regions were sequenced to enrich and investigate the prokaryotic and fungal diversity profiles from different anatomical regions within the sea cucumber’s intestinal biology.

16S↗

DOE OSTI2026

Soil microbiome resilience to short-term (30 days, 90 days) and long-term (1000 days) drought

This dataset contains data used for the paper "Drought duration does not impact soil microbiome resilience". The Related References will be updated with a full citation when available. Increasing global droughts exert large but poorly understood effects on the microbial communities and ecology of soil. Microbial communities generally show resilience and return to pre-drought conditions when short-term droughted soils are rewet; soils exposed to long-term drought, however, often show a lag upon rewetting, after which microbial communities may or may not return to their pre-stressed conditions. Though short-term droughts have been widely studied, long-term drought manipulation experiments remain rare, especially those that compare microbial response to short-term and long-term drought in tandem. We conducted a 1000-day drought simulation in controlled laboratory conditions with soil cores collected from a tidal freshwater ecosystem in Washington state, USA, and subsequently exposed them to rewetting for two weeks. We also included short-term (30-day and 90-day) drought and rewet treatments to directly compare microbial community and organic matter responses across drought durations. We found distinct microbial taxa belonging to Firmicutes and Actinobacteria enriched after the 1000-day drought, but not after the short-term droughts. While we hypothesized that the microbial community would recover from a short-term drought after rewetting to resemble pre-drought conditions, our results revealed community dissimilarities between rewet and pre-drought conditions across all drought durations. These findings suggest unique microbial life history strategies within certain microbial phyla that make them successful colonizers during an extended drought period, and the influence of environmental and physiological context on microbial responses to rewetting. The 16SrRNA gene amplicon dataset contains processed DNA sequences in the form of an ASV table with raw unrarefied read counts and representative sequences in .fasta format as described in the ESS-DIVE amplicon sequence reporting format (https://ess-dive.gitbook.io/amplicon-sequencing-reporting-format/instructions). The Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FTICR-MS) dataset consists of processed files containing presence absence data of molecular formulae and molecular characterization of FTICR resolved peaks. The Nuclear Magnetic Resonance (NMR) dataset contains files relevant to NMR spectra and peaks. A sample key file and a sample metadata file is included for the FTICR/NMR and 16S dataset respectively.

1000-day drought↗

DOE OSTI2025

Lost and Found: Rediscovering Microbiome-Associated Phenotypes that Reshape Agricultural Sustainability

Overview Code and data repository for NIL Manuscript. Documentation includes sequence processing examples and data analysis. Supplemental sequence processing and R statistical analysis for publication, which compares the microbiome of teosinte-B73 Near Isogenic Lines. Sample Data Amplicon sequence data for 16S rRNA genes, the fungal ITS2 region, and nitrogen-cycling functional genes are available through the NCBI Sequence Read Archive (SRA) under accession number PRJNA1042643(https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1042643). Raw metabolomic data are available on Metabolomics Workbench, Project ID: PR002654. This study is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org where it has been assigned Study ID ST004211. The data can be accessed directly via its Project DOI: http://dx.doi.org/10.21228/M8KV8T.

Near Isogeneic Lines↗

DOE OSTI2026

Enhancing climate-smart crop performance in arid agrivoltaics systems: effects of photovoltaic shading and soil amendments on tepary bean growth, yield, and associated soil microbiome

As climate change expands the world’s arid and semiarid regions, sustainable systems that integrate food and energy production are becoming increasingly critical. Agrivoltaics—co-locating crops with photovoltaic (PV) panels—offers a dual land-use strategy that mitigates environmental stress by shading crops, conserving soil moisture, and enhancing PV efficiency. While climate-smart crops like the tepary bean ( Phaseolus acutifolius ) are well adapted to heat and drought, little is known about how these crops and their associated soil microbiomes respond to the unique microclimates created by PV shading. This study evaluated tepary bean performance and plant–microbial interactions under PV-shade vs. no shade across three soil amendment treatments at two experimental sites. We assessed plant traits including germination, phenology, biomass, height, as well as yield and bean morphology, alongside shifts in soil microbial composition and functional potential. Plants grown under PV-shade were generally taller, with extended reproductive periods and higher yields: 42% of shaded plants produced beans compared to only 8% under full sun. Shaded plants also produced rounder, higher-quality beans, whereas non-shaded plants yielded flatter, less developed beans. Microbial community composition was more strongly influenced by amendment and site conditions than by shading alone. Key microbial taxa (e.g., Glomeromycetes, Desulfobacterota ) and predicted functions (e.g., denitrification, nitrogen-respiration, sulfate reduction) were associated with differences in plant performance. Finally, combining agrivoltaic systems with targeted soil amendments can enhance crop yield and soil microbial functionality—offering a promising strategy for sustainable agriculture in arid landscapes.

14 SOLAR ENERGY↗

DOE OSTI2025

A global soil plasmidome resource unveils functional and ecological roles of plasmids in soil microbiomes

Plasmids play significant roles in microbial adaptation to ecosystems, yet their dynamics remain poorly understood due to identification challenges. We present the Global Soil Plasmidome Resource (GSPR), a comprehensive dataset of 98,728 plasmid sequences amassed from 6860 terrestrial microbial communities and isolates. We explore this resource through various computational approaches, including phylogenetic diversity analysis, host prediction, and extensive functional annotation, to understand the contribution of plasmids to the genetic and functional diversity in soil, correlating these findings with sample type, as well as the soil habitat they were retrieved from. Our analysis reveals insights into plasmid-encoded functions such as effector modules, quorum sensing, and stress resistance, which may contribute to their persistence and microbial adaptation in soil. Furthermore, CRISPR analysis suggests a prevalent role of these elements related to intra-plasmid competition. By contrasting plasmids from cultivated and uncultivated organisms, we identify important functions that expand existing knowledge of plasmid roles in these habitats. This study represents a notable step forward in elucidating plasmid diversity and function within soil microbiomes and establishes a foundational framework for exploring their roles in natural environments.

Fiamenghi, Mateus B↗

DOE OSTI2026

Viromics approaches for the study of viral diversity and ecology in microbiomes

Viruses are found across all ecosystems and infect every type of organism on Earth. Traditional culture-based methods have proven insufficient to explore this viral diversity at scale, driving the development of viromics, the sequence-based analysis of uncultivated viruses. Viromics approaches have been particularly useful for studying viruses of microorganisms, which can act as crucial regulators of microbiomes across ecosystems. They have already revealed the broad geographic distribution of viral communities and are progressively uncovering the expansive genetic and functional diversity of the global virome. Moving forward, large-scale viral ecogenomics studies combined with new experimental and computational approaches to identify virus activity and host interactions will enable a more complete characterization of global viral diversity and its effects.

Ecology↗

DOE OSTI2024

Agricultural practices influence soil microbiome assembly and interactions at different depths identified by machine learning

Agricultural practices affect soil microbes which are critical to soil health and sustainable agriculture. To understand prokaryotic and fungal assembly under agricultural practices, we use machine learning-based methods. We show that fertility source is the most pronounced factor for microbial assembly especially for fungi, and its effect decreases with soil depths. Fertility source also shapes microbial co-occurrence patterns revealed by machine learning, leading to fungi-dominated modules sensitive to fertility down to 30 cm depth. Tillage affects soil microbiomes at 0-20 cm depth, enhancing dispersal and stochastic processes but potentially jeopardizing microbial interactions. Cover crop effects are less pronounced and lack depth-dependent patterns. Machine learning reveals that the impact of agricultural practices on microbial communities is multifaceted and highlights the role of fertility source over the soil depth. Machine learning overcomes the linear limitations of traditional methods and offers enhanced insights into the mechanisms underlying microbial assembly and distributions in agriculture soils.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2025

Microbiome Adaptation Could Amplify Modeled Projections of Global Soil Carbon Loss With Climate Warming

Warming alters soil microbial traits through ecological and evolutionary processes, directly influencing the decomposition of organic matter, which significantly affects global soil carbon emissions. Yet, soil carbon models largely ignore these processes and their implications for global responses to warming. Here, we incorporate eco-evolutionary theory into a mechanistic model describing microbial soil carbon decomposition to address the question of whether such processes could have consequential effects on climate carbon feedbacks globally. We assume that a key trait of microbes, their resource allocation to production of exoenzymes (which facilitate decomposition of organic matter)—is optimized to environmental temperatures by natural selection. We find that eco-evolutionary optimization results in microbes allocating more resources to enzyme production under warming. When applied at the global scale, eco-evolutionary optimization enhances the biological realism of soil carbon models and significantly amplifies global soil carbon loss by 2100. Our results highlight the significant potential of microbial eco-evolutionary responses to influence carbon cycle feedbacks to climate change, and motivate an urgent need for more comprehensive data to accurately quantify the adaptive potential of microbiomes in response to climate change.

Biodiversity & Conservation↗

DOE OSTI2026

Animal-associated jumbo phages as widespread and active modulators of gut microbiome ecology and metabolism

Huge phages are widespread in the biosphere, yet their prevalence and ecology in the human gut remain poorly characterized. Here, we report Jug (jumbo gut) phages with genomes of 360 to 402 kilobase pairs that comprise ~1.1% of the reads in human gut metagenomes, and are predicted to infect Bacteroides and/or Phocaeicola. Although three of the four major groups of Jug phages shared >90% genome-wide sequence identity, their large terminase subunits exhibited only 38 to 57% identity, suggesting horizontal acquisition from other phages. Over 1500 genomes of Jug phages were recovered from human and animal gut metagenomes, revealing their broad distribution, with largely shared gene content suggestive of frequent cross-animal-host transmission. Jug phages displayed high gene transcription activities, including the gene for a calcium-translocating P-type ATPase not detected previously in phages. These findings broaden our understanding of huge phages and highlight Jug phages as potential major players in gut microbiome ecology.

Chen, LinXing [University of Science and Technolog↗

DOE OSTI2024

Soil microbiome interventions for carbon sequestration and climate mitigation

Mitigating climate change in soil ecosystems involves complex plant and microbial processes regulating carbon pools and flows. Here, we advocate for the use of soil microbiome interventions to help increase soil carbon stocks and curb greenhouse gas emissions from managed soils. Direct interventions include the introduction of microbial strains, consortia, phage, and soil transplants, whereas indirect interventions include managing soil conditions or additives to modulate community composition or its activities. Approaches to increase soil carbon stocks using microbially catalyzed processes include increasing carbon inputs from plants, promoting soil organic matter (SOM) formation, and reducing SOM turnover and production of diverse greenhouse gases. Marginal or degraded soils may provide the greatest opportunities for enhancing global soil carbon stocks. Among the many knowledge gaps in this field, crucial gaps include the processes influencing the transformation of plant-derived soil carbon inputs into SOM and the identity of the microbes and microbial activities impacting this transformation. As a critical step forward, we encourage broadening the current widespread screening of potentially beneficial soil microorganisms to encompass functions relevant to stimulating soil carbon stocks. Moreover, in developing these interventions, we must consider the potential ecological ramifications and uncertainties, such as incurred by the widespread introduction of homogenous inoculants and consortia, and the need for site-specificity given the extreme variation among soil habitats. Incentivization and implementation at large spatial scales could effectively harness increases in soil carbon stocks, helping to mitigate the impacts of climate change.

54 ENVIRONMENTAL SCIENCES↗