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All sourcesNASA NTRSDOE OSTI
DOE OSTI2026

Single nucleotide variants drive evolutionary phage-host arms race in anaerobic carbon dioxide-converting microbiome

Microbial bioconversions are shaped by environmental perturbations and the adaptation of resident microbiomes. Prokaryotes coexist with bacteriophages, yet their coevolutionary trajectories remain underexplored. Here, we investigate the effects of a cultivation vessel leak on an anaerobic consortium performing carbon dioxide reduction. Using time-series shotgun metagenomic sequencing, we reconstruct microbial and viral genomes to track community shifts. We further apply single-nucleotide variant profiling and CRISPR array analysis to monitor viral microdiversity and host defense mechanisms. After bioaugmentation restores bioconversion efficiency, the consortium undergoes pronounced restructuring, with new dominant taxa emerging from the rare biosphere. We identify patterns consistent with phage predation selectively removing certain species, while others exhibit resilience to infection. This shift aligns with a widespread viral outbreak and a transient increased frequency of single nucleotide variants in bacterial CRISPR–Cas defense genes. Expansion of CRISPR spacers further supports that CRISPR-mediated processes influence microbial resilience. Concurrently, phages infecting resilient hosts exhibited adaptive evolution, marked by high genetic heterogeneity. Selective pressure varies across their genomes, targeting infectivity genes and protospacer-adjacent motifs. These findings highlight a dynamic evolutionary arms race driven by the selection of beneficial genetic variants, providing a mechanistic framework for multi-omics investigations, and informing biotechnological applications, including phage-based microbiome manipulation.

Ghiotto, G↗
DOE OSTI2025

STREAMS guidelines: standards for technical reporting in environmental and host-associated microbiome studies

The interdisciplinary nature of microbiome research, coupled with the generation of complex multi-omics data, makes knowledge sharing challenging. The Strengthening the Organization and Reporting of Microbiome Studies (STORMS) guidelines provide a checklist for the reporting of study information, experimental design and analytical methods within a scientific manuscript on human microbiome research. Here, in this Consensus Statement, we present the standards for technical reporting in environmental and host-associated microbiome studies (STREAMS) guidelines. The guidelines expand on STORMS and include 67 items to support the reporting and review of environmental (for example, terrestrial, aquatic, atmospheric and engineered), synthetic and non-human host-associated microbiome studies in a standardized and machine-actionable manner. Based on input from 248 researchers spanning 28 countries, we provide detailed guidance, including comparisons with STORMS, and case studies that demonstrate the usage of the STREAMS guidelines. In conclusion, STREAMS, like STORMS, will be a living community resource updated by the Consortium with consensus-building input of the broader community.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

A genomic perspective on fungal diversity and evolution

Originating from aquatic unicellular ancestors, over the course of ~1 billion years, the fungi have evolved to occupy nearly all aerobic environments on the planet, diversified into millions of different ‘species’ and have developed complex multicellular structures. Their relatively small, simple genomes have facilitated massive-scale sequencing and allowed us to explore genome evolution across an ancient eukaryotic kingdom. With thousands of genomes from diverse lineages now available, this Review will discuss insights into fungal biology and evolution gleaned with genomics and other multi-omics approaches. Using published genomes available through GenBank and the Joint Genome Institute’s MycoCosm platform, we generated kingdom-wide phylogenies and used them to highlight how fungal genomes have changed over time. With this phylogeny as a guide, we also discuss major evolutionary transitions that occurred across the fungal kingdom. Although progress has been made, these efforts are hampered by biases in genome representation and limited characterization of gene functions. Here, in this study, we discuss these challenges and possible future directions to address them, including initiatives to characterize conserved genes of unknown function and scale up sequencing towards 10,000 annotated fungal genomes.

Mondo, Stephen J. [USDOE Joint Genome Institute (J↗
DOE OSTI2024

Adding labile carbon to peatland soils triggers deep carbon breakdown

Peatlands store vast amounts of carbon, with deep peat carbon remaining stable due to limited thermodynamic energy and transport. However, climate change-induced increases in labile carbon inputs could destabilize these stores. Here, we combined DNA stable isotope probing with stable isotope-assisted metabolomics employing a multi-platform approach to investigate microbial dynamics driving deep peat carbon degradation upon labile carbon (e.g., glucose) amendment. Our findings highlight the vulnerability of deep peat carbon, as glucose addition triggers the breakdown of older organic matter. By uniquely integrating these techniques, we identified active glucose metabolizers to specific microbial populations and mapped carbon flow through microbial networks, elucidating their role in priming recalcitrant carbon mineralization. This multi-omics approach offers crucial insights into how changing resources reshape the peatland microbiome, enhancing our understanding of deep carbon processing, and refining model parameterization to predict microbial responses and carbon cycle feedbacks under global change pressures.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

The evolution of analytical techniques for multiplex analysis of protein biomarkers

Introduction: The landscape of biomarker development has evolved with advanced analytical technologies, particularly affinity- and mass spectrometry-based techniques. These advancements have deepened our understanding of disease mechanisms, enabling the development of precise diagnostic tools and personalized medicine. Protein biomarkers, which play pivotal roles in biological processes, have become invaluable in diagnosing and monitoring diseases, aided by their presence in various biological samples and the availability of established detection methods. Areas covered: This review covers the role of protein biomarkers in clinical practice, the development and dimensionality of protein biomarkers, advancements in detection technologies, a comparison of these technologies, and future directions in biomarker discovery and disease mechanism elucidation. Expert opinion: Advances in biomarker technologies have the potential to transform diagnostics and personalized treatment but face challenges such as high costs and technical complexity. Enhancing reproducibility and integrating multi-omics approaches may offer better insights. In conclusion, the field should evolve toward high-throughput, automated methods, continuously adapting research, and clinical practices.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

miss-SNF: a multimodal patient similarity network integration approach to handle completely missing data sources

Abstract Motivation Precision medicine leverages patient-specific multimodal data to improve prevention, diagnosis, prognosis, and treatment of diseases. Advancing precision medicine requires the non-trivial integration of complex, heterogeneous, and potentially high-dimensional data sources, such as multi-omics and clinical data. In the literature, several approaches have been proposed to manage missing data, but are usually limited to the recovery of subsets of features for a subset of patients. A largely overlooked problem is the integration of multiple sources of data when one or more of them are completely missing for a subset of patients, a relatively common condition in clinical practice. Results We propose miss-Similarity Network Fusion (miss-SNF), a novel general-purpose data integration approach designed to manage completely missing data in the context of patient similarity networks. miss-SNF integrates incomplete unimodal patient similarity networks by leveraging a non-linear message-passing strategy borrowed from the SNF algorithm. miss-SNF is able to recover missing patient similarities and is “task agnostic”, in the sense that can integrate partial data for both unsupervised and supervised prediction tasks. Experimental analyses on nine cancer datasets from The Cancer Genome Atlas (TCGA) demonstrate that miss-SNF achieves state-of-the-art results in recovering similarities and in identifying patients subgroups enriched in clinically relevant variables and having differential survival. Moreover, amputation experiments show that miss-SNF supervised prediction of cancer clinical outcomes and Alzheimer’s disease diagnosis with completely missing data achieves results comparable to those obtained when all the data are available. Availability and implementation miss-SNF code, implemented in R, is available at https://github.com/AnacletoLAB/missSNF.

Biochemistry & Molecular Biology↗
DOE OSTI2024

Gut microbiota carbon and sulfur metabolisms support Salmonella infections

Abstract Salmonella enterica serovar Typhimurium is a pervasive enteric pathogen and ongoing global threat to public health. Ecological studies in the Salmonella impacted gut remain underrepresented in the literature, discounting microbiome mediated interactions that may inform Salmonella physiology during colonization and infection. To understand the microbial ecology of Salmonella remodeling of the gut microbiome, we performed multi-omics on fecal microbial communities from untreated and Salmonella-infected mice. Reconstructed genomes recruited metatranscriptomic and metabolomic data providing a strain-resolved view of the expressed metabolisms of the microbiome during Salmonella infection. These data informed possible Salmonella interactions with members of the gut microbiome that were previously uncharacterized. Salmonella-induced inflammation significantly reduced the diversity of genomes that recruited transcripts in the gut microbiome, yet increased transcript mapping was observed for seven members, among which Luxibacter and Ligilactobacillus transcript read recruitment was most prevalent. Metatranscriptomic insights from Salmonella and other persistent taxa in the inflamed microbiome further expounded the necessity for oxidative tolerance mechanisms to endure the host inflammatory responses to infection. In the inflamed gut lactate was a key metabolite, with microbiota production and consumption reported amongst members with detected transcript recruitment. We also showed that organic sulfur sources could be converted by gut microbiota to yield inorganic sulfur pools that become oxidized in the inflamed gut, resulting in thiosulfate and tetrathionate that support Salmonella respiration. This research advances physiological microbiome insights beyond prior amplicon-based approaches, with the transcriptionally active organismal and metabolic pathways outlined here offering intriguing intervention targets in the Salmonella-infected intestine.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Time‐series multi‐omics analysis of micronutrient stress in Sorghum bicolor reveals iron and zinc crosstalk and regulatory network conservation

Micronutrient stress impacts growth, biomass production, and grain yield in crops. Multi-omics studies are valuable resources in identifying genes for functional studies and trait improvement, such as accumulation of Fe or Zn under deficient or excess conditions for bioenergy or grain agriculture. We conducted transcriptomics and ionomics analyses on Sorghum bicolor BTx623, grown under Fe and Zn limited and excess conditions over a 21-day period. To identify early and late transcriptional response in roots and leaves, 180 RNAseq libraries were sequenced for differential expression and co-expression network analyses. Fe and Zn accumulation was measured using ICP-MS at each time point, and a fluorometer was used to estimate chlorophyll content in leaves. Among the four treatments, Fe limitation and Zn excess resulted in the largest phenotypic effects and transcriptional response in roots and leaves. Several of the reduction (Strategy I) and chelation (Strategy II) strategy genes that improve bioavailability of Fe and Zn in plant roots often used by non-grass and grass species, respectively, were differentially expressed. Gene regulatory network (GRN) analysis of roots revealed enrichment of genes from Fe limiting and Zn excess which strongly connect to homologues of SbFIT, SbPYE, and SbBTS as hub genes. The GRN for leaf responses showed homologues of SbPYE and SbBTS as hubs connecting genes for chloroplast biosynthesis, Fe-S cluster assembly, photosynthesis, and ROS scavenging. Expression analyses suggest sorghum uses Strategy II genes for Fe and Zn uptake, as expected, but can also utilize Strategy I genes, which may be advantageous in variable moisture environments. We found strong overlap between Fe and Zn responsive GRNs, indicative of micronutrient crosstalk. We also found conservation of root and leaf GRNs, and known homologous genes suggest strong constraints on homeostasis networks in plants. These data will provide a resource for functional genetics to enhance micronutrient transport in sorghum, and opportunities to conduct further comparative GRN analysis across diverse crops species.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Capacity of Arctic fjord sediments to degrade carbohydrates from permafrost active layer

The degradation of organic matter (OM) by microorganisms in thawing permafrost produces greenhouse gases. Terrestrial OM is transported into fjords through hydrological runoff, but it is unclear whether the microbial mechanisms of OM degradation on land persist after soils enter marine environments, which differ greatly in conditions and microbial communities. This question is particularly relevant for low-OM soils, which dominate Arctic landscapes and are more exposed to oxidants. Here, we compared OM-degrading capacity in permafrost-affected active layer soils and adjacent fjord sediments from Kongsfjorden, Svalbard, focusing on carbohydrate-active enzymes (CAZymes), which target some of the most abundant types of organic matter in soils. Using multi-omics approaches—metagenomics, metagenome-assembled genomes (MAGs), metabolomics, metatranscriptomics, and metaproteomics—we examined CAZyme presence, distribution, and activity. Despite environmental differences, both soils and sediments harbored diverse glycoside hydrolases and polysaccharide lyases, most of which showed evidence of activity. Verrucomicrobia expressed the highest number of CAZyme transcripts, indicating that they dominated active carbohydrate degradation in fjord sediments, while Acidobacteria and Actinobacteria were more active in soils. Notably, CAZymes in fjord sediments targeted primarily soil-derived OM, and the proportions of enzymes degrading terrestrial OM, marine OM, and microbial necromass—remnants of dead microbial cells were similar across both environments. These results suggest that microbial communities in both soils and fjord sediments are equipped to degrade carbohydrates, and that burial of terrestrial-derived OM in fjord sediments may not protect it from microbial breakdown under Arctic warming.

Chukwufumnanya, Abuah [University of Tennessee, Kn↗
DOE OSTI2024

Device and Method for Parallel Measurement of Phosphoproteome and Proteome from Single Cells

We present the development of an immobilized metal affinity chromatography (IMAC) chip designed to enable nanoscale phosphopeptide enrichment within microfabricated nanowells. This novel platform leverages surface chemistry to immobilize high-density Nickel-Nitrilotriacetic Acid (Ni-NTA) molecules on nanowells, followed by applying Fe 3+ . The nanowell surface serves as a capture media to enrich phosphopeptides based on IMAC. The system's efficiency was validated using ß-casein as a model protein, demonstrating the chip’s capability to significantly enrich phosphopeptides. Future applications of this technology are anticipated to enable the detection of over 100 phosphopeptides from individual cells and more than 500 phosphopeptides from pools of 100 cells, offering exciting potential for single-cell phosphoproteomics. We will next apply an integrated proteomics workflow to perform multi-omics measurements, including single-cell isolation, protein digestion, and phosphopeptide enrichment, followed by LC-MS analysis of both the global proteome and phosphoproteome. Future research will explore the use of this technology to study phosphorylation dynamics in cancer cells, enhancing our understanding of cellular signaling and disease mechanisms.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Enhanced Resistance Pines for Improved Renewable Biofuel and Chemical Production (Technical Report)

We completed phenotyping constitutive and inducible oleoresin flow across two seasons, constitutive resin canal number and density and wood terpene content in our ADEPT2 and CCLONES populations. We completed genetic association between 19 oleoresin phenotypes and a total of 523,192 SNP markers from ADEPT2 and 13,883 SNP markers in CCLONES using four mixed linear models. A total of 293 significant SNPs (FDR = 0.20) were identified. We used the MENTOR tool to mine mechanistic connections from a multiplex network constructed from poplar multi-omic data to construct a conceptual model for a subset of these significant SNPs. Our model contains 6 transcriptional regulators in addition to 3 monoterpene synthases. To generate more lines of evidence for these significant SNPs, we completed a time course RNAseq experiment after inducing vascular zone cells to differentiate into new resin canals with a methyl jasmonate treatment, a single nuclei RNAseq that identified differentiating resin canal epithelial cells and are completing analysis for a QTL study in a hybrid pine population. The time course identified 4634 significantly down and 1890 significantly up regulated transcripts after treatment with methyl jasmonate, an inducer of new resin canal formation in the vascular cambial meristem. To analyze this large set of differentially regulated genes, we created a predictive expression network and analyzed it with random walk restart using 6 seed genes coding for transcription factors regulating xylem differentiation in poplar. Of the top ranked 200 transcripts, 119 transcripts were significant differentially expressed supporting these transcripts as potential candidates regulating resin canal formation. Analysis of single nuclei sequencing of shoot tips that contain differentiating resin canals, identified 10 clusters. One cluster was highly enriched in transcripts coding for 9 of the enzymes in the MEP pathway 3 prenyl synthetases, and 3 monoterpene synthases strongly suggesting that this cluster represents resin canal epithelial cells. We are mining the additional transcripts to create a trajectory analysis. In summary, we have identified > 10 novel genes that are strongly supported candidates for further analysis in breeding lines and for genetic engineering over- and under- expressing lines to increase wood terpene content to improve resistance to insect and fungal pathogens while simultaneously increasing terpene supplies for renewable chemicals and biofuels.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Creating multifunctional synthetic lichen platforms for sustainable biosynthesis of biofuel precursors

In this project, we were creating a sustainable platform for biofuel production, utilizing carbon-fixing autotrophs to supply oxygen and organic substrates to heterotrophic partners, which in turn produce carbon dioxide to feed the autotrophs. This symbiotic lichen community could lower the input cost, optimize metabolic exchanges and improve the generation of biofuel precursors through multi-omics driven genetic engineering. The cyanobacteria Synechococcus elongatus (S. elongatus) was used as the primary autotroph to provide oxygen and organic substrates, especially sucrose, to a co-culture system. The strain with overexpression of sucrose transporter cscB demonstrated a significant increase in sucrose production under salt stress as what we expected. We also implemented 13C metabolic flux analysis on the sucrose secreting strain S. elongatus cscB-NaCl. Next, transporters proteins like glutamate exporter mscCG from Corynebacterium glutamicum was overexpressed in S. elongatus to improve metabolite exchange.

Betenbaugh, Michael↗
DOE OSTI2025

Moisture Metaphenome Incubation Analysis Results

The Birch effect, a pulse of CO2 release that occurs when dry soil is rewet, is commonly observed, yet the underlying biogeochemistry remains elusive. Using multi-omics data, real-time mass spectrometry and modeling approaches, we investigated the molecular response to rewetting of a soil microbiome exposed to drought for one and two weeks. The microbiome response was evaluated through analysis of transcript, protein, metabolite, and respiration profiles and metabolic modeling using an enhanced version of the Metabolite Expression Metabolic Network Integration for Pathway Identification and Selection (MEMPIS) algorithm (Roy Chowdhury et al, mSystems, 2019).

Lipton, Mary S [Pacific Northwest National Laborat↗
DOE OSTI2025

Charting the state of GEMs in microalgae: progress, challenges, and innovations

Genome-scale metabolic models (GEMs) provide a systems-level framework for understanding and engineering microalgal metabolism. This review explores the evolution of GEMs in microalgae, highlighting advances in light modeling, automation, and multi-omics integration. Special emphasis is placed on Chlamydomonas reinhardtii as a model species. Limitations of current models, particularly for microalgae, are discussed, alongside promising developments in dynamic modeling and machine learning. Together, these innovations chart a path toward more predictive, adaptable GEMs that can accelerate biotechnological applications of microalgae in sustainable production systems.

Plant Sciences↗
DOE OSTI2025

Altering translation allows E. coli to overcome G-quadruplex stabilizers

The data included in this Dryad submission was collected in order to understand how the model organism* Escherichia coli* overcomes stabilized G-quadruplexes. This work involved a multi-omics approach to studying how the G-quadruplex stabilizers NMM and Braco-19 impact growth, gene importance, and mRNA/proteomic abundance in G-quadruplex stabilizing conditions.

Bacteria↗
DOE OSTI2026

Ampk alpha2 T172 activation dictates exercise performance and energy transduction in skeletal muscle

Adenosine 5′-monophosphate–activated protein kinase (AMPK) is an energetic sensor for metabolic regulation and integration. Here, we used CRISPR-Cas9 to generate nonactivatable Ampkα knock-in (KI) mice with mutation of threonine-172 phosphorylation site to alanine (T172A), circumventing the limitations of previous genetic interventions that disrupt the protein stoichiometry. KI mice of Ampkα2, but not Ampkα1, demonstrated phenotypic changes with increased fat-to-lean mass, impaired endurance exercise capacity, and diminished mitochondrial maximal respiration and conductance in skeletal muscle. Integrated temporal multiomics analysis (proteomics/phosphoproteomics/metabolomics) in skeletal muscle at rest and during exercise establishes a pleiotropic yet imperative role of Ampkα2 T172 activation for glycolytic and oxidative metabolism, mitochondrial respiration, and contractile function. There is a substantial overlap of skeletal muscle proteomic changes in Ampkα2 T172A KI mice with that of patients with type 2 diabetes. Our findings suggest that Ampkα2 T172 activation is critical for exercise performance and energy transduction in skeletal muscle and may serve as a therapeutic target for type 2 diabetes.

Bioenergetics↗
DOE OSTI2024

Clostridium autoethanogenum alters cofactor synthesis, redox metabolism, and lysine-acetylation in response to elevated H 2 :CO feedstock ratios for enhancing carbon capture efficiency

Clostridium autoethanogenum is an acetogenic bacterium that autotrophically converts carbon monoxide (CO) and carbon dioxide (CO 2 ) gases into bioproducts and fuels via the Wood–Ljungdahl pathway (WLP). To facilitate overall carbon capture efficiency, the reaction stoichiometry requires supplementation of hydrogen at an increased ratio of H 2 :CO to maximize CO 2 utilization; however, the molecular details and thus the ability to understand the mechanism of this supplementation are largely unknown. In order to elucidate the microbial physiology and fermentation where at least 75% of the carbon in ethanol comes from CO 2 , we established controlled chemostats that facilitated a novel and high (11:1) H 2 :CO uptake ratio. We compared and contrasted proteomic and metabolomics profiles to replicate continuous stirred tank reactors (CSTRs) at the same growth rate from a lower (5:1) H 2 :CO condition where ~ 50% of the carbon in ethanol is derived from CO 2 . Our hypothesis was that major changes would be observed in the hydrogenases and/or redox-related proteins and the WLP to compensate for the elevated hydrogen feed gas. Our analyses did reveal protein abundance differences between the two conditions largely related to reduction–oxidation (redox) pathways and cofactor biosynthesis, but the changes were more minor than we would have expected. While the Wood–Ljungdahl pathway proteins remained consistent across the conditions, other post-translational regulatory processes, such as lysine-acetylation, were observed and appeared to be more important for fine-tuning this carbon metabolism pathway. Metabolomic analyses showed that the increase in H 2 :CO ratio drives the organism to higher carbon dioxide utilization resulting in lower carbon storages and accumulated fatty acid metabolite levels. This research delves into the intricate dynamics of carbon fixation in C. autoethanogenum, examining the influence of highly elevated H 2 :CO ratios on metabolic processes and product outcomes. The study underscores the significance of optimizing gas feed composition for enhanced industrial efficiency, shedding light on potential mechanisms, such as post-translational modifications (PTMs), to fine-tune enzymatic activities and improve desired product yields.

09 BIOMASS FUELS↗