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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Engineering Methanogenic Microbiomes to Redirect Flux to Biomass

In this study, we present a method for acquiring and characterizing novel microbial consortia that regulates methanogens and methanotrophs through selective cultivation and metagenomic analysis of indigenous microorganisms in the environment. In addition, we present the work performed as part of this project to model the pathways that act as limiting factors in microbial methane metabolism based on a carbon cycle model. In this report, we describe the methods for selective cultivation of methane-metabolism-related microorganisms from environmental samples, the method for monitoring their methane consumption performance, and the method and results for verifying their functions using quantitative PCR and metagenomics techniques. The microbial consortia containing methanotrophs were obtained through selective cultivation and molecular biological verification, and their methane consumption performance was evaluated. In addition, the potential of the existence of bacteriophages interacting with methane metabolism-related microorganisms was identified through metagenomic sequencing.

09 BIOMASS FUELS↗
DOE OSTI2021

Omics-to-Reactive-Transport (ORT): A workflow linking genome-scale metabolic models with reactive transport codes

Motivation: Nutrient and contaminant behavior in the subsurface are governed by multiple coupled hydrobiogeochemical processes which occur across different temporal and spatial scales. Accurate description of macroscopic system behavior requires accounting for the effects of microscopic and especially microbial processes. Microbial processes mediate precipitation and dissolution and change aqueous geochemistry, all of which impacts macroscopic system behavior. As `omics data describing microbial processes is increasingly affordable and available, novel methods for using this data quickly and effectively for improved ecosystem models are needed. Results: We propose a workflow (`Omics to Reactive Transport – ORT) for utilizing metagenomic and environmental data to describe the effect of microbiological processes in macroscopic reactive transport models. This workflow utilizes and couples two open-source software packages: KBase (a software platform for systems biology) and PFLOTRAN (a reactive transport modeling code). We describe the architecture of ORT and demonstrate an implementation using metagenomic and geochemical data from a river system. Our demonstration uses microbiological drivers of nitrification and denitrification to predict nitrogen cycling patterns which agree with those provided with generalized stoichiometries. While our example uses data from a single measurement, our workflow can be applied to spatiotemporal metagenomic datasets to allow for iterative coupling between KBASE and PFLOTRAN. Live, interactive models, which incorporate the results from this narrative into a PFLOTRAN simulation, are available (without login) at https://pflotranmodeling.paf.subsurfaceinsights.com/pflotran-simple-model/.

Rubinstein, Rebecca L↗
DOE OSTI2021

DRAM example narrative

DRAM example narrative DRAM on KBase let's anyone run annotations using DRAM in the cloud. DRAM is an annotation tool that can annotate bacterial, archaeal and viral genomes and distills those annotatios into represetations of the functional genomic potential of those organisms. If you want to read more about DRAM you can check out the GitHub, wiki and journal article. DRAM annotate assemblies In KBase Assembly objects contain nucleotide sequences from genomes or metagenomes. DRAM can predict genes and annotate their function from KBase Assembly objects which may be microbial isolate genomes, metagenome assembled genomes or metagenomes. This is done with the Annotate and Distill Assemblies with DRAM app. This app can also anntoate AssemblySet objects which contain collection of Assembly objects. It also generates a Genome object and a GenomeSet object which can be used for further analysis with other KBase apps. The full annotations and other DRAM files are also available for download in the app.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Morphological and Genomic Features of the New Klosneuvirinae Isolate Fadolivirus IHUMI-VV54

Since the discovery of Mimivirus, viruses with large genomes encoding components of the translation machinery and other cellular processes have been described as belonging to the nucleocytoplasmic large DNA viruses. Recently, genome-resolved metagenomics led to the discovery of more than 40 viruses that have been grouped together in a proposed viral subfamily named Klosneuvirinae. Members of this group had genomes of up to 2.4Mb in size and featured an expanded array of translation system genes. Yet, despite the large diversity of the Klosneuvirinae in metagenomic data, there are currently only two isolates available. Here, we report the isolation of a novel giant virus known as Fadolivirus from an Algerian sewage site and provide morphological data throughout its replication cycle in amoeba and a detailed genomic characterization. The Fadolivirus genome, which is more than 1.5Mb in size, encodes 1,452 predicted proteins and phylogenetic analyses place this viral isolate as a near relative of the metagenome assembled Klosneuvirus and Indivirus. The genome encodes for 66 tRNAs, 23 aminoacyl-tRNA synthetases and a wide range of transcription factors, surpassing Klosneuvirus and other giant viruses. The Fadolivirus genome also encodes putative vacuolar-type proton pumps with the domains D and A, potentially constituting a virus-derived system for energy generation. The successful isolation of Fadolivirus will enable future hypothesis-driven experimental studies providing deeper insights into the biology of the Klosneuvirinae.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Expanding standards in viromics: in silico evaluation of dsDNA viral genome identification, classification, and auxiliary metabolic gene curation

Viruses influence global patterns of microbial diversity and nutrient cycles. Though viral metagenomics (viromics), specifically targeting dsDNA viruses, has been critical for revealing viral roles across diverse ecosystems, its analyses differ in many ways from those used for microbes. To date, viromics benchmarking has covered read pre-processing, assembly, relative abundance, read mapping thresholds and diversity estimation, but other steps would benefit from benchmarking and standardization. Here we use in silico-generated datasets and an extensive literature survey to evaluate and highlight how dataset composition (i.e., viromes vs bulk metagenomes) and assembly fragmentation impact (i) viral contig identification tool, (ii) virus taxonomic classification, and (iii) identification and curation of auxiliary metabolic genes (AMGs). The in silico benchmarking of five commonly used virus identification tools show that gene-content-based tools consistently performed well for long (≥3 kbp) contigs, while k -mer- and blast-based tools were uniquely able to detect viruses from short (≤3 kbp) contigs. Notably, however, the performance increase of k -mer- and blast-based tools for short contigs was obtained at the cost of increased false positives (sometimes up to ~5% for virome and ~75% bulk samples), particularly when eukaryotic or mobile genetic element sequences were included in the test datasets. Furthermore, for viral classification, variously sized genome fragments were assessed using gene-sharing network analytics to quantify drop-offs in taxonomic assignments, which revealed correct assignations ranging from ~95% (whole genomes) down to ~80% (3 kbp sized genome fragments). A similar trend was also observed for other viral classification tools such as VPF-class, ViPTree and VIRIDIC, suggesting that caution is warranted when classifying short genome fragments and not full genomes. Finally, we highlight how fragmented assemblies can lead to erroneous identification of AMGs and outline a best-practices workflow to curate candidate AMGs in viral genomes assembled from metagenomes. Together, these benchmarking experiments and annotation guidelines should aid researchers seeking to best detect, classify, and characterize the myriad viruses ‘hidden’ in diverse sequence datasets.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

DOE new players Carbon cycle (2016-2021)

The overarching scientific goals of our multidisciplinary grant was to further expand understanding about the key microorganisms (players), metabolic strategies (processes), and interspecies relationships (interactions) involved in the formation and oxidation of methane in the environment. This research applied novel environmental metagenomics, transcriptomics, and proteomic techniques, state-of-the-art analytical imaging, stable isotope geochemistry, and reaction-transport modeling to address these goals and develop an ‘ecosystems level’ understanding of the factors which regulate microbial methane cycling in anoxic sedimentary ecosystems. For decades, it was believed that the obligate step in methanogenesis catalyzed by methyl coenzyme M reductase (Mcr) was limited to a specific branch of the archaeal Domain, formerly known as the Euryarchaeota. Less than a decade ago co-I Tyson’s team published a surprising metagenomic-based discovery of divergent Mcr genes in a novel uncultured phylum (Bathyarchaeota), catalyzing a major shift in thinking about the diversity of microorganisms that encode the potential for methane (or higher alkane) metabolism in anoxic environments (Evans et al., 2015). In our work here, we further expand on the groups of archaea harboring the genomic potential for methane or hydrocarbon metabolism using environmental metagenomics and new gene targeted bioinformatics techniques. We additionally advanced understanding about the terminal electron acceptors and metabolic potential supporting the anaerobic oxidation of methane (AOM) in terrestrial ecosystems, specifically focused on new lineages of ANME archaea capable of respiring manganese oxides with methane presumably using large extracellular multi-heme cytochrome complexes. New details about specific syntrophic mechanisms underlying the exchange of electrons during sulfate-coupled methane oxidation between ANME-2 archaea and their sulfate-reducing bacterial partners were also elucidated as part of this funded project. Through a series of experimental ‘omics and single cell stable isotope probing studies with incubated environmental sediment samples and a cultured model electrogenic microorganism combined with model-based predictions. Combined, this work provides strong support for the hypothesis of direct interspecies electron transfer (DIET) is the dominant syntrophic mechanism controlling the anaerobic oxidation of methane with sulfate over other proposed mechanisms and additionally illustrates important spatial constraints and the underlying physico-chemical factors influencing AOM syntrophic consortia structure for DIET and extracellular metal respiration. This collaborative multi-institutional project successfully advanced several of our milestone goals including the identification of new microbial players containing methyl coenzyme M reductases hypothesized to be central to methane or hydrocarbon cycling in anoxic environments and enhancing fundamental knowledge about the role extracellular electron transfer plays in the ecophysiology of methanotrophic archaea respiring metal oxides and in the physical and metabolic structuring of syntrophic interactions in methane-rich sedimentary ecosystems.

03 NATURAL GAS↗
DOE OSTI2021

Genomic reconstruction of fossil and living microorganisms in ancient Siberian permafrost

Abstract Background Total DNA (intracellular, iDNA and extracellular, eDNA) from ancient permafrost records the mixed genetic repository of the past and present microbial populations through geological time. Given the exceptional preservation of eDNA under perennial frozen conditions, typical metagenomic sequencing of total DNA precludes the discrimination between fossil and living microorganisms in ancient cryogenic environments. DNA repair protocols were combined with high throughput sequencing (HTS) of separate iDNA and eDNA fraction to reconstruct metagenome-assembled genomes (MAGs) from ancient microbial DNA entrapped in Siberian coastal permafrost. Results Despite the severe DNA damage in ancient permafrost, the coupling of DNA repair and HTS resulted in a total of 52 MAGs from sediments across a chronosequence (26–120 kyr). These MAGs were compared with those derived from the same samples but without utilizing DNA repair protocols. The MAGs from the youngest stratum showed minimal DNA damage and thus likely originated from viable, active microbial species. Many MAGs from the older and deeper sediment appear related to past aerobic microbial populations that had died upon freezing. MAGs from anaerobic lineages, including Asgard archaea, however exhibited minimal DNA damage and likely represent extant living microorganisms that have become adapted to the cryogenic and anoxic environments. The integration of aspartic acid racemization modeling and metaproteomics further constrained the metabolic status of the living microbial populations. Collectively, combining DNA repair protocols with HTS unveiled the adaptive strategies of microbes to long-term survivability in ancient permafrost. Conclusions Our results indicated that coupling of DNA repair protocols with simultaneous sequencing of iDNA and eDNA fractions enabled the assembly of MAGs from past and living microorganisms in ancient permafrost. The genomic reconstruction from the past and extant microbial populations expanded our understanding about the microbial successions and biogeochemical alterations from the past paleoenvironment to the present-day frozen state. Furthermore, we provided genomic insights into long-term survival mechanisms of microorganisms under cryogenic conditions through geological time. The combined strategies in this study can be extrapolated to examine other ancient non-permafrost environments and constrain the search for past and extant extraterrestrial life in permafrost and ice deposits on Mars.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

Gulf of Mexico blue hole harbors high levels of novel microbial lineages

Exploration of oxygen-depleted marine environments has consistently revealed novel microbial taxa and metabolic capabilities that expand our understanding of microbial evolution and ecology. Marine blue holes are shallow karst formations characterized by low oxygen and high organic matter content. They are logistically challenging to sample, and thus our understanding of their biogeochemistry and microbial ecology is limited. We present a metagenomic and geochemical characterization of Amberjack Hole on the Florida continental shelf (Gulf of Mexico). Dissolved oxygen became depleted at the hole’s rim (32 m water depth), remained low but detectable in an intermediate hypoxic zone (40–75 m), and then increased to a secondary peak before falling below detection in the bottom layer (80–110 m), concomitant with increases in nutrients, dissolved iron, and a series of sequentially more reduced sulfur species. Microbial communities in the bottom layer contained heretofore undocumented levels of the recently discovered phylum Woesearchaeota (up to 58% of the community), along with lineages in the bacterial Candidate Phyla Radiation (CPR). Thirty-one high-quality metagenome-assembled genomes (MAGs) showed extensive biochemical capabilities for sulfur and nitrogen cycling, as well as for resisting and respiring arsenic. One uncharacterized gene associated with a CPR lineage differentiated hypoxic from anoxic zone communities. Overall, microbial communities and geochemical profiles were stable across two sampling dates in the spring and fall of 2019. The blue hole habitat is a natural marine laboratory that provides opportunities for sampling taxa with under-characterized but potentially important roles in redox-stratified microbial processes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Development of a field-deployable qPCR assay for real-time pest monitoring in algal cultivation systems

Outdoor cultivation is commonly used to produce algal biomass for a variety of bioproducts including food, feed, fuel, pharmaceuticals, and nutraceuticals. Outdoor cultivation ponds are highly susceptible to pest pressures that may lead to periods of low productivity or even entire loss of the algal crop. Consequently, there is a need for rapid, real-time tracking of pests for early intervention to mitigate crop loss. In this work, we describe the development of a field deployable, low-cost qPCR assay for detecting both known and novel pests of a farmed eukaryotic alga species, Nannochloropsis sp. We performed a proximity guided metagenome deconvolution approach (ProxiMeta™) to discover novel pests that temporally correspond to periods of reduced pond productivity. This approach provided high-quality metagenome assemblies that were used to design qPCR probes to detect specific pests of interest. The portable qPCR assay, designed to be deployed at remote field locations, enables low-cost surveillance with a rapid (2 h) turn-around time. Frequent sampling allows for early detection and prompts intervention strategies to remedy infected ponds to minimize crop loss. The qPCR assay was used to successfully detect a known predatory bacterium within the order Bdellovibrionales both in the lab and at a remote field location. Furthermore, we assembled the genome of two novel, site-specific pests in the Saprospiraceae family and successfully designed qPCR probes that differentially detected their presence in two different pond locations. Ultimately, this assay has the potential to monitor multiple pests simultaneously and tailor targets to match likely pest infections that differ across geographical locations, helping to mitigate crop loss on a large scale.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Insight into co-hosts of nitrate reduction genes and antibiotic resistance genes in an urban river of the qinghai-tibet plateau

Microbial co-hosts of nitrate reduction genes (NRGs) and antibiotic resistance genes (ARGs) have been recently reported, but their ecology and biochemical role in urban waterways remain largely unknown. Here, we collected 69 surface water and sediment samples in the Huangshui River on the Qinghai-Tibet Plateau during the wet and dry season associated with comparatively low and high antibiotic levels. Using metagenomic sequencing, we retrieved 278 medium-to-high-quality metagenome-assembled genomes (MAGs) of NRG-ARG co-hosts, mainly belonging to the phyla Proteobacteria, Actinobacteriota, and Bacteroidota. Of microorganisms carrying ARGs, a high proportion (75.3?94.9%) also encoded NRGs, supporting nitrate reducing bacteria as dominant hosts of ARGs. Seasonal changes in antibiotic levels corresponded to significant variation in the relative abundance and community composition of NRG-ARG co-host in both water and sediments, resulting in a concomitant change in antibiotic resistance pathways. In contrast, the contribution of NRG-ARG co-hosts to nitrate reduction was stable between seasons. Further, we identify specific antibiotics (e.g., sulphonamides) and microbial taxa (e.g., Acinetobacter and Hafnia) that may disproportionately impact these relationships to serve as a basis for laboratory investigations into bioremediation strategies. Our study suggests that highly abundant nitrate reducing microorganisms in contaminated environments may also directly impact human health as carriers of antibiotic resistance.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Molecular and Dual-Isotopic Profiling of the Microbial Controls on Nitrogen Leaching in Agricultural Soils under Managed Aquifer Recharge

Nitrate (NO 3 – ) leaching is a serious health and ecological concern in global agroecosystems, particularly those under the application of agricultural-managed aquifer recharge (Ag-MAR); however, there is an absence of information on microbial controls affecting NO 3 – leaching outcomes. We combine natural dual isotopes of NO 3 – ( 15 N/ 14 N and 18 O/ 16 O) with metagenomics, quantitative polymerase chain reaction (PCR), and a threshold indicator taxa analysis (TITAN) to investigate the activities, taxon profiles, and environmental controls of soil microbiome associated with NO 3 – leaching at different depths from Californian vineyards under Ag-MAR application. The isotopic signatures demonstrated a significant priming effect (P < 0.01) of Ag-MAR on denitrification activities in the topsoil (0–10 cm), with a 12–25-fold increase of 15 N–NO 3 – and 18 O–NO 3 – after the first 24 h of flooding, followed by a sharp decrease in the enrichment of both isotopes with ~80% decline in denitrification activities thereafter. In contrast, deeper soils (60–100 cm) showed minimal or no denitrification activities over the course of Ag-MAR application, thus resulting in 10–20-fold of residual NO 3 – being leached. Metagenomic profiling and laboratory microcosm demonstrated that both nitrifying and denitrifying groups, responsible for controlling NO 3 – leaching, decreased in abundance and potential activity rates with soil depth. TITAN suggested that Nitrosocosmicus and Bradyrhizobium, as the major nitrifier and denitrifier, had the highest and lowest tipping points with regard to the NO 3 – changes (P < 0.05), respectively. Overall, our study provides new insight into specific depth limitations of microbial controls on soil NO 3 – leaching in agroecosystems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Chemolithoautotroph distributions across the subsurface of a convergent margin

Subducting oceanic crusts release fluids rich in biologically relevant compounds into the overriding plate, fueling subsurface chemolithoautotrophic ecosystems. To understand the impact of subsurface geochemistry on microbial communities, we collected fluid and sediments from 14 natural springs across a ~200 km transect across the Costa Rican convergent margin and performed shotgun metagenomics. The resulting 404 metagenome-assembled genomes (MAGs) cluster into geologically distinct regions based on MAG abundance patterns: outer forearc-only (25% of total relative abundance), forearc/arc-only (38% of total relative abundance), and delocalized (37% of total relative abundance) clusters. In the outer forearc, Thermodesulfovibrionia, Candidatus Bipolaricaulia, and Firmicutes have hydrogenotrophic sulfate reduction and Wood-Ljungdahl (WL) carbon fixation pathways. In the forearc/arc, Anaerolineae, Ca. Bipolaricaulia, and Thermodesulfovibrionia have sulfur oxidation, nitrogen cycling, microaerophilic respiration, and WL, while Aquificae have aerobic sulfur oxidation and reverse tricarboxylic acid carbon fixation pathway. Transformation-based canonical correspondence analysis shows that MAG distribution corresponds to concentrations of aluminum, iron, nickel, dissolved inorganic carbon, and phosphate. While delocalized MAGs appear surface-derived, the subsurface chemolithoautotrophic, metabolic, and taxonomic landscape varies by the availability of minerals/metals and volcanically derived inorganic carbon. Furthermore, the WL pathway persists across all samples, suggesting that this versatile, energy-efficient carbon fixation pathway helps shape convergent margin subsurface ecosystems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Plant microbiomes harbor potential to promote nutrient turnover in impoverished substrates of a Brazilian biodiversity hotspot

The substrates of the Brazilian campos rupestres, a grassland ecosystem, have extremely low concentrations of phosphorus and nitrogen, imposing restrictions to plant growth. Despite that, this ecosystem harbors almost 15% of the Brazilian plant diversity, raising the question of how plants acquire nutrients in such a harsh environment. Here, we set out to uncover the taxonomic profile, the compositional and functional differences and similarities, and the nutrient turnover potential of microbial communities associated with two plant species of the campos rupestres-dominant family Velloziaceae that grow over distinct substrates (soil and rock). Using amplicon sequencing data, we show that, despite the pronounced composition differentiation, the plant-associated soil and rock communities share a core of highly efficient colonizers that tend to be highly abundant and is enriched in 21 bacterial families. Functional investigation of metagenomes and 522 metagenome-assembled genomes revealed that the microorganisms found associated to plant roots are enriched in genes involved in organic compound intake, and phosphorus and nitrogen turnover. We show that potential for phosphorus transport, mineralization, and solubilization are mostly found within bacterial families of the shared microbiome, such as Xanthobacteraceae and Bryobacteraceae. We also detected the full repertoire of nitrogen cycle-related genes and discovered a lineage of Isosphaeraceae that acquired nitrogen-fixing potential via horizontal gene transfer and might be also involved in nitrification via a metabolic handoff association with Binataceae. We highlight that plant-associated microbial populations in the campos rupestres harbor a genetic repertoire with potential to increase nutrient availability and that the microbiomes of biodiversity hotspots can reveal novel mechanisms of nutrient turnover.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Seasonal activities of the phyllosphere microbiome of perennial crops

Understanding the interactions between plants and microorganisms can inform microbiome management to enhance crop productivity and resilience to stress. Here, we apply a genome-centric approach to identify ecologically important leaf microbiome members on replicated plots of field-grown switchgrass and miscanthus, and to quantify their activities over two growing seasons for switchgrass. We use metagenome and metatranscriptome sequencing and curate 40 medium- and high-quality metagenome-assembled-genomes (MAGs). We find that classes represented by these MAGs (Actinomycetia, Alpha- and Gamma- Proteobacteria, and Bacteroidota) are active in the late season, and upregulate transcripts for short-chain dehydrogenase, molybdopterin oxidoreductase, and polyketide cyclase. Stress-associated pathways are expressed for most MAGs, suggesting engagement with the host environment. We also detect seasonally activated biosynthetic pathways for terpenes and various non-ribosomal peptide pathways that are poorly annotated. Our findings support that leaf-associated bacterial populations are seasonally dynamic and responsive to host cues.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Complex subsurface hydrothermal fluid mixing at a submarine arc volcano supports distinct and highly diverse microbial communities

Hydrothermally active submarine volcanoes are mineral-rich biological oases contributing significantly to chemical fluxes in the deep sea, yet little is known about the microbial communities inhabiting these systems. Here we investigate the diversity of microbial life in hydrothermal deposits and their metagenomics-inferred physiology in light of the geological history and resulting hydrothermal fluid paths in the subsurface of Brothers submarine volcano north of New Zealand on the southern Kermadec arc. From metagenome-assembled genomes we identified over 90 putative bacterial and archaeal genomic families and nearly 300 previously unknown genera, many potentially endemic to this submarine volcanic environment. While magmatically influenced hydrothermal systems on the volcanic resurgent cones of Brothers volcano harbor communities of thermoacidophiles and diverse members of the superphylum “DPANN,” two distinct communities are associated with the caldera wall, likely shaped by two different types of hydrothermal circulation. The communities whose phylogenetic diversity primarily aligns with that of the cone sites and magmatically influenced hydrothermal systems elsewhere are characterized predominately by anaerobic metabolisms. These populations are probably maintained by fluids with greater magmatic inputs that have interacted with different (deeper) previously altered mineral assemblages. However, proximal (a few meters distant) communities with gene-inferred aerobic, microaerophilic, and anaerobic metabolisms are likely supported by shallower seawater-dominated circulation. Furthermore, mixing of fluids from these two distinct hydrothermal circulation systems may have an underlying imprint on the high microbial phylogenomic diversity. Collectively our results highlight the importance of considering geologic evolution and history of subsurface processes in studying microbial colonization and community dynamics in volcanic environments.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Codon bias, nucleotide selection, and genome size predict in situ bacterial growth rate and transcription in rewetted soil

In soils, the first rain after a prolonged dry period represents a major pulse event impacting soil microbial community function, yet we lack a full understanding of the genomic traits associated with the microbial response to rewetting. Genomic traits such as codon usage bias and genome size have been linked to bacterial growth in soils—however, often through measurements in culture. Here, we used metagenome-assembled genomes (MAGs) with 18 O-water stable isotope probing and metatranscriptomics to track genomic traits associated with growth and transcription of soil microorganisms over one week following rewetting of a grassland soil. We found that codon bias in ribosomal protein genes was the strongest predictor of growth rate. We also found higher growth rates in bacteria with smaller genomes, suggesting that reduced genome size enables a faster response to pulses in soil bacteria. Faster transcriptional upregulation of ribosomal protein genes was associated with high codon bias and increased nucleotide skew. We found that several of these relationships existed within phyla, indicating that these associations between genomic traits and activity could be generalized characteristics of soil bacteria. Finally, we used publicly available metagenomes to assess the distribution of codon bias across a pH gradient and found that microbial communities in higher pH soils—which are often more water limited and pulse driven—have higher codon usage bias in their ribosomal protein genes. Together, these results provide evidence that genomic characteristics affect soil microbial activity during rewetting and pose a potential fitness advantage for soil bacteria where water and nutrient availability are episodic.

59 BASIC BIOLOGICAL SCIENCES↗
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 OSTI2024

Potential for functional divergence in ectomycorrhizal fungal communities across a precipitation gradient

Abstract Functional traits influence the assembly of microbial communities, but identifying these traits in the environment has remained challenging. We studied ectomycorrhizal fungal (EMF) communities inhabiting Populus trichocarpa roots distributed across a precipitation gradient in the Pacific Northwest, USA. We profiled these communities using taxonomic (meta-barcoding) and functional (metagenomic) approaches. We hypothesized that genes involved in fungal drought-stress tolerance and fungal mediated plant water uptake would be most abundant in drier soils. We were unable to detect support for this hypothesis; instead, the abundance of genes involved in melanin synthesis, hydrophobins, aquaporins, trehalose-synthases, and other gene families exhibited no significant shifts across the gradient. Finally, we studied variation in sequence homology for certain genes, finding that fungal communities in dry soils are composed of distinct aquaporin and hydrophobin gene sequences. Altogether, our results suggest that while EMF communities exhibit significant compositional shifts across this gradient, coupled functional turnover, at least as inferred using community metagenomics is limited. Accordingly, the consequences of these distinct EMF communities on plant water uptake remain critically unknown, and future studies targeting the expression of genes involved in drought stress tolerance are required.

59 BASIC BIOLOGICAL SCIENCES↗