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DOE OSTI2022

Genome-Resolved Metaproteomics Decodes the Microbial and Viral Contributions to Coupled Carbon and Nitrogen Cycling in River Sediments

Rivers have a significant role in global carbon and nitrogen cycles, serving as a nexus for nutrient transport between terrestrial and marine ecosystems. Although rivers have a small global surface area, they contribute substantially to worldwide greenhouse gas emissions through microbially mediated processes within the river hyporheic zone. Despite this importance, research linking microbial and viral communities to specific biogeochemical reactions is still nascent in these sediment environments. To survey the metabolic potential and gene expression underpinning carbon and nitrogen biogeochemical cycling in river sediments, we collected an integrated data set of 33 metagenomes, metaproteomes, and paired metabolomes. We reconstructed over 500 microbial metagenome-assembled genomes (MAGs), which we dereplicated into 55 unique, nearly complete medium- and high-quality MAGs spanning 12 bacterial and archaeal phyla. We also reconstructed 2,482 viral genomic contigs, which were dereplicated into 111 viral MAGs (vMAGs) of >10 kb in size. As a result of integrating gene expression data with geochemical and metabolite data, we created a conceptual model that uncovered new roles for microorganisms in organic matter decomposition, carbon sequestration, nitrogen mineralization, nitrification, and denitrification. We show how these metabolic pathways, integrated through shared resource pools of ammonium, carbon dioxide, and inorganic nitrogen, could ultimately contribute to carbon dioxide and nitrous oxide fluxes from hyporheic sediments. Further, by linking viral MAGs to these active microbial hosts, we provide some of the first insights into viral modulation of river sediment carbon and nitrogen cycling.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2023

Testing the contribution of dispersal to microbial succession following a wildfire

ABSTRACT Given increased wildfire activity, there is growing interest in understanding the drivers of microbial succession after fire. Dispersal may be especially important to post-fire succession as biotic communities can be more susceptible to invasion following a disturbance. Here, we experimentally manipulated dispersal into disturbed leaf litter communities collected following a wildfire and tracked bacterial and fungal dispersal assemblages over time. We show that the identity and source of microbes immigrating into the soil surface post-fire change across time with seasonal shifts and the reemergence of aboveground vegetation. Further, dispersal significantly contributed to the reassembly of leaf litter microbial communities after the fire, producing an increasingly distinct assembly trajectory. The effect of dispersal on α-diversity and β-diversity was ecosystem dependent but, unexpectedly, influenced bacterial and fungal communities in a similar manner within ecosystems. Collectively, these results demonstrate that dispersal explicitly alters the course of microbial community succession following a wildfire and may impact bacteria and fungi in parallel ways, despite differing in traits expected to alter dispersal patterns. IMPORTANCE Identifying the mechanisms underlying microbial community succession is necessary for predicting how microbial communities, and their functioning, will respond to future environmental change. Dispersal is one mechanism expected to affect microbial succession, yet the difficult nature of manipulating microorganisms in the environment has limited our understanding of its contribution. Using a dispersal exclusion experiment, this study isolates the specific effect of environmental dispersal on bacterial and fungal community assembly over time following a wildfire. The work demonstrates the potential to quantify dispersal impacts on soil microbial communities over time and test how dispersal might further interact with other assembly processes in response to environmental change.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Proteomes reveal metabolic capabilities of Yarrowia lipolytica for biological upcycling of polyethylene into high-value chemicals

ABSTRACT Polyolefins derived from plastic wastes are recalcitrant for biological upcycling. However, chemical depolymerization of polyolefins can generate depolymerized plastic (DP) oil, comprising of a complex mixture of saturated, unsaturated, even, and odd hydrocarbons suitable for biological conversion. While DP oil contains a rich carbon and energy source, it is inhibitory to cells. Understanding and harnessing robust metabolic capabilities of microorganisms to upcycle the hydrocarbons in DP oil, both naturally and unnaturally occurring, into high-value chemicals are limited. Here, we discovered that an oleaginous yeast, Yarrowia lipolytica, undergoing short-term adaptation to DP oil robustly utilized a wide range of hydrocarbons for cell growth and production of citric acid and neutral lipids. When growing on hydrocarbons, Y. lipolytica partitioned into planktonic and oil-bound cells with each exhibiting distinct proteomes and amino acid distributions invested in establishing these proteomes. Significant proteome reallocation toward energy and lipid metabolism, belonging to 2 of the 23 Eukaryotic Orthologous Groups classes C and I, enabled robust growth of Y. lipolytica on hydrocarbons, with n-hexadecane as the preferential substrate. This investment was even higher for growth on DP oil where classes C and I were ranked one and two, respectively, and many associated proteins and pathways were expressed and upregulated including the hydrocarbon degradation pathway, Krebs cycle, glyoxylate shunt and, unexpectedly, propionate metabolism. However, a reduction in proteome allocation for protein biosynthesis, at the expense of the observed increase toward energy and lipid metabolisms, might have caused the inhibitory effect of DP oil on cell growth. IMPORTANCE Sustainable processes for biological upcycling of plastic wastes in a circular bioeconomy are needed to promote decarbonization and reduce environmental pollution due to increased plastic consumption, incineration, and landfill storage. Strain characterization and proteomic analysis revealed the robust metabolic capabilities of Yarrowia lipolytica to upcycle polyethylene into high-value chemicals. Significant proteome reallocation toward energy and lipid metabolisms was required for robust growth on hydrocarbons with n-hexadecane as the preferential substrate. However, an apparent over-investment in these same categories to utilize complex depolymerized plastic (DP) oil came at the expense of protein biosynthesis, limiting cell growth. Taken together, this study elucidates how Y. lipolytica activates its metabolism to utilize DP oil and establishes Y. lipolytica as a promising host for the upcycling of plastic wastes.

1-alkenes↗

DOE OSTI2025

Floodplain nitrifiers harbor the genetic potential for utilizing a wide range of organic nitrogen compounds

Organic compounds such as urea and cyanate can serve as nitrogen (N) sources for nitrifying microorganisms, including ammonia-oxidizing archaea (AOA) and bacteria (AOB), complete ammonia-oxidizing (comammox) bacteria, and nitrite-oxidizing bacteria (NOB). Here we investigated metagenome-assembled genomes (MAGs) for all four nitrifier guilds generated from hydrologically variable floodplain sediments of the Wind River Basin (WRB; Riverton, WY, USA) for their genetic potential to utilize organic N compounds. A vast majority of WRB nitrifier MAGs harbored urease (ure) and at least one urea transporter ( utp, urt, dur3 ). AOA were the most abundant and phylogenetically diverse nitrifiers in WRB floodplain sediments. Several AOA MAGs encoded cyanase ( cynS ), nitrilase ( nit1 ), omega-amidase ( nit2 ), nitrile hydratase ( nthA ), and genes related to purine degradation, including biuret hydrolase ( biuH ), oxamic transcarbamylase ( allFGH ), and catabolic carbamate kinase ( allK ). AOA often encoded an uncharacterized amidohydrolase collocated with biuH , rather than allophanate hydrolase ( atzF ). A small number of AOA encoded atzF , functioning in an unknown pathway. AOB and comammox were of relatively low abundance and taxonomic diversity and were present only at certain depths in WRB; however, they encoded triuret/biuret degradation genes ( trtA, biuH , and atzH ), and in comammox, these genes were also collocated with allFGHK . The genetic potential of ammonia oxidizers in the WRB floodplain suggests that organic N may support nitrification in this system. The proposed pathways for utilizing purine degradation products other than urea potentially expand the known metabolic capabilities of AOA, AOB, and comammox bacteria and reveal the possibility for cryptic N cycling between microbial community members.

floodplain↗

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↗

DOE OSTI2024

Single-colony MALDI mass spectrometry imaging reveals spatial differences in metabolite abundance between natural and cultured Trichodesmium morphotypes

Trichodesmium, a globally significant N 2 -fixing marine cyanobacterium, forms extensive surface blooms in nutrient-poor ocean regions. These blooms consist of a dynamic assemblage of Trichodesmium species that form distinct colony morphotypes and are inhabited by diverse microorganisms. Trichodesmium colony morphotypes vary in ecological niche, nutrient uptake, and organic molecule release, differentially impacting ocean carbon and nitrogen biogeochemical cycles. Here, we assessed the poorly studied spatial abundance of metabolites within and between three morphologically distinct Trichodesmium colonies collected from the Red Sea. We also compared these results with two morphotypes of the cultivable Trichodesmium strain IMS101. Using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) coupled with liquid extraction surface analysis (LESA) tandem mass spectrometry (MS2), we identified and localized a wide range of small metabolites associated with single-colony Trichodesmium morphotypes. Our untargeted MALDI-MSI approach revealed 80 unique features (metabolites) shared between Trichodesmium morphotypes. Discrimination analysis showed spatial variations in 57 shared metabolites, accounting for 62% of the observed variation between morphotypes. The greatest variations in metabolite abundance were observed between the cultured morphotypes compared to the natural colony morphotypes, suggesting substantial differences in metabolite production between the cultivable strain IMS101 and the naturally occurring colony morphotypes that the cultivable strain is meant to represent. This study highlights the variations in metabolite abundance between natural and cultured Trichodesmium morphotypes and provides valuable insights into metabolites common to morphologically distinct Trichodesmium colonies, offering a foundation for future targeted metabolomic investigations.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Drivers of stability and transience in composition-functioning links during serial propagation of litter-decomposing microbial communities

ABSTRACT Biotic factors that influence the temporal stability of microbial community functioning are an emerging research focus for the control of natural and engineered systems. The discovery of common features within community ensembles that differ in functional stability over time is a starting point to explore biotic factors. We serially propagated a suite of soil microbial communities through five generations of 28-day microcosm incubations to examine microbial community compositional and functional stability during plant litter decomposition. Using dissolved organic carbon (DOC) abundance as a target function, we hypothesized that microbial diversity, compositional stability, and associated changes in interactions would explain the relative stability of the ecosystem function between generations. Communities with initially high DOC abundance tended to converge towards a “low DOC” phenotype within two generations, but across all microcosms, functional stability between generations was highly variable. By splitting communities into two cohorts based on their relative DOC functional stability, we found that compositional shifts, diversity, and interaction network complexity were associated with the stability of DOC abundance between generations. Further, our results showed that legacy effects were important in determining compositional and functional outcomes, and we identified taxa associated with high DOC abundance. In the context of litter decomposition, achieving functionally stable communities is required to utilize soil microbiomes to increase DOC abundance and long-term terrestrial DOC sequestration as one solution to reduce atmospheric carbon dioxide concentrations. Identifying factors that stabilize function for a community of interest may improve the success of microbiome engineering applications. IMPORTANCE Microbial community functioning can be highly dynamic over time. Identifying and understanding biotic factors that control functional stability is of significant interest for natural and engineered communities alike. Using plant litter–decomposing communities as a model system, this study examined the stability of ecosystem function over time following repeated community transfers. By identifying microbial community features that are associated with stable ecosystem functions, microbial communities can be manipulated in ways that promote the consistency and reliability of the desired function, improving outcomes and increasing the utility of microorganisms.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Transcriptional Regulation of Plant Biomass Degradation and Carbohydrate Utilization Genes in the Extreme Thermophile Caldicellulosiruptor bescii

To develop functional metabolic engineering platforms for nonmodel microorganisms, a comprehensive understanding of the physiological and metabolic characteristics is critical. Caldicellulosiruptor bescii and other species in this genus have untapped potential for conversion of unpretreated plant biomass into industrial fuels and chemicals. The highly interactive and complex machinery used by C. bescii to acquire and process complex carbohydrates contained in lignocellulose was elucidated here to complement related efforts to develop a metabolic engineering platform with this bacterium.

comparative genomics↗

DOE OSTI2026

Distinguishing Leptothrix and Sphaerotilus genera by an integrated genomic-phenotypic analysis supported by new Leptothrix genomes

The Sphaerotilus-Leptothrix group of bacteria includes one of the first described microorganisms, Leptothrix ochracea, an uncultured type strain, plus isolates of Leptothrix and Sphaerotilus. This group is unified by the ability to form sheaths and oxidize metals, although L. ochracea exhibits obvious ecological, morphological, and functional differences from the rest of Sphaerotilus-Leptothrix. Recently, there have been calls to combine the group into one genus, Sphaerotilus; however, these studies lacked adequate genomic representation of L. ochracea. Here, we present a comprehensive comparative genomic analysis of the Sphaerotilus-Leptothrix group, including expanded representation of L. ochracea, a closely related novel species, Leptothrix toolikensis, and two new isolates (Leptothrix mechoopdaensis). Analysis of 38 genomes resolves three phylogenetic and functional groups: the ochracea-type Leptothrix (Group 1), the mobilis-type Leptothrix (Group 2), and Sphaerotilus (Group 3). Group 1 genomes form a separate genus based on average nucleotide identity and alignment fraction. The genomes clearly diverge from the rest of Sphaerotilus-Leptothrix in phylogeny, size, and metabolic potential. Group 1 genomes are much smaller (2.59–3.04 Mb) than those of Groups 2 (4.55–6.06 Mb) and 3 (3.94–5.07 Mb), while encoding more metal oxidases and fewer carbohydrate-active enzymes. Group 2 clusters with Group 3 phylogenetically and is similar in organic carbon metabolisms but maintains more metal oxidation genes. Group 2 members lack homogeneity in phenotype and genotype, suggesting that additional isolates and genomes are needed for confident classification. However, Group 1 genomes (L. ochracea and L. toolikensis) show clear divergence, precluding their inclusion in Sphaerotilus and supporting the retention of the genus Leptothrix.

Leptothrix↗

DOE OSTI2024

Identification and characterization of a skin microbiome on Caenorhabditis elegans suggests environmental microbes confer cuticle protection

ABSTRACT In the wild, C. elegans are emersed in environments teeming with a veritable menagerie of microorganisms. The C. elegans cuticular surface serves as a barrier and first point of contact with their microbial environments. In this study, we identify microbes from C. elegans natural habitats that associate with its cuticle, constituting a simple “skin microbiome.” We rear our animals on a modified CeMbio, mCeMbio, a consortium of ecologically relevant microbes. We first combine standard microbiological methods with an adapted micro skin-swabbing tool to describe the skin-resident bacteria on the C. elegans surface. Furthermore, we conduct 16S rRNA gene sequencing studies to identify relative shifts in the proportion of mCeMbio bacteria upon surface-sterilization, implying distinct skin- and gut-microbiomes. We find that some strains of bacteria, including Enterobacter sp. JUb101 , are primarily found on the nematode skin, while others like Stenotrophomonas indicatrix JUb19 and Ochrobactrum vermis MYb71 are predominantly found in the animal’s gut. Finally, we show that this skin microbiome promotes host cuticle integrity in harsh environments. Together, we identify a skin microbiome for the well-studied nematode model and propose its value in conferring host fitness advantages in naturalized contexts. IMPORTANCE The genetic model organism C. elegans has recently emerged as a tool for understanding host–microbiome interactions. Nearly all of these studies either focus on pathogenic or gut-resident microbes. Little is known about the existence of native, nonpathogenic skin microbes or their function. We demonstrate that members of a modified C. elegans model microbiome, mCeMbio, can adhere to the animal's cuticle and confer protection from noxious environments. We combine a novel micro-swab tool, the first 16S microbial sequencing data from relatively unperturbed C. elegans , and physiological assays to demonstrate microbially mediated protection of the skin. This work serves as a foundation to explore wild C. elegans skin microbiomes and use C. elegans as a model for skin research.

16S RNA↗

DOE OSTI2024

A shift between mineral and nonmineral sources of iron and sulfur causes proteome-wide changes in Methanosarcina barkeri

ABSTRACT Iron (Fe) and sulfur (S) are required elements for life, and changes in their availability can limit the ecological distribution and function of microorganisms. In anoxic environments, soluble Fe typically exists as ferrous iron [Fe(II)] and S as sulfide (HS − ). These species exhibit a strong affinity that ultimately drives the formation of sedimentary pyrite (FeS 2 ). Recently, paradigm-shifting studies indicate that Fe and S in FeS 2 can be made bioavailable by methanogens through a reductive dissolution process. However, the impact of the utilization of FeS 2 , as opposed to canonical Fe and S sources, on the phenotype of cells is not fully understood. Here, shotgun proteomics was utilized to measure changes in the phenotype of Methanosarcina barkeri MS grown with FeS 2 , Fe(II)/HS − , or Fe(II)/cysteine. Shotgun proteomics tracked 1,019 proteins overall, with 307 observed to change between growth conditions. Functional characterization and pathway analyses revealed these changes to be systemic and largely tangential to Fe/S metabolism. As a final step, the proteomics data were viewed with respect to previously collected transcriptomics data to deepen the analysis. Presented here is evidence that M. barkeri adopts distinct phenotypes to exploit specific sources of Fe and S in its environment. This is supported by observed protein abundance changes across broad categories of cellular biology. DNA adjacent metabolism, central carbon metabolism methanogenesis, metal trafficking, quorum sensing, and porphyrin biosynthesis pathways are all features in the phenotypic differentiation. Differences in trace metal availability attributed to complexation with HS − , either as a component of the growth medium [Fe(II)/HS − ] or generated through reduction of FeS 2 , were likely a major factor underpinning these phenotypic differences. IMPORTANCE The methanogenic archaeon Methanosarcina barkeri holds great potential for industrial bio-mining and energy generation technologies. Much of the biochemistry of this microbe is poorly understood, and its characterization will provide a glimpse into biological processes that evolved close to life’s origin. The discovery of its ability to extract iron and sulfur from bulk, solid-phase minerals shifted a longstanding paradigm that these elements were inaccessible to biological systems. The full elucidation of this process has the potential to help scientists and engineers extract valuable metals from low-grade ore and mine waste generating energy in the form of methane while doing so.

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 OSTI2023

Microbial Drivers of Plant Performance during Drought Depend upon Community Composition and the Greater Soil Environment

The increasing occurrence of drought is a global challenge that threatens food security through direct impacts to both plants and their interacting soil microorganisms. Plant growth promoting microbes are increasingly being harnessed to improve plant performance under stress. However, the magnitude of microbiome impacts on both structural and physiological plant traits under water limited and water replete conditions are not well-characterized. Using two microbiomes sourced from a ponderosa pine forest and an agricultural field, we performed a greenhouse experiment that used a crossed design to test the individual and combined effects of the water availability and the soil microbiome composition on plant performance. Specifically, we studied the structural and leaf functional traits of maize that are relevant to drought tolerance. We further examined how microbial relationships with plant phenotypes varied under different combinations of microbial composition and water availability. We found that water availability and microbial composition affected plant structural traits. Surprisingly, they did not alter leaf function. Maize grown in the forest-soil microbiome produced larger plants under well-watered and water-limited conditions, compared to an agricultural soil community. Although leaf functional traits were not significantly different between the watering and microbiome treatments, the bacterial composition and abundance explained significant variability in both plant structure and leaf function within individual treatments, especially water-limited plants. Our results suggest that bacteria-plant interactions that promote plant performance under stress depend upon the greater community composition and the abiotic environment.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Application of quasimetagenomics methods to define microbial diversity and subtype Listeria monocytogenes in dairy and seafood production facilities

Microorganisms frequently colonize surfaces within food production facilities. Detection of Listeria monocytogenes in this setting relies on culture-dependent methods, but the complex dynamics of bacterial interactions within these environments and their impact on pathogen detection remain largely unexplored. To address this challenge, we applied both 16S rRNA and shotgun quasimetagenomic (enriched microbiome) sequencing of swab culture enrichments from five seafood and seven dairy production environments. Utilizing 16S rRNA amplicon sequencing, we observed variability between 355 samples taken from these 12 production facilities and a distinctive microbiome for each environment. With shotgun quasimetagenomic sequencing, we were able to assemble L. monocytogenes metagenome-assembled genomes (MAGs) from 28 of the 32 culture-positive samples. We compared these MAGs to their corresponding whole-genome sequencing assemblies, which resulted in two polyphyletic clades consisting of L. monocytogenes lineages I and II with 13,195 and 25,556 single-nucleotide polymorphism sites, respectively. The remaining four MAGs did not produce sufficient genome coverage. To understand and establish limits for pathogen detection and subtyping using shotgun quasimetagenomics, these same data sets were downsampled in slilico to produce a titration series of abundances of L. monocytogenes and analyzed. Pathogen detection was achieved for all downsampled data sets, even those with only 3× genome coverage. This study contributes to the understanding of microbial diversity within food production environments and presents insights into the level of genome coverage needed in a metagenome sequencing data set to detect, subtype, and source track a foodborne pathogen.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Diversity and Composition of Methanotroph Communities in Caves

Recent observations have shown the atmospheric greenhouse gas methane (CH 4 ) is consumed by microorganisms (methanotrophs) in caves at rates comparable to CH 4 oxidation in surface soils. Caves are abundant in karst landscapes that comprise 14% of Earth’s land surface area, and therefore may represent a potentially important, but overlooked, CH 4 sink.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Proteomic Analysis of Methanococcus voltae Grown in the Presence of Mineral and Nonmineral Sources of Iron and Sulfur

Iron sulfur (Fe-S) proteins are essential and ubiquitous across all domains of life, yet the mechanisms underpinning assimilation of iron (Fe) and sulfur (S) and biogenesis of Fe-S clusters are poorly understood. This is particularly true for anaerobic methanogenic archaea, which are known to employ more Fe-S proteins than other prokaryotes. Here, we utilized a deep proteomics analysis of Methanococcus voltae A3 cultured in the presence of either synthetic pyrite (FeS 2 ) or aqueous forms of ferrous iron and sulfide to elucidate physiological responses to growth on mineral or nonmineral sources of Fe and S. The liquid chromatography-mass spectrometry (LCMS) shotgun proteomics analysis included 77% of the predicted proteome. Through a comparative analysis of intra- and extracellular proteomes, candidate proteins associated with FeS 2 reductive dissolution, Fe and S acquisition, and the subsequent transport, trafficking, and storage of Fe and S were identified. The proteomic response shows a large and balanced change, suggesting that M. voltae makes physiological adjustments involving a range of biochemical processes based on the available nutrient source. Among the proteins differentially regulated were members of core methanogenesis, oxidoreductases, membrane proteins putatively involved in transport, Fe-S binding ferredoxin and radical S-adenosylmethionine proteins, ribosomal proteins, and intracellular proteins involved in Fe-S cluster assembly and storage. This work improves our understanding of ancient biogeochemical processes and can support efforts in biomining of minerals. Clusters of iron and sulfur are key components of the active sites of enzymes that facilitate microbial conversion of light or electrical energy into chemical bonds. The proteins responsible for transporting iron and sulfur into cells and assembling these elements into metal clusters are not well understood. Using a microorganism that has an unusually high demand for iron and sulfur, we conducted a global investigation of cellular proteins and how they change based on the mineral forms of iron and sulfur. Understanding this process will answer questions about life on early earth and has application in biomining and sustainable sources of energy.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Fungal Hyphosphere Microbiomes Are Distinct from Surrounding Substrates and Show Consistent Association Patterns

Fungi and bacteria are the most abundant and diverse organisms in soils (perhaps more so than any other habitat on earth), and together these microorganisms contribute to broad soil ecosystem processes. There is a suite of bacteria that appears consistently within the physical space called the hyphosphere, the area of influence surrounding fungal hyphae.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Accelerating large scale de novo metagenome assembly using GPUs

Metagenomic workflows involve studying uncultured microorganisms directly from the environment. These environmental samples when processed by modern sequencing machines yield large and complex datasets that exceed the capabilities of metagenomic software. The increasing sizes and complexities of datasets make a strong case for exascale-capable metagenome assemblers. However, the underlying algorithmic motifs are not well suited for GPUs. This poses a challenge since the majority of next-generation supercomputers will rely primarily on GPUs for computation. In this paper we present the first of its kind GPU-Accelerated implementation of the local assembly approach that is an integral part of a widely used large-scale metagenome assembler, MetaHipMer. Local assembly uses algorithms that induce random memory accesses and non-deterministic workloads, which make GPU offloading a challenging task. Our GPU implementation outperforms the CPU version by about 7x and boosts the performance of MetaHipMer by 42% when running on 64 Summit nodes.

Awan, Muaaz Gul↗