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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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At least 1,333 records · Page 74
All sourcesNASA NTRSDOE OSTI
DOE OSTI2017

A eukaryotic nicotinate-inducible gene cluster: convergent evolution in fungi and bacteria

Nicotinate degradation has hitherto been elucidated only in bacteria. In the ascomycete Aspergillus nidulans, six loci, hxnS/AN9178 encoding the molybdenum cofactor-containing nicotinate hydroxylase, AN11197 encoding a Cys2/His2 zinc finger regulator HxnR, together with AN11196/hxnZ, AN11188/hxnY, AN11189/hxnP and AN9177/hxnT, are clustered and stringently co-induced by a nicotinate derivative and subject to nitrogen metabolite repression mediated by the GATA factor AreA. These genes are strictly co-regulated by HxnR. Within the hxnR gene, constitutive mutations map in two discrete regions. Aspergillus nidulans is capable of using nicotinate and its oxidation products 6-hydroxynicotinic acid and 2,5-dihydroxypyridine as sole nitrogen sources in an HxnR-dependent way. HxnS is highly similar to HxA, the canonical xanthine dehydrogenase (XDH), and has originated by gene duplication, preceding the origin of the Pezizomycotina. This cluster is conserved with some variations throughout the Aspergillaceae. Our results imply that a fungal pathway has arisen independently from bacterial ones. Significantly, the neo-functionalization of XDH into nicotinate hydroxylase has occurred independently from analogous events in bacteria. This work describes for the first time a gene cluster involved in nicotinate catabolism in a eukaryote and has relevance for the formation and evolution of co-regulated primary metabolic gene clusters and the microbial degradation of N-heterocyclic compounds.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Genetic Transfer in Action: Uncovering DNA Flow in an Extremophilic Microbial Community

ABSTRACT Horizontal genetic transfer (HGT) is a significant driver of genomic novelty in all domains of life. HGT has been investigated in many studies however, the focus has been on conspicuous protein‐coding DNA transfers that often prove to be adaptive in recipient organisms and are therefore fixed longer‐term in lineages. These results comprise a subclass of HGTs and do not represent exhaustive (coding and non‐coding) DNA transfer and its impact on ecology. Uncovering exhaustive HGT can provide key insights into the connectivity of genomes in communities and how these transfers may occur. In this study, we use the term frequency‐inverse document frequency (TF‐IDF) technique, that has been used successfully to mine DNA transfers within real and simulated high‐quality prokaryote genomes, to search for exhaustive HGTs within an extremophilic microbial community. We establish a pipeline for validating transfers identified using this approach. We find that most DNA transfers are within‐domain and involve non‐coding DNA. A relatively high proportion of the predicted protein‐coding HGTs appear to encode transposase activity, restriction‐modification system components, and biofilm formation functions. Our study demonstrates the utility of the TF‐IDF approach for HGT detection and provides insights into the mechanisms of recent DNA transfer.

Microbiology↗
DOE OSTI2024

Developing a pipeline to expand the genetic code of diverse bacteria for microbial engineering

Microbial biotechnologies are key to addressing grand challenges to promote human health, reverse carbon emissions, recycle mixed plastic waste, remediate contaminated soils, and achieve sustainable economies. Synthetic biology has enabled design of diverse microbes and their proteins for useful purposes, but the narrowness of the natural genetic code limits functional diversity (e.g., biosynthesis) of engineered microbes. The natural genetic code defines the fundamental rules of translating genetic information into proteins comprised of 22 ‘canonical’ amino acids. However, using a technique called genetic code expansion (GCE), the chemical properties and therefore functions of proteins can be transformed by incorporation of one or more of ~200 chemically diverse ‘non-canonical’ amino acids. The effective application of genetic code expansion in diverse microbes has the potential to revolutionize biotechnology. However, despite over 50 years of research and its transformative potential, the application of genetic code expansion has been limited to a handful of bacterial species. In this project, we will perform three tasks to both overcome the barriers that prevent wide spread adoption of GCE as molecular tool and demonstrate its potential for biotechnological applications. Specifically, we will (1) develop a genetic engineering methodology that will enable use of GCE in a broad range of bacterial hosts, (2) use high-throughput functional genomics methods to identify physiological responses to both genetic code expansion and exposure to non-canonical amino acids in three different bacteria, and (3) demonstrate an application of GCE by selectively incorporate non-canonical amino acids into surface displayed peptides such as those used for biomining.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

Metatranscriptomics and Amplicon Sequencing Reveal Mutualisms in Seagrass Microbiomes

Terrestrial plants benefit from many well-understood mutualistic relationships with root and leaf-associated microbiomes, but relatively little is known about these relationships for seagrass and other aquatic plants. We used 16S rRNA gene amplicon sequencing and metatranscriptomics to assess potential mutualisms between microorganisms and the seagrasses Zostera marina and Zostera japonica collected from mixed beds in Netarts Bay, OR, United States. The phylogenetic composition of leaf-, root-, and water column-associated bacterial communities were strikingly different, but these communities were not significantly different between plant species. Many taxa present on leaves were related to organisms capable of consuming the common plant metabolic waste product methanol, and of producing agarases, which can limit the growth of epiphytic algae. Taxa present on roots were related to organisms capable of oxidizing toxic sulfur compounds and of fixing nitrogen. Metatranscriptomic sequencing identified expression of genes involved in all of these microbial metabolic processes at levels greater than typical water column bacterioplankton, and also identified expression of genes involved in denitrification and in bacterial synthesis of the plant growth hormone indole-3-acetate. These results provide the first evidence using metatranscriptomics that seagrass microbiomes carry out a broad range of functions that may benefit their hosts, and imply that microbe–plant mutualisms support the health and growth of aquatic plants.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Migration and Distribution of Natural Organic Matter Injected into Subsurface Systems - 16523

The Savannah River Site (SRS) was one of the major U.S. Department of Energy facilities that produced large amounts of radioactive and hazardous waste during the Cold War era, a legacy of material production for defense purposes. The unlined F-Area seepage basins received approximately 1.8 billion gallons of acidic waste solutions containing low-level radioactivity from numerous isotopes generated by uranium slug and irradiated fuel processing. Sufficient quantities of uranium isotopes, I-129, Tc-99, and tritium migrated into the groundwater, creating an acidic plume with a pH between 3 and 5.5. Humic substances (HS) are complex heterogeneous mixtures of poly-dispersed materials formed by biochemical and chemical reactions during the decay and transformation of plant and microbial remains. They are known for their excellent binding capacity for metals, making them a strong candidate for remediation efforts to reduce the mobility of uranium (VI) in the subsurface. Studies showed that HS are an important ion exchange and metal-complexing ligand, carrying a large number of functional groups with high complexing capacity that can greatly affect the mobility behavior of actinides in natural systems. Hence, it is a potential in situ amendment for treating groundwater contaminated with these constituents. Column experiments were conducted to address the complexities of HS in flow-through porous media representative of the acidic aquifer at the SRS F-Area Seepage Basins. The humate source used, Huma-K, is high in humic and fulvic compounds. The column studies were done using the soil obtained from SRS's FAW-1 core-hole at a depth interval of 60'-70'. A bromide tracer test was performed to obtain the pore volume (PV), variance and Peclet number. The columns were saturated with an artificial groundwater solution prepared to mimic the groundwater at SRS that was pH adjusted to 3.5 and 5 until the effluent pH reached equilibrium. Approximately one (1) PV of 5000 ppm of Huma-K was injected into the columns at 2 ml/min to study the sorption of Huma-K on the sediment. Desorption of Huma-K was studied by injecting approximately four (4) PVs of the pH adjusted artificial groundwater solution. A faster increase in pH was observed in column 1 (saturated with pH 3.5 AGW) compared to column 2 (saturated with pH 5 AGW). After injection of 2 PV of AGW, the pH of the columns reached 6.5 and 7, respectively, for columns 1 and 2. After injecting an additional PV of AGW, the pH remained relatively stable. With an increase in pH from 3.5 to 5.0, the overall retention of HA increased from 461 mg/kg for the column with pH 3.5 to 642 mg/kg for the column with pH 5. This suggests that retention is not due to adsorption alone, but adsorption coupled with precipitation and re-dissolution of Huma-K as it flows through the columns is a possible explanation for greater retention at pH 5. This negates the use of a simple Langmuir isotherm to explain humate behavior in the acidic environment of column 1. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗
DOE OSTI2023

Engineering Rhodosporidium toruloides for production of 3-hydroxypropionic acid from lignocellulosic hydrolysate

Microbial production of valuable bioproducts is a promising route towards green and sustainable manufacturing. The oleaginous yeast, Rhodosporidium toruloides, has emerged as an attractive host for the production of biofuels and bioproducts from lignocellulosic hydrolysates. 3-hydroxypropionic acid (3HP) is an attractive platform molecule that can be used to produce a wide range of commodity chemicals. This study focuses on establishing and optimizing the production of 3HP in R. toruloides. As R. toruloides naturally has a high metabolic flux towards malonyl-CoA, we exploited this pathway to produce 3HP. Upon finding the yeast capable of catabolizing 3HP, we then implemented functional genomics and metabolomic analysis to identify the catabolic pathways. Deletion of a putative malonate semialdehyde dehydrogenase gene encoding an oxidative 3HP pathway was found to significantly reduce 3HP degradation. We further explored monocarboxylate transporters to promote 3HP transport and identified a novel 3HP transporter in Aspergillus pseudoterreus by RNA-seq and proteomics. Combining these engineering efforts with media optimization in a fed-batch fermentation resulted in 45.4 g/L 3HP production. This represents one of the highest 3HP titers reported, and the highest titer produced from lignocellulosic hydrolysate to the best of our knowledge. This work demonstrates microbial production of 3HP in R. toruloides from lignocellulosic hydrolysate at high titers, and it represents a significant step toward enabling industrial production of 3HP in the future.

3-Hydroxypropionic acid↗
DOE OSTI2014

Extracting data from the muck: deriving biological insight from complex microbial communities and non-model organisms with next generation sequencing

It is becoming increasingly clear that microbes within microbial communities, for which cultured isolates have not yet been obtained, have an immense, untapped reservoir of enzymes that could help address grand challenges in human health, energy, and sustainability. Despite the obstacles associated with culturing these microbes, recent advances in next-generation sequencing (NGS) have made it possible to explore complex microbial communities in their native context for the first time. Key to extracting meaning from rapidly growing NGS datasets are bioinformatics tools that assemble the sequence data, annotate homologous sequences and interrogate it to reveal regulatory patterns. Complementing this are advances in proteomics that can link NGS data to biological function. Finally, this combination of next generation sequencing, proteomics and bioinformatic analysis forms a powerful tool to study non-model microbes, which will transform what we know about these dynamic systems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Microbial diversity drives carbon use efficiency in a model soil

Abstract Empirical evidence for the response of soil carbon cycling to the combined effects of warming, drought and diversity loss is scarce. Microbial carbon use efficiency (CUE) plays a central role in regulating the flow of carbon through soil, yet how biotic and abiotic factors interact to drive it remains unclear. Here, we combine distinct community inocula (a biotic factor) with different temperature and moisture conditions (abiotic factors) to manipulate microbial diversity and community structure within a model soil. While community composition and diversity are the strongest predictors of CUE, abiotic factors modulated the relationship between diversity and CUE, with CUE being positively correlated with bacterial diversity only under high moisture. Altogether these results indicate that the diversity × ecosystem-function relationship can be impaired under non-favorable conditions in soils, and that to understand changes in soil C cycling we need to account for the multiple facets of global changes.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2016

Identification and genome reconstruction of abundant distinct taxa in microbiomes from one thermophilic and three mesophilic production-scale biogas plants

Biofuel production from conversion of biomass is indispensable in the portfolio of renewable energies. Complex microbial communities are involved in the anaerobic digestion process of plant material, agricultural residual products and food wastes. Analysis of the genetic potential and microbiology of communities degrading biomass to biofuels is considered to be the key to develop process optimisation strategies. Hence, due to the still incomplete taxonomic and functional characterisation of corresponding communities, new and unknown species are of special interest. Three mesophilic and one thermophilic production-scale biogas plants (BGPs) were taxonomically profiled using high-throughput 16S rRNA gene amplicon sequencing. All BGPs shared a core microbiome with the thermophilic BGP featuring the lowest diversity. However, the phyla Cloacimonetes and Spirochaetes were unique to BGPs 2 and 3, Fusobacteria were only found in BGP3 and members of the phylum Thermotogae were present only in the thermophilic BGP4. Taxonomic analyses revealed that these distinctive taxa mostly represent so far unknown species. The only exception is the dominant Thermotogae OTU featuring 16S rRNA gene sequence identity to Defluviitoga tunisiensis L3, a sequenced and characterised strain. To further investigate the genetic potential of the biogas communities, corresponding metagenomes were sequenced in a deepness of 347.5 Gbp in total. A combined assembly comprised 80.3 % of all reads and resulted in the prediction of 1.59 million genes on assembled contigs. Genome binning yielded genome bins comprising the prevalent distinctive phyla Cloacimonetes, Spirochaetes, Fusobacteria and Thermotogae. Comparative genome analyses between the most dominant Thermotogae bin and the very closely related Defluviitoga tunisiensis L3 genome originating from the same BGP revealed high genetic similarity. This finding confirmed applicability and reliability of the binning approach. The four highly covered genome bins of the other three distinct phyla showed low or very low genetic similarities to their closest phylogenetic relatives, and therefore indicated their novelty. In this study, the 16S rRNA gene sequencing approach and a combined metagenome assembly and binning approach were used for the first time on different production-scale biogas plants and revealed insights into the genetic potential and functional role of so far unknown species.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Genomic composition and dynamics among Methanomicrobiales predict adaptation to contrasting environments

Members of the order Methanomicrobiales are abundant, and sometimes dominant, hydrogenotrophic (H 2 -CO 2 utilizing) methanoarchaea in a broad range of anoxic habitats. In spite of their key roles in greenhouse gas emissions and waste conversion to methane, little is known about the physiological and genomic bases for their widespread distribution and abundance. In this work, we compared the genomes of nine diverse Methanomicrobiales strains, examined their pangenomes, reconstructed gene flow and identified genes putatively mediating their success across different habitats. Most strains slowly increased gene content whereas one, Methanocorpusculum labreanum, evidenced genome downsizing. Peat-dwelling Methanomicrobiales showed adaptations centered on improved transport of scarce inorganic nutrients and likely use H + rather than Na + transmembrane chemiosmotic gradients during energy conservation. In contrast, other Methanomicrobiales show the potential to concurrently use Na + and H + chemiosmotic gradients. Analyses also revealed that the Methanomicrobiales lack a canonical electron bifurcation system (MvhABGD) known to produce low potential electrons in other orders of hydrogenotrophic methanogens. Additional putative differences in anabolic metabolism suggest that the dynamics of interspecies electron transfer from Methanomicrobiales syntrophic partners can also differ considerably. Altogether, our findings suggest profound differences in electron trafficking in the Methanomicrobiales compared with other hydrogenotrophs, and warrant further functional evaluations.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Molecular dissection of the glutamine synthetase-GlnR nitrogen regulatory circuitry in Gram-positive bacteria

How bacteria sense and respond to nitrogen levels are central questions in microbial physiology. In Gram-positive bacteria, nitrogen homeostasis is controlled by an operon encoding glutamine synthetase (GS), a dodecameric machine that assimilates ammonium into glutamine, and the GlnR repressor. GlnR detects nitrogen excess indirectly by binding glutamine-feedback-inhibited-GS (FBI-GS), which activates its transcription-repression function. The molecular mechanisms behind this regulatory circuitry, however, are unknown. Here we describe biochemical and structural analyses of GS and FBI-GS-GlnR complexes from pathogenic and non-pathogenic Gram-positive bacteria. The structures show FBI-GS binds the GlnR C-terminal domain within its active-site cavity, juxtaposing two GlnR monomers to form a DNA-binding-competent GlnR dimer. The FBI-GS-GlnR interaction stabilizes the inactive GS conformation. Strikingly, this interaction also favors a remarkable dodecamer to tetradecamer transition in some GS, breaking the paradigm that all bacterial GS are dodecamers. These data thus unveil unique structural mechanisms of transcription and enzymatic regulation.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Bioavailable iron titrations reveal oceanic Synechococcus ecotypes optimized for different iron availabilities

Abstract The trace metal iron (Fe) controls the diversity and activity of phytoplankton across the surface oceans, a paradigm established through decades of in situ and mesocosm experimental studies. Despite widespread Fe-limitation within high-nutrient, low chlorophyll (HNLC) waters, significant contributions of the cyanobacterium Synechococcus to the phytoplankton stock can be found. Correlations among differing strains of Synechococcus across different Fe-regimes have suggested the existence of Fe-adapted ecotypes. However, experimental evidence of high- versus low-Fe adapted strains of Synechococcus is lacking, and so we investigated the transcriptional responses of microbial communities inhabiting the HNLC, sub-Antarctic region of the Southern Ocean during the Spring of 2018. Analysis of metatranscriptomes generated from on-deck incubation experiments reflecting a gradient of Fe-availabilities reveal transcriptomic signatures indicative of co-occurring Synechococcus ecotypes adapted to differing Fe-regimes. Functional analyses comparing low-Fe and high-Fe conditions point to various Fe-acquisition mechanisms that may allow persistence of low-Fe adapted Synechococcus under Fe-limitation. Comparison of in situ surface conditions to the Fe-titrations indicate ecological relevance of these mechanisms as well as persistence of both putative ecotypes within this region. This Fe-titration approach, combined with transcriptomics, highlights the short-term responses of the in situ phytoplankton community to Fe-availability that are often overlooked by examining genomic content or bulk physiological responses alone. These findings expand our knowledge about how phytoplankton in HNLC Southern Ocean waters adapt and respond to changing Fe supply.

Gilbert, Naomi E. (ORCID:0000000256915547)↗
DOE OSTI2014

Metaproteomics reveals differential modes of metabolic coupling among ubiquitous oxygen minimum zone microbes

Oxygen minimum zones (OMZs) are intrinsic water column features arising from respiratory oxygen demand during organic matter degradation in stratified marine waters. Currently OMZs are expanding due to global climate change. This expansion alters marine ecosystem function and the productivity of fisheries due to habitat compression and changes in biogeochemical cycling leading to fixed nitrogen loss and greenhouse gas production. Here we use metaproteomics to chart spatial and temporal patterns of gene expression along defined redox gradients in a seasonally anoxic fjord, Saanich Inlet to better understand microbial community responses to OMZ expansion. The expression of metabolic pathway components for nitrification, anaerobic ammonium oxidation (anammox), denitrification and inorganic carbon fixation predominantly co-varied with abundance and distribution patterns of Thaumarchaeota, Nitrospira, Planctomycetes and SUP05/ARCTIC96BD-19 Gammaproteobacteria. Within these groups, pathways mediating inorganic carbon fixation and nitrogen and sulfur transformations were differentially expressed across the redoxcline. Nitrification and inorganic carbon fixation pathways affiliated with Thaumarchaeota dominated dysoxic waters and denitrification, sulfur-oxidation and inorganic carbon fixation pathways affiliated with SUP05 dominated suboxic and anoxic waters. Nitrite-oxidation and anammox pathways affiliated with Nitrospina and Planctomycetes respectively, also exhibited redox partitioning between dysoxic and suboxic waters. The differential expression of these pathways under changing water column redox conditions has quantitative implications for coupled biogeochemical cycling linking different modes of inorganic carbon fixation with distributed nitrogen and sulfur-based energy metabolism extensible to coastal and open ocean OMZs.

Environmental Molecular Sciences Laboratory↗
DOE OSTI2022

Bacterial Community Assembly, Succession, and Metabolic Function during Outdoor Cultivation of Microchloropsis salina

Outdoor cultivation of microalgae has promising potential for renewable bioenergy, but there is a knowledge gap on the structure and function of the algal microbiome that coinhabits these ecosystems. Here, we describe the assembly mechanisms, taxonomic structure, and metabolic potential of bacteria associated with Microchloropsis salina cultivated outdoors. Open mesocosms were inoculated with algal cultures that were either free of bacteria or coincubated with one of two different strains of alga-associated bacteria and were sampled across five time points taken over multiple harvesting rounds of a 40-day experiment. Using quantitative analyses of metagenome-assembled genomes (MAGs), we tracked bacterial community compositional abundance and taxon-specific functional capacity involved in algal-bacterial interactions. One of the inoculated bacteria (Alteromonas sp.) persisted and dispersed across mesocosms, whereas the other inoculated strain (Phaeobacter gallaeciensis) disappeared by day 17 while a taxonomically similar but functionally distinct Phaeobacter strain became established. The inoculated strains were less abundant than 6 numerically dominant newly recruited taxa with functional capacities for mutualistic or saprophytic lifestyles, suggesting a generalist approach to persistence. This includes a highly abundant unclassified Rhodobacteraceae species that fluctuated between 25% and 77% of the total community. Overall, we did not find evidence for priority effects exerted by the distinct inoculum conditions; all mesocosms converged with similar microbial community compositions by the end of the experiment. Instead, we infer that the 15 total populations were retained due to host selection, as they showed high metabolic potential for algal-bacterial interactions such as recycling alga-produced carbon and nitrogen and production of vitamins and secondary metabolites associated with algal growth and senescence, including B vitamins, tropodithietic acid, and roseobacticides.

rhodobacteraceae↗
DOE OSTI2019

A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products

Diterpenoids form a diverse class of small molecule natural products that are widely distributed across the kingdoms of life and have critical biological functions in developmental processes, interorganismal interactions, and environmental adaptation. Due to these various bioactivities, many diterpenoids are also of economic importance as pharmaceuticals, food additives, biofuels, and other bioproducts. Advanced genomics and biochemical approaches have enabled a rapid increase in the knowledge of diterpenoid-metabolic genes, enzymes, and pathways. However, the structural complexity of diterpenoids and the narrow taxonomic distribution of individual compounds in often only a single species remain constraining factors for their efficient production. Availability of a broader range of metabolic enzymes now provide resources for producing diterpenoids in sufficient titers and purity to facilitate a deeper investigation of this important metabolite group. Drawing on established tools for microbial and plant-based enzyme co-expression, we present an easily operated and customizable protocol for the enzymatic production of diterpenoids in either Escherichia coli or Nicotiana benthamiana, and the purification of desired products via silica chromatography and semi-preparative HPLC. Using the group of maize (Zea mays) dolabralexin diterpenoids as an example, we highlight how modular combinations of diterpene synthase (diTPS) and cytochrome P450 monooxygenase (P450) enzymes can be used to generate different diterpenoid scaffolds. Purified compounds can be used in various downstream applications, such as metabolite structural analyses, enzyme structure-function studies, and in vitro and in planta bioactivity experiments.

09 BIOMASS FUELS↗