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At least 55 records · Page 3
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Bulk and Spatially Resolved Extracellular Metabolome of Free-Living Nitrogen Fixation

Free-living nitrogen-fixing bacteria contribute significantly to terrestrial nitrogen availability; however, the forms of nitrogen contributed by this process are poorly understood. This is in part because of inherent challenges to studying soil microorganisms in situ , such as vast differences in scale between microorganism and ecosystem and complexities of the soil system (e.g., opacity, chemical complexity).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Metabolic Remodeling during Nitrogen Fixation in Zymomonas mobilis

Biofuels and bioproducts have the potential to serve as environmentally sustainable replacements for petroleum-derived fuels and commodity molecules. Advanced fuels such as higher alcohols and isoprenoids are more suitable gasoline replacements than bioethanol.

09 BIOMASS FUELS↗
DOE OSTI2020

Associative nitrogen fixation linked with three perennial bioenergy grasses in field and greenhouse experiments

Abstract Associative nitrogen (N 2 )‐fixation (ANF) by bacteria in the root‐zone of perennial bioenergy grasses has the potential to replace or supplement N fertilizer and support sustainable production of biomass, but its application in marginal ecosystems requires further evaluation. In this study, we first combined both greenhouse and field experiments, to explore the N 2 fixation effects of three temperate feedstocks Miscanthus × giganteus (giant miscanthus, Freedom), Panicum virgatum (switchgrass, Alamo), and Saccharum sp. (energycane, Ho 02‐147). In field studies across three growing seasons, plant and soil pools of candidate feedstocks were partially composed of N derived from the atmosphere (Ndfa). Energycane, giant miscanthus, and switchgrass were estimated to derive >30%, %Ndfa. Greenhouse studies were also performed to trace isotopically labeled 15 N 2 into plant biomass and soil pools. Evidence for Ndfa was detected in all three feedstock grasses (using reference 15 N of soil, chicory, and sorghum, δ 15 N~+7.0). Isotopically labeled 15 N 2 was traced into biomass (during grass elongation stage) and soil pools. Extrapolation of rates during the 24 hr labeling period to 50 days estimated 30%–55% of plant Ndfa, with the greatest Ndfa for energycane. The findings of the field natural abundance and greenhouse 15 N 2 feeding experiments provided complementary evidence that perennial bioenergy grasses have the potential to support relatively high rates of ANF, and accumulate diazotroph‐derived N into biomass when grown on non‐fertilized soil.

09 BIOMASS FUELS↗
DOE OSTI2020

Nitrogen fixation and resorption efficiency differences among twelve upland and lowland switchgrass cultivars

In nitrogen (N)-limited terrestrial ecosystems, plants employ various strategies to acquire and conserve N, including translocation of N in perennial tissues and stimulation of N fixation in roots and soils. Switchgrass (Panicum virgatum L.) is a genotypically and phenotypically diverse perennial grass with two distinct ecotypes (lowland and upland) and numerous genotypes. It grows well in low-N soils, likely because of its ability to translocate N and to associate with N-fixing microbes, but little is known about variation in these traits among cultivars or even ecotypes. We measured N translocation, N fixation potential in roots and soils, soil net N mineralization, soil net nitrification, and biomass yields in 12 switchgrass cultivars grown in a replicated block experiment in southwestern Michigan, USA. Lowland cultivars had higher yields, rates of N translocation, soil net N mineralization, and N fixation potentials on washed, non-sterile roots, while upland cultivars exhibited higher N fixation potentials in root-free soil. N resorption efficiencies averaged 53±5 [SE] % for lowland versus 29±3% for upland cultivars. Additionally, there were significant among-cultivar differences for all response variables except mineralization and nitrification, with differences likely explained by cultivar-specific physiologies and microbial communities. Here, the ideal cultivar for biofuels is one that can maintain high yields with minimal fertilizer addition and there appear to be several cultivars that meet these criteria. In addition, results suggest substantial N cycle differences among cultivars that might be exploited by breeders to create new or improved high-yielding, N-conserving switchgrass lines.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Influence of PRE-emergence herbicides on soybean development, root nodulation and symbiotic nitrogen fixation

We report widespread occurrence of herbicide-resistant weeds has further limited effective POST-emergence herbicide options in soybean (Glycine max (L.) Merr.) leading to an increased adoption of PRE-emergence herbicides. The objective of this study was to investigate the influence of 11 commonly used PRE herbicides on soybean development, root nodulation, and symbiotic N fixation. Soybean plants were grown under greenhouse conditions in pots (10 L; 4 plants per pot) filled with silt loam soil and treated one day after planting with a labeled field rate of imazethapyr, chlorimuron-ethyl, cloransulam-methyl, metribuzin, sulfentrazone, flumioxazin, saflufenacil, acetochlor, S-metolachlor, dimethenamid-P, pyroxasulfone and no herbicide (nontreated control). Sulfentrazone reduced soybean canopy at the VC growth stage but no canopy reduction was observed at the V2 growth stage from any of the herbicide treatments. At the R2 growth stage, herbicides had no effect on soybean development (root and shoot biomass), root nodulation (# nodule per plant, nodule diameter, and nodule biomass) and symbiotic N fixation (acetylene reduction assay and 15 N natural abundance). According to our findings, although PRE herbicides may slightly affect early-season soybean development, the impacts on plant growth, root nodulation, and symbiotic N fixation were negligible. Thus, when sprayed according to the label, the benefits of PRE herbicides for weed control likely outweigh any potential concern regarding soybean development, root nodulation, and N fixation.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2020

A CRISPRi-dCas9 System for Archaea and Its Use To Examine Gene Function during Nitrogen Fixation by Methanosarcina acetivorans

CRISPR-based systems are emerging as the premier method to manipulate many cellular processes. In this study, a simple and efficient CRISPR interference (CRISPRi) system for targeted gene repression in archaea was developed. The Methanosarcina acetivorans CRISPR-Cas9 system was repurposed by replacing Cas9 with the catalytically dead Cas9 (dCas9) to generate a CRISPRi-dCas9 system for targeted gene repression. To test the utility of the system, genes involved in nitrogen (N2) fixation were targeted for dCas9-mediated repression. First, the nif operon (nifHI 1 I 2 DKEN) that encodes molybdenum nitrogenase was targeted by separate guide RNAs (gRNAs), one targeting the promoter and the other targeting nifD. Remarkably, growth of M. acetivorans with N 2 was abolished by dCas9-mediated repression of the nif operon with each gRNA. The abundance of nif transcripts was >90% reduced in both strains expressing the gRNAs, and NifD was not detected in cell lysate. Next, we targeted NifB, which is required for nitrogenase cofactor biogenesis. Expression of a gRNA targeting the coding sequence of NifB decreased nifB transcript abundance >85% and impaired but did not abolish growth of M. acetivorans with N2. Finally, to ascertain the ability to study gene regulation using CRISPRi-dCas9, nrpR1, encoding a subunit of the repressor of the nif operon, was targeted. The nrpR1 repression strain grew normally with N2 but had increased nif operon transcript abundance, consistent with NrpR1 acting as a repressor. Furthermore, these results highlight the utility of the system, whereby a single gRNA when expressed with dCas9 can block transcription of targeted genes and operons in M. acetivorans.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Nitrogen fixation as NO x using air plasma coupled with heterogeneous catalysis at atmospheric pressure

Here, this study presents insights into the use of activated Al 2 O 3 catalysts to improve the energy efficiency of NO x production in atmospheric pressure air plasma. The introduction of catalysts in the direct current glow discharge system reduces the energy cost of NO x production by up to 45% at low gas flow rates. Notably, even when positioned away from the plasma zone, the catalyst enhanced NO x production, suggesting a significant role for the catalytic activation of downstream neutral species. The study also introduced a novel approach involving an air plasma jet infused with floating catalyst powder. This method significantly increased energy efficiency at higher discharge currents, with an associated energy cost of 2.9 MJ/mol for NO x production.

70 PLASMA PHYSICS AND FUSION TECHNOLOGY↗
DOE OSTI2022

Uptake of individual and mixed per- and polyfluoroalkyl substances (PFAS) by soybean and their effects on functional genes related to nitrification, denitrification, and nitrogen fixation

Here in this study, we set up a soil-microbe-soybean system spiked with PFOA, PFOS, or a PFAS mixture of eight PFAS and investigated the distribution of PFAS in the system and impacts on the abundance and expression level of genes involved in the nitrogen (N) cycle. When soybean was exposed to the PFAS mixtures, synergistic uptake by shoots was detected. PFAS exhibited remarkable impacts on abundance of nitrification and denitrification genes in both bulk soil and rhizosphere as well as expression of N fixation gene in soybean nodules. The abundance of nitrification genes AOA and AOB amoA and denitrification gene nirK was significantly reduced (p < 0.05) in almost all treatments in bulk soil, except PFOA at 10 μg/kg. The abundance of other functional genes, such as nirS and norZ was affected differently depending on PFAS concentrations and sample location, either bulk soil or the rhizosphere. Interestingly, the N fixation gene nifH in soybean nodules was overexpressed by a PFAS mixture at 100 μg/kg. Hence, this work provided in-depth knowledge regarding the distribution of PFAS and their impacts on the N cycle for the studied system. Results from this study provide insights on assessing risks posed by individual or mixed PFAS to soybean.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Rainfall events stimulate episodic associative nitrogen fixation in switchgrass

Abstract Associative N 2 fixation (ANF) is widespread but poorly characterized, limiting our ability to estimate global inputs from N 2 fixation. In some places, ANF rates are at or below detection most of the time but occasionally and unpredictably spiking to very high rates. Here we test the hypothesis that plant phenology and rainfall events stimulate ANF episodes. We measured ANF in intact soil cores in switchgrass ( Panicum virgatum L.) in Michigan, USA. We used rain exclusion shelters to impose three rainfall treatments with each receiving 60 mm of water over a 20-day period but at different frequencies. We concurrently established a treatment that received ambient rainfall, and all four treatments were replicated four times. To assess the effects of plant phenology, we measured ANF at key phenological stages in the ambient treatment. To assess the effects of rainfall, we measured ANF immediately before and immediately after each wetting event in each treatment involving rainfall manipulation. We found that the previous day’s rainfall could explain 29% of the variation in ANF rates within the ambient treatment alone, and that bulk soil C:N ratio was also positively correlated with ANF, explaining 18% of the variation alone. Wetting events increased ANF and the magnitude of response to wetting increased with the amount of water added and decreased with the amount of inorganic N added in water. ANF episodes thus appear to be driven primarily by wetting events. Wetting events likely increase C availability, promote microbial growth, and make rhizosphere conditions conducive to ANF.

Vizza, Carmella (ORCID:0000000292690357)↗
DOE OSTI2026

Function, Structure, and Regulation of Nitrogen Fixation-like Metalloproteins for Nitrogen, Energy, Carbon, and Sulfur Metabolism

Nitrogenases (N 2 ases) and nitrogen fixation-like (NFL) systems play distinct roles in nitrogen, carbon, sulfur, and energy metabolism based on their fundamental differences in structure and metallocofactor identity. As new NFL systems have recently been identified and characterized, striking parallels and differences compared to N 2 ase structure, catalysis, and regulation have emerged. NFL systems use metallocofactors that span from simple [4Fe-4S] clusters to complex clusters akin to FeMo-co, previously only thought to occur in N 2 ase. This review describes the present state of knowledge on the function, structure, catalytic mechanisms, and regulation of NFL systems that perform distinct biological roles across all three domains of life. Recent advancements in N 2 ase spectroscopic techniques for probing metallocofactor structure and electronic states guide current and future work on how each NFL system catalyzes its specific biological reaction(s). Key knowledge gaps and needed areas of research for uncovering the specific metallocofactors and structural motifs that are at the heart of NFL system reaction specificity, along with how these systems are regulated, are discussed.

Bacteria↗
DOE OSTI2020

Microbial functional genes commonly respond to elevated carbon dioxide

Atmospheric CO 2 concentration is increasing, largely due to anthropogenic activities. Previous studies of individual free-air CO 2 enrichment (FACE) experimental sites have shown significant impacts of elevated CO 2 (eCO 2 ) on soil microbial communities; however, no common microbial response patterns have yet emerged, challenging our ability to predict ecosystem functioning and sustainability in the future eCO 2 environment. Here we analyzed 66 soil microbial communities from five FACE sites, and showed common microbial response patterns to eCO 2 , especially for key functional genes involved in carbon and nitrogen fixation (e.g., pcc/acc for carbon fixation, nifH for nitrogen fixation), carbon decomposition (e.g., amyA and pulA for labile carbon decomposition, mnp and lcc for recalcitrant carbon decomposition), and greenhouse gas emissions (e.g., mcrA for methane production, norB for nitrous oxide production) across five FACE sites. Also, the relative abundance of those key genes was generally increased and directionally associated with increased biomass, soil carbon decomposition, and soil moisture. In addition, a further literature survey of more disparate FACE experimental sites indicated increased biomass, soil carbon decay, nitrogen fixation, methane and nitrous oxide emissions, plant and soil carbon and nitrogen under eCO 2 . A conceptual framework was developed to link commonly responsive functional genes with ecosystem processes, such as pcc/acc vs. soil carbon storage, amyA/pulA/mnp/lcc vs. soil carbon decomposition, and nifH vs. nitrogen availability, suggesting that such common responses of microbial functional genes may have the potential to predict ecosystem functioning and sustainability in the future eCO 2 environment.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Regulatory response to a hybrid ancestral nitrogenase in Azotobacter vinelandii

Biological nitrogen fixation, the microbial reduction of atmospheric nitrogen to bioavailable ammonia, represents both a major limitation on biological productivity and a highly desirable engineering target for synthetic biology. However, the engineering of nitrogen fixation requires an integrated understanding of how the gene regulatory dynamics of host diazotrophs respond across sequence-function space of its central catalytic metalloenzyme, nitrogenase. Here, we interrogate this relationship by analyzing the transcriptome of Azotobacter vinelandii engineered with a phylogenetically inferred ancestral nitrogenase protein variant. The engineered strain exhibits reduced cellular nitrogenase activity but recovers wild-type growth rates following an extended lag period. We find that expression of genes within the immediate nitrogen fixation network is resilient to the introduced nitrogenase sequence-level perturbations. Rather the sustained physiological compatibility with the ancestral nitrogenase variant is accompanied by reduced expression of genes that support trace metal and electron resource allocation to nitrogenase. Our results spotlight gene expression changes in cellular processes adjacent to nitrogen fixation as productive engineering considerations to improve compatibility between remodeled nitrogenase proteins and engineered host diazotrophs.

59 BASIC BIOLOGICAL SCIENCES↗