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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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46 records · Page 3
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Choreographing root architecture and rhizosphere interactions through synthetic biology

Abstract Climate change is driving extreme changes to the environment, posing substantial threats to global food security and bioenergy. Given the direct role of plant roots in mediating plant-environment interactions, engineering the form and function of root systems and their associated microbiota may mitigate these effects. Synthetic genetic circuits have enabled sophisticated control of gene expression in microbial systems for years and a surge of advances has heralded the extension of this approach to multicellular plant species. Targeting these tools to affect root structure, exudation, and microbe activity on root surfaces provide multiple strategies for the advancement of climate-ready crops.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Hypophosphite Is a Naturally Occurring Selective Inhibitor of Syntrophic Methanogenesis

Microbial methanogenesis is a major contributor to global warming, and methane fluxes represent a loss of energy and electrons from industrial ecosystems. The chemical space of methane control strategies is still underexplored. Most known methanogenesis inhibitors target methanogenic archaeal enzymes. However, interference with the exchange of syntrophic electron carriers (H 2 or formate) in methanogenic systems presents an additional target for methane control. Here, we show that hypophosphite (H 2 PO 2 − ), an inorganic formate analogue, is a potent and selective inhibitor of syntrophic methanogenesis versus primary fermentation in rice field sediments and cattle rumens. Hypophosphite is also generally recognized as safe and relatively nontoxic to plants and animals. Genetic screens and physiological assays in the model methanogen Methanococcus maripaludis S2 implicate formate metabolism as the target of hypophosphite inhibition. Currently, there is no known biological pathway for anaerobic hypophosphite oxidation, and hypophosphite is stable in anoxic sediments for weeks to months. Given its widespread natural occurrence, we propose that hypophosphite may modulate the carbon cycle in natural environments. Taken together, our results suggest that hypophosphite could be used as a safe, inexpensive strategy for methane control in syntrophic methanogenic ecosystems.

chemical inhibitor↗
DOE OSTI2024

TbsP and TrmB jointly regulate gapII to influence cell development phenotypes in the archaeon Haloferax volcanii

Abstract Microbial cells must continually adapt their physiology in the face of changing environmental conditions. Archaea living in extreme conditions, such as saturated salinity, represent important examples of such resilience. The model salt‐loving organism Haloferax volcanii exhibits remarkable plasticity in its morphology, biofilm formation, and motility in response to variations in nutrients and cell density. However, the mechanisms regulating these lifestyle transitions remain unclear. In prior research, we showed that the transcriptional regulator, TrmB, maintains the rod shape in the related species Halobacterium salinarum by activating the expression of enzyme‐coding genes in the gluconeogenesis metabolic pathway. In Hbt. salinarum , TrmB‐dependent production of glucose moieties is required for cell surface glycoprotein biogenesis. Here, we use a combination of genetics and quantitative phenotyping assays to demonstrate that TrmB is essential for growth under gluconeogenic conditions in Hfx. volcanii . The ∆ trmB strain rapidly accumulated suppressor mutations in a gene encoding a novel transcriptional regulator, which we name t rm B s u p pressor, or TbsP (a.k.a. “tablespoon”). TbsP is required for adhesion to abiotic surfaces (i.e., biofilm formation) and maintains wild‐type cell morphology and motility. We use functional genomics and promoter fusion assays to characterize the regulons controlled by each of TrmB and TbsP, including joint regulation of the glucose‐dependent transcription of gapII , which encodes an important gluconeogenic enzyme. We conclude that TrmB and TbsP coregulate gluconeogenesis, with downstream impacts on lifestyle transitions in response to nutrients in Hfx. volcanii .

Biochemistry & Molecular Biology↗
DOE OSTI2022

In vivo Functional Characterization of Hydrophilic X2 Modules in the Cellulosomal Scaffolding Protein

As part of free cellulases or scaffolding proteins in cellulosomes, the hydrophilic non-catalytic X2 module is widely distributed in cellulolytic Clostridia or other Firmicutes bacteria. Previous biochemical studies suggest that X2 modules might increase the solubility and substrate binding affinity of X2-bearing proteins. However, their in vivo biological functions remain elusive. Here we employed CRISPR-Cas9 editing to genetically modify X2 modules by deleting the conserved motif (NGNT) from the CipC scaffoldin. Both single and double X2 mutants (X2-N: near the N terminus of CipC; X2-C: near the C terminus of CipC) presented similar stoichiometric compositions in isolated cellulosomes as the wildtype strain (WT). These X2 mutants had an elongated adaptation stage during growth on cellulose compared to cellobiose. Compared to WT, the double mutant ΔX2-NC reduced cellulose degradation by 15% and the amount of released soluble sugars by 63%. Since single X2 mutants did not present such obvious physiological changes as ΔX2-NC, there seems to be a functional redundancy between X2 modules in CipC. The in vivo adhesion assay revealed that ΔX2-NC decreased cell attachment to cellulose by 70% but a weaker effect was also overserved in single X2 mutants. These results highlight the in vivo biological role of X2 in increasing cellulose degradation efficiency by enhancing the binding affinity between cells and cellulose, which provides new perspectives for microbial engineering.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Subacute safety assessment of recombinant Lactococcus lactis on the gut microbiota of male Sprague–Dawley rats

Abstract BACKGROUND Lactococcus lactis strain pGSMT/MG1363 is a genetically modified microorganism (GMM) that constitutively expresses human metallothionein‐I fusion protein to combine with intracellular lead. Unlike traditional probiotics, pGSMT/MG1363 lacks a history of safe use in food. Administration of microorganism could influence the gut microbial community and consequently confer health benefits or cause disadvantages to the host. To date, little has been done to assess the influence of recombinant strain pGSMT/MG1363 on the stability of gut microbiota. RESULTS Liver, testis and kidney sections of male Sprague–Dawley rats orally administered pGSMT/MG1363 for 6 weeks showed normal structure and no pathological damage. There were no adverse effects on the analyzed serum biochemical parameters between the pGSMT/MG1363 group and the MG1363 group. Principal coordinate analysis showed that, compared with the MG1363 group, the 6‐week‐old fecal gut microbiota of rats fed with pGSMT/MG1363 was not significantly different (Adonis, P = 0.802). pGSMT/MG1363 treatment for 6 weeks did not significantly change the relative abundance of gut microbiota at the phylum and genus levels in comparison with MG1363 treatment. CONCLUSION Compared to the non‐GM strain MG1363 group, administration of the recombinant strain pGSMT/MG1363 for 6 weeks showed no adverse effects on the analyzed physiological parameters and gut microbial compositions of male Sprague–Dawley rats. The results suggested that, in terms of gut microbiota stability, pGSMT/MG1363 could be considered as safe as MG1363, at least for short‐term intake. Further toxicological evaluations still need to be considered before drawing a definite conclusion concerning the safe use of pGSMT/MG1363. © 2021 Society of Chemical Industry.

Chen, Guowei↗
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 OSTI2022

Ecosystems and Networks Integrated with Genes and Molecular Assemblies (ENIGMA). Final report

The goal of the Stahl lab is to use a combination of isotopic, genomic, and activity-based analyses in the field, of selected field isolates, and defined synthetic communities to identify sources and sinks of different nitrogen species in the field, focusing on the FRC at the ORNL. The most general research objective is to characterize variants in the pathway for denitrification among field isolates and synthetic microbial assemblies as a foundation for predicting the biotic and abiotic controls of nitrogen and carbon transformations in the field. Specifically, we have focused on characterizing environmental controls of nitrous oxide emissions, an important greenhouse gas and ozone depleting molecule. Studies have mapped the flux of nitrous oxide and controlling genetic elements within the FRC subsurface, indicating the importance of different enzyme variants in nitrous oxide production and consumption in controlling surface emissions. We have developed methods to quantify and catalog the isotopic signatures imparted on nitrous oxide by different nitrous oxide reductase variants in isolated organisms and model synthetic communities. We have developed methods to quantify and catalog the nitrous oxide affinity of field relevant isolates and their biomass yield when respiring nitrous oxide on different carbon substrates. We continue to expand the collection of field-relevant microbial isolates within ENIGMA by focused enrichment and isolation from field material. These objectives have been facilitated by the development of new technologies, including: 1) the development of a multiplexed automated optical density measurement system for quantifying growth kinetics of microorganisms dependent on gaseous substrates in monoculture or gas exchange in synthetic communities and 2) the development of a microcalorimetric analysis format to quantify the impact of environmental stress on microbial maintenance energy. Since field relevant processes are mediated by assemblies of organisms interacting with each other and their local environment, we have also collaborated with other ENIGMA investigators on the assembly and characterization of synthetic communities. These studies have evaluated the role of organisms lacking a complete pathway for denitrification in the control of field processes, measuring process rates of assemblies of organisms that individually lack the full denitrification pathway but mediate the complete denitrification when growing together. These efforts have so far resulted in the publication of 18 peer reviewed papers ranging from ecological and physiological drivers of pathway optimization to field-scale process characterization.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Physiological and genomic evidence of cysteine degradation and aerobic hydrogen sulfide production in freshwater bacteria

The sulfur-containing amino acid cysteine is abundant in the environment, including in freshwater lakes. Biological cysteine degradation can result in hydrogen sulfide (H 2 S), a toxic and ecologically relevant compound that is a central player in biogeochemical cycling in aquatic environments. Here, we investigated the ecological significance of cysteine in oxic freshwater, using isolated cultures, controlled experiments, and multiomics. We screened bacterial isolates enriched from natural lake water for their ability to produce H 2 S when provided cysteine. We identified 29 isolates (Bacteroidota, Proteobacteria, and Actinobacteria) that produced H 2 S. To understand the genomic and genetic basis for cysteine degradation and H 2 S production, we further characterized three isolates using whole-genome sequencing (using a combination of short-read and long-read sequencing) and tracked cysteine and H 2 S levels over their growth ranges: Stenotrophomonas maltophilia (Gammaproteobacteria), S. bentonitica (Gammaproteobacteria), and Chryseobacterium piscium (Bacteroidota). Cysteine decreased and H 2 S increased, and all three genomes had genes involved in cysteine degradation. Finally, to assess the presence of these organisms and genes in the environment, we surveyed a 5-year time series of metagenomic data from the same isolation source (Lake Mendota, Madison, WI, USA) and identified their presence throughout the time series. Overall, our study shows that diverse isolated bacterial strains can use cysteine and produce H 2 S under oxic conditions, and we show evidence using metagenomic data that this process may occur more broadly in natural freshwater lakes. Future considerations of sulfur cycling and biogeochemistry in oxic environments should account for H 2 S production from the degradation of organosulfur compounds.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Eukaryotic genomes from a global metagenomic data set illuminate trophic modes and biogeography of ocean plankton

ABSTRACT Metagenomics is a powerful method for interpreting the ecological roles and physiological capabilities of mixed microbial communities. Yet, many tools for processing metagenomic data are neither designed to consider eukaryotes nor are they built for an increasing amount of sequence data. EukHeist is an automated pipeline to retrieve eukaryotic and prokaryotic metagenome-assembled genomes (MAGs) from large-scale metagenomic sequence data sets. We developed the EukHeist workflow to specifically process large amounts of both metagenomic and/or metatranscriptomic sequence data in an automated and reproducible fashion. Here, we applied EukHeist to the large-size fraction data (0.8–2,000 µm) from Tara Oceans to recover both eukaryotic and prokaryotic MAGs, which we refer to as TOPAZ (Tara Oceans Particle-Associated MAGs). The TOPAZ MAGs consisted of >900 environmentally relevant eukaryotic MAGs and >4,000 bacterial and archaeal MAGs. The bacterial and archaeal TOPAZ MAGs expand upon the phylogenetic diversity of likely particle- and host-associated taxa. We use these MAGs to demonstrate an approach to infer the putative trophic mode of the recovered eukaryotic MAGs. We also identify ecological cohorts of co-occurring MAGs, which are driven by specific environmental factors and putative host-microbe associations. These data together add to a number of growing resources of environmentally relevant eukaryotic genomic information. Complementary and expanded databases of MAGs, such as those provided through scalable pipelines like EukHeist, stand to advance our understanding of eukaryotic diversity through increased coverage of genomic representatives across the tree of life. IMPORTANCE Single-celled eukaryotes play ecologically significant roles in the marine environment, yet fundamental questions about their biodiversity, ecological function, and interactions remain. Environmental sequencing enables researchers to document naturally occurring protistan communities, without culturing bias, yet metagenomic and metatranscriptomic sequencing approaches cannot separate individual species from communities. To more completely capture the genomic content of mixed protistan populations, we can create bins of sequences that represent the same organism (metagenome-assembled genomes [MAGs]). We developed the EukHeist pipeline, which automates the binning of population-level eukaryotic and prokaryotic genomes from metagenomic reads. We show exciting insight into what protistan communities are present and their trophic roles in the ocean. Scalable computational tools, like EukHeist, may accelerate the identification of meaningful genetic signatures from large data sets and complement researchers’ efforts to leverage MAG databases for addressing ecological questions, resolving evolutionary relationships, and discovering potentially novel biodiversity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Global Archaeal Diversity Revealed Through Massive Data Integration: Uncovering Just Tip of Iceberg

The domain of Archaea has gathered significant interest for its ecological and biotechnological potential and its role in helping us to understand the evolutionary history of Eukaryotes. In comparison to the bacterial domain, the number of adequately described members in Archaea is relatively low, with less than 1000 species described. It is not clear whether this is solely due to the cultivation difficulty of its members or, indeed, the domain is characterized by evolutionary constraints that keep the number of species relatively low. Based on molecular evidence that bypasses the difficulties of formal cultivation and characterization, several novel clades have been proposed, enabling insights into their metabolism and physiology. Given the extent of global sampling and sequencing efforts, it is now possible and meaningful to question the magnitude of global archaeal diversity based on molecular evidence. To do so, we extracted all sequences classified as Archaea from 500 thousand amplicon samples available in public repositories. After processing through our highly conservative pipeline, we named this comprehensive resource the ‘Global Archaea Diversity’ (GAD), which encompassed nearly 3 million molecular species clusters at 97% similarity, and organized it into over 500 thousand genera and nearly 100 thousand families. Saline environments have contributed the most to the novel taxa of this previously unseen diversity. The majority of those 16S rRNA gene sequence fragments were verified by matches in metagenomic datasets from IMG/M. These findings reveal a vast and previously overlooked diversity within the Archaea, offering insights into their ecological roles and evolutionary importance while establishing a foundation for the future study and characterization of this intriguing domain of life.

59 BASIC BIOLOGICAL SCIENCES↗