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209 records · Page 12

Climate adaptation and sustainability in switchgrass: exploring plant-microbe-soil interactions across continental scale environmental gradients

Less carbon-intensive energy sources are needed to reduce greenhouse gas emissions and their predicted role in climate change. There is growing interest in the potential of biofuels for meeting this need. A critical question is whether large-scale biofuel production can be sustainable over the time scales needed to mitigate our carbon debt from fossil fuel consumption. The carbon balance and ultimately the sustainability of biofuel feedstock production is the result of complex climate-coupled interactions between carbon fixation, sequestration, and release through combustion. Similarly, the long-term productivity of biofuels depends on the environmental factors limiting plant growth. These factors are often related to soil resources which involve complex interactions at the plant-microbe-soil interface impacting their availability and cycling. Our collaborative project addressed sustainable switchgrass (Panicum virgatum) production by exploring Plant Systems, Plant-Microbiome Interactions, and Ecosystem Processes through the integrating lens of Multi-Scale Modeling. Our research was based on detailed characterization of genetically diverse switchgrass genotypes planted in common gardens across a continental latitudinal gradient. The underlying theme of our Plant Systems research was the use of locally adapted plant material to explore plant function, to understand the mechanistic basis of environmental interactions, and to discover the plant genes important for adaptation and sustainability in the face of climate change. Our Plant-Microbiome Interaction project characterized the microbial communities associated with switchgrass using genomic tools. Our Ecosystem Processes research focused on carbon cycle responses at the ecosystem level using stand level plantings. Finally, our Multi-Scale Modeling helped to define conditions of a sustainable biofuel system and identify key tradeoffs between genetic diversity, productivity, and ecosystem services. Genome-wide association analyses were used to identify alleles that contribute to successful establishment and biomass production across North America. Together, our work provided a baseline analyses of the potential of switchgrass as a biofuel feedstock. Our project resulted in a number of successful outcomes. First, we were successful in collecting switchgrass germplasm across the species range, propagating the material, and establishing common garden experiments across the species range. In collaboration with DOE JGI, we successfully assembled the first tetraploid switchgrass genome and published this resource with an analyses of the genetic basis local adaptation from our gardens (Lowry et al. 2019, Lovell et al. 2021). The gardens were used to characterize the genetic architecture for a number of important plant phenotypes. Our project also conducted extensive sampling and sequencing to characterize the bacterial and fungal associates of switchgrass roots and leaves. We showed that host genotype, location, and harvesting practices can play a role in microbiome assembly (Singer et al. 2019 & 2022, Van Wallendael et al. 2020 & 2022, Edwards et al. 2023). Our ecosystem processes work created baseline dataset of carbon and nutrient cycling in realistic stand plantings of switchgrass. Data from this experiment provided new insight into the role of plant traits, phenology, and local environments in ecosystem processes like soil respiration, net-ecosystem exchange, and dynamics of soil and plant nutrients (Ricketts et al. 2023). Finally, our crop modelling experiments help to characterize the sensitivity of common modeling frameworks to parameters, identify key limiters of productivity across large geographic scales, and leverage patterns of local adaptation in prediction. Ultimately, these studies help to identify critical plant-microbe-soil traits that may be manipulated, through breeding or agronomic management, to improve the sustainability of biofuel feedstocks.

09 BIOMASS FUELS↗

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↗

Effects of Spaceflight Relevant Carbon Dioxide Levels on Pathogenesis Related Microbial Characteristics

BACKGROUND Multiple stressors in the spaceflight environment have the potential to alter microbial pathogenesis and virulence characteristics, including microgravity, radiation, and hostile/confined conditions. This combination of factors may lead to a stacking of risks, potentially creating unexpected increase in the risk of infectious disease. One understudied variable is the higher CO2 atmospheric concentration. The current Spaceflight Maximum Allowance Concentration (SMAC) for 24-hour average CO2 is 0.4% (3 mm Hg). These elevated ambient CO2 levels aboard the ISS could potentially influence the diversity and phenotypic responses of the resident microbial communities from both the spacecraft environment (air, surface, water) and crewmembers (gut, nasal, skin microbiomes). METHODS For these studies, we will evaluate the response of Staphylococcus aureus, Streptococcus pneumoniae and Enterobacter aerogenes, which are opportunistic pathogens that are of medical significance and have been or are likely to be found aboard spacecraft. These microorganisms will be cultured in both spaceflight analog and control conditions at CO2levels representing terrestrial atmospheric concentration of 0.04% (0.3 mmHg), ISS elevated concentration of 0.4% (3 mm Hg), and an unexpected elevated concentration of 1 % (7.6 mmHg) to represent a worst case scenario. DISCUSSION Overwhelming evidence from studies performed in food microbiology, marine biology and terrestrial environmental biology suggest altered gene expression, selective bacterial inhibition, increased growth and diversity, and increased antibiotic resistance of bacterial communities individually and in biofilm formation when exposed to increased levels of CO2. While many of these findings are based on levels of CO2 higher than would be found during spaceflight exploration missions, the potential for subtle changes in CO2 levels to exacerbate infectious disease risks warrants an evaluation of pathogen responses to spaceflight CO2 conditions. Our study will provide answers to spaceflight relevant atmospheric conditions, as described in the Risk of Adverse Health Effects Due to Host-Microorganism Interaction (GapMicro 103).

A A Medina-Colorado↗

Salt Distribution from Freezing Intrusions in Ice Shells on Ocean Worlds: Application to Europa

Several icy moons and dwarf planets appear to have hosted subsurface liquid water. Liquid water intruding upwards into the icy outer shells of these worlds freezes, forming ice and (from ocean solutes) non-ice solids. Here, we model concentrated aqueous solutions below 273 K to simulate the compositional evolution of freezing spherical intrusions. Starting solutions are based on five previously reported compositional end members for Europa's ocean. For moderate-pH end members dominated by chloride, sulfate, and/or carbonate, the solids formed include Ca-, Mg-, and Na-sulfates and -carbonates, as well as Na- and K-chlorides. For silica-rich, high-pH end members, abundant amorphous silica forms with, potentially, similarly abundant NaOH and KOH. We further develop a new numerical model to compute the spatial distribution of the formed solids and residual brine as freezing progresses. If non-ice solids settle to the bottom, their deposits tend to have stacked hourglass shapes, widening each time the crystallization temperature of a new solid is reached. We discuss the applicability of this model to vertical fractures and global freezing of a subsurface ocean. These results inform (i) how compositional heterogeneities may affect the thermophysical properties of ice shells, which in turn influence convective and cryovolcanic transport, (ii) the compatibility of brine pockets with physicochemical conditions suitable for microbial life, and (iii) possible measurements of compositional heterogeneities within ice shells by spacecraft such as NASA's Europa Clipper and ESA's JUICE missions. The methodology developed here is applicable to other ice-covered ocean worlds.

Planetary structure↗

Engineered plants for the production of the antioxidants arbutin and gallate

The shikimate pathway is a crucial metabolic route for the biosynthesis of numerous valuable chemicals. In this study, we engineered the shikimate pathway in plants via expression of microbial enzymes to produce the two important antioxidants gallate and arbutin. The engineered pathways utilize the aromatics protocatechuate and 4-hydroxybenzoate as metabolic intermediates. Through transient expression in Nicotiana benthamiana leaves, we first identified biosynthetic routes for the production of gallate from either chorismate or 3-dehydroshikimate. Gallate production was then achieved in Arabidopsis using a genetic background that overproduces protocatechuate and via expression of a mutated version of the 4-hydroxybenzoate hydroxylase PobA from Pseudomonas sp. Arbutin production was obtained in Arabidopsis using a genetic background that overproduces 4-hydroxybenzoate and via expression of the monooxygenase MNX1 from Candida parapsilosis. The best Arabidopsis transgenic lines accumulated gallate and arbutin in the range of 0.25 and 0.93 dry weight % (dwt%), respectively. Using sorghum for large-scale in planta production, the titers of gallate and arbutin produced from the intermediate 4-hydroxybenzoate reached 0.58 dwt% and 0.50 dwt%, respectively, in mature transgenic plants, surpassing levels typically observed in plants that naturally produce these compounds. Gallate and arbutin were readily extracted from plant tissues using methanol solvent. Analysis of extractive-free biomass showed only trace amounts of gallate and its precursors 4-hydroxybenzoate and protocatechuate crosslinked to cell walls, suggesting that they mainly occur as soluble conjugated forms stored in the vacuole. This study presents alternative synthesis routes using plant hosts for the eco-friendly production of gallate and arbutin.

Antioxidant↗

A call for caution in the biological interpretation of viral auxiliary metabolic genes

Virus-encoded auxiliary metabolic genes (AMGs) are non-essential genes that increase viral fitness by maintaining or manipulating host metabolism during infection. AMGs are intriguing from an evolutionary perspective, as most viral genomes are highly compact and have limited coding capacity for accessory genes. Advances in viral (meta)genomics have expanded the detection of putative AMGs from viruses in diverse environments. However, this has also led to many instances of misannotation due to the limitations of annotation tools, resulting in misinterpretations about the roles of some viral genes. Here, we highlight studies that support claims about AMGs with more than just function predictions for guidance on best practices. We then propose the adoption of an expanded, inclusive view of all genes auxiliary to core viral functions with the term ‘auxiliary viral genes’ (AVGs), alongside an associated eco-evolutionary framework for considering the types of analyses that can better support claims made about AVGs.

Environmental microbiology↗

GLBRC Soil Yearlong Incubation 13C-SIP-Lipidomics

Data package for Lipids represent a dynamic, yet stable pool of microbially-derived soil carbon This data is published under a CC0 license. The authors encourage data reuse and request attribution by referencing the below citations for the data packages and associated manuscript. Please cite as: Rempfert KR, Bell SL, Kasanke CP, Kyle JE, Hofmockel KS. 2025. GLBRC Soil Yearlong Incubation 13C-SIP-Lipidomics. [Data Set] PNNL DataHub. doi: Rempfert KR, Bell SL, Kasanke CP, Kyle JE, Hofmockel KS. 2025. MSV000097435: GLBRC soil yearlong incubation 13C-SIP-Lipidomics [Data Set] MassIVE. doi:10.25345/C57659T3K Rempfert KR, Bell SL, Kasanke CP, Kyle JE, Hofmockel KS. 2025. Lipids represent a dynamic, yet stable pool of microbially-derived soil carbon. In Prep This data package consists of compound-specific 13C SIP-lipidomics data from a yearlong tracer incubation experiment designed to investigate microbial lipid persistence in switchgrass bioenergy crop soils. In order to explore how lipid structure may modulate the persistence of C in soil lipids, we leveraged soils from two sites (Michigan - sandy texture, Wisconsin - silty texture) operated by the U.S. Department of Energy-funded Great Lakes Bioenergy Research Center (GLBRC). These sites had comparable climates, identical management practices, but contrasting soil textures, allowing us to assess the variability of lipid accrual or degradation in soils as well as provide insight regarding the degree to which edaphic properties may regulate the retention of soil lipids. Untargeted lipidomics analyses were performed to identify 13C-labeled lipids in the soil microbiome after long-term incubation. Soils were supplemented with 100 micrograms glucose per gram dry soil (99 atom % 13C or natural abundance for paired control) and incubated; samples were collected two months and one year after glucose addition. Lipid extracts (MPLEx) were analyzed by LC-MS/MS and identified using LIQUID. Calculation of isotopic enrichment of lipids was performed by targeted approach using TarMet to quantify lipid isotopologues and IsoCorrectoR to correct for natural abundance isotopes. Contents: Data package contents reported here are the first version and contain downstream analysis files for the raw LC-MS mass spectrometry files (.mzXML) deposited at the MassIVE database repository under accession MSV000097435 (80 experimental runs; 5.85 GB) | MassIVE DOI: 10.25345/C57659T3K. Support files include the additional data download 'Read Me' file containing data descriptor information. Reported data download contents are structured for compliance with project data sharing guidelines, community standards initiatives, and sponsor stakeholder policies supporting FAIR data principles. Data processing software, analysis tools, and data workflows are listed below corresponding to the host repository long-term location. Available Data Downloads (0.3 GB): "GLBRC soil yearlong incubation 13C-SIP-Lipidomics_readme.txt" - 'Read Me' data package content file (txt) "GLBRC_DataPackage_analysis files" - Data processing files (Rmd) and saved intermediate data processing outputs (rds, csv, xlsx) "GLBRC_13C_lipidomics_dataset.xlsx" - processed data in tabular format (xlsx) Linked Software: LIQUID LC-MS Analysis Software | 10.5281/zenodo.6459462 Lipid Mini-On Software Tools | 10.5281/zenodo.1492803 pmartR Omics Statistical Software | 10.5281/zenodo.6108667 xcms (v4.3.3) TarMet (v1.1.1) IsoCorrectoR (1.24.0) Funding Acknowledgments: This research was supported by an Early Career Research Program award funded by the U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research (OBER) Genomic Science program under FWP 68292, FWP 07880 and EMSL Exploratory Research Project 51095. A portion of this work was performed in the William R. Wiley Environmental Molecular Sciences Laboratory, a national scientific user facility sponsored by OBER and located at Pacific Northwest National Laboratory (PNNL). PNNL is a multi-program national laboratory operated by Battelle for the DOE under Contract DE-AC05-76RLO1830.

Rempfert, Kaitlin R [Pacific Northwest National La↗

Fabricated devices for performing bacterial-fungal interaction experiments across scales

Diverse and complex microbiomes are found in virtually every environment on Earth. Bacteria and fungi often co-dominate environmental microbiomes, and there is growing recognition that bacterial-fungal interactions (BFI) have significant impacts on the functioning of their associated microbiomes, environments, and hosts. Investigating BFI in vitro remains a challenge, particularly when attempting to examine interactions at multiple scales of system complexity. Fabricated devices can provide control over both biotic composition and abiotic factors within an experiment to enable the characterization of diverse BFI phenotypes such as modulation of growth rate, production of biomolecules, and alterations to physical movements. Engineered devices ranging from microfluidic chips to simulated rhizosphere systems have been and will continue to be invaluable to BFI research, and it is anticipated that such devices will continue to be developed for diverse applications in the field. This will allow researchers to address specific questions regarding the nature of BFI and how they impact larger microbiome and environmental processes such as biogeochemical cycles, plant productivity, and overall ecosystem resilience. Devices that are currently used for experimental investigations of bacteria, fungi, and BFI are discussed herein along with some of the associated challenges and several recommendations for future device design and applications.

59 BASIC BIOLOGICAL SCIENCES↗

NASA GeneLab: Open Science for Life in Space

The NASA GeneLab project capitalizes on multi-omic technologies to maximize the return on spaceflight experiments. To do this, GeneLab maintains a publicly accessible database (GLDS) that houses spaceflight and spaceflight relevant multi-omics data and collaborates with NASA principal investigators and projects to generate additional omics data. GeneLab houses more than 350 transcriptomic, proteomic, metabolomic and epigenomic datasets from plant, animal and microbial experiments, with a growing number of these having been produced by the GeneLab Sequencing Lab. The GLDS contains rich metadata about each experiment and has integrated radiation dosimetry data from experiments flown on the Space Shuttle, International Space Station, and Free Flying spacecrafts. With the increasing amount and complexity of omics data being generated, GeneLab utilizes community-defined, common models for metadata and terminology so that omics data and results are discoverable and reliably reproducible. GeneLab uses the ISA-Tab specification and semantic model for organizing and representing omics metadata. In addition to metadata standards, data files must be open-source file or common exchange formats to ensure accessibility and usability by all users. To ease data ingestion and transfer, the web-based submission tool allows PIs a user-friendly user interface to curate, organize, and publish their space relevant omics data. In the more recent years, data curation and submission portal has incorporated the FAIR principles making data findable, accessible, interoperable, and reusable. To increase reusability of data, GeneLab has implemented an effort to present processed data in the GLDS in addition to the raw omics data. The processed data will enable interpretation of the data by a larger group of students, scientists and the general public. Standard pipelines for the transformation of raw data into visualizations were developed by four GeneLab Analysis Working Groups (animals, plants, microbes, multi-omics) comprised of over 200 scientists from NASA, industry, and academia. To explore the data, the GLDS provides users various tools for data analysis, collaborative workspace for file storage and sharing, and a visualization portal. The analysis platform built using the Galaxy toolshed provides access to a broad variety of users including those with limited bioinformatics experience and students to learn how to analyze spaceflight omics data. The visualization portal takes GeneLab one step closer to data democratization by removing all bioinformatics requisites to interpret transcriptomics data hosted in the repository. To train the next generation of scientists, NASA offers training programs such as GeneLab 4 High School (GL4HS) and GeneLab 4 Universities. NLM Curation at a Scale Workshop 2022 | NASA GeneLab (GL4U) to teach students bioinformatics and computational biology methods to analyze omics data. Discoveries made using GeneLab have begun and will continue to deepen our understanding of biology, advance the field of genomics, and help to discover cures for diseases, create better diagnostic tools, and ultimately allow astronauts to better withstand the rigors of long-duration spaceflight.

GeneLab↗

Local adaptation of both plant and pathogen: an arms‐race compromise in switchgrass rust

In coevolving species, parasites locally adapt to host populations as hosts locally adapt to resist parasites. Parasites often outpace host local adaptation since they have rapid life cycles, but host diversity, the strength of selection, and external environmental influence can result in complex outcomes. To better understand local adaptation in host–parasite systems, we examined locally adapted switchgrass (Panicum virgatum), and its leaf rust pathogen (Puccinia novopanici) across a latitudinal range in North America. We grew switchgrass genotypes in 10 replicated multiyear common gardens, measuring rust severity from natural infection in a ‘host reciprocal transplant’ framework for testing local adaptation. We conducted genome-wide association mapping to identify genetic loci associated with rust severity. Genetically differentiated rust populations were locally adapted to northern and southern switchgrass, despite host local adaptation to environmental conditions in the same regions. Rust resistance was polygenic, and distinct loci were associated with rust severity in the north and south. We narrowed a previously identified large-effect quantitative trait locus for rust severity to a candidate YELLOW STRIPE-LIKE gene and linked numerous other loci to defense-related genes. Overall, our results suggest that both hosts and parasites can be simultaneously locally adapted, especially when parasites impose less selection than other environmental factors.

59 BASIC BIOLOGICAL SCIENCES↗

Host resistance of CD18 knockout mice against systemic infection with Listeria monocytogenes

Mice with targeted mutations of CD18, the common beta2 subunit of CD11/CD18 integrins, have leukocytosis, impaired transendothelial neutrophil emigration, and reduced host defense to Streptococcus pneumoniae, a gram-positive extracellular bacterium. Previous studies using blocking monoclonal antibodies suggested roles for CD18 and CD11b in hepatic neutrophil recruitment and host innate response to Listeria monocytogenes, a gram-positive intracellular bacterium. We induced systemic listeriosis in CD18 knockout (CD18-ko) and wild-type (WT) mice by tail vein injection with Listeria. By 14 days postinjection (dpi), 8 of 10 WT mice died, compared with 2 of 10 CD18-ko mice (P < 0.01). Quantitative organ culture showed that numbers of Listeria organisms in livers and spleens were similar in both groups at 20 min postinfection. By 3, 5, and 7 dpi, however, numbers of Listeria organisms were significantly lower in livers and spleens of CD18-ko mice than in WT mice. Histopathology showed that following Listeria infection, CD18-ko mice had milder inflammatory and necrotizing lesions in both spleens and livers than did WT mice. Cytokine assays indicated that baseline interleukin-1beta and granulocyte colony-stimulating factor (G-CSF) levels were higher in CD18-ko mice than in WT mice and that CD18-ko splenocytes produced higher levels of interleukin-1beta and G-CSF than WT splenocytes under the same amount of Listeria stimulation. These findings show that CD18 is not an absolute requirement for antilisterial innate immunity or hepatic neutrophil recruitment. We propose that the absence of CD18 in the mice results in the priming of innate immunity, as evidenced by elevated cytokine expression, and neutrophilic leukocytosis, which augments antilisterial defense.

Non-NASA Center↗