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DOE OSTI2024

An integrated metagenomic, metabolomic and transcriptomic survey of Populus across genotypes and environments

Abstract Bridging molecular information to ecosystem-level processes would provide the capacity to understand system vulnerability and, potentially, a means for assessing ecosystem health. Here, we present an integrated dataset containing environmental and metagenomic information from plant-associated microbial communities, plant transcriptomics, plant and soil metabolomics, and soil chemistry and activity characterization measurements derived from the model tree species Populus trichocarpa . Soil, rhizosphere, root endosphere, and leaf samples were collected from 27 different P. trichocarpa genotypes grown in two different environments leading to an integrated dataset of 318 metagenomes, 98 plant transcriptomes, and 314 metabolomic profiles that are supported by diverse soil measurements. This expansive dataset will provide insights into causal linkages that relate genomic features and molecular level events to system-level properties and their environmental influences.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

GROWdb US River Systems - Samples

GROW Overview We developed the Genome Resolved Open Watersheds database (GROWdb), which aims to increase genomic sampling and understanding of global river microbiomes. An emphasis of GROWdb is to create a publicly available and ever-expanding microbial genome database that is focused on rivers while being interoperable with databases from other ecosystems. GROWdb is based on a network-of-networks approach to move beyond a small collection of well-studied rivers, towards a spatially distributed, global network of systematic observations. GROWdb represents the first microbial, river-focused resource parsed at various scales from genes to MAGs to community level including expression and potential based measurements that will be of interest to microbiologists, ecologists, geochemists, hydrologists, and modelers. Dataset Acknowledgement GROWdb contains data from various research campaigns, please acknowledge the following data generators, as appropriate: WHONDRS derived genomes or samples - include this statement in your acknowledgements: “This study used data from the Worldwide Hydrobiogeochemistry Observation Network for Dynamic River Systems (WHONDRS) under the River Corridor Science Focus Area (SFA) at the Pacific Northwest National Laboratory (PNNL) that was generated at the U.S. Department of Energy (DOE) Joint Genome Institute User Facility. PNNL is operated by Battelle Memorial Institute for the U.S. DOE under Contract No. DE-AC05-76RL01830. The SFA is supported by the U.S. DOE, Office of Biological and Environmental Research (BER), Environmental System Science (ESS) Program.” Total Samples loaded onto this Narrative: 178 Note: Not all GROW samples may be loaded into KBase Data Availability The data underlying GROWdb are accessible across various platforms to ensure all levels of data structure are widely available. First, all reads and MAGs are publicly hosted on National Center for Biotechnology (NCBI) under Bioproject PRJNA946291. Second, all data related data presented here including MAG annotations, extended data tables, phylogenetic tree files, antibiotic resistance gene database files, and MAG abundance tables are available in Zenodo (link). Beyond the flat database files listed above, our aim for GROWdb was to maximize data use by making the data available in searchable and interactive platforms including the National Microbiome Data Collaborative (NMDC) data portal, the Department of Energy’s Systems Biology Knowledgebase (KBase), and a GROW specific user interface released here, GROWdb Explorer. Each platform provides different ways to interact with GROWdb: NMDC GROWdb formed a pilot project for the NMDC. Specifically, individual GROWdb datasets (metagenomes, metatranscriptomes, etc) are easily accessible and searchable through the NMDC data portal, where they are systematically connected to each other and to a rich suite of sample information and standard analysis results, following Findable, Accessible, Interoperable, and Reusable (FAIR) data practices. KBase GROWdb is publicly available within KBase, including samples (this Narrative), MAGs, and corresponding genome scale metabolic models. Access within KBase allows for immediate access and reuse of data, including comparison to private data using KBase’s 500+ analysis tools. Other linked narratives in KBase: GROW Metagenome Assembled Genomes (MAGs) GROW Metabolic Models GROWdb Explorer GROWdb data is also explorable through a graphical user interface built through the Colorado State University Geospatial Centroid (https://geocentroid.shinyapps.io/GROWdatabase/), allowing users to search and graph microbial and spatial data simultaneously. In summary, this microbial genome resource represents the first publicly available genome collection from rivers and offers data that can be leveraged across microbiome studies. GROWdb is an expanding repository to incorporate and unify global river multi-omic data for the future.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Long-term CRISPR locus dynamics and stable host-virus co-existence in subsurface fractured shales

Viruses are the most ubiquitous biological entities on Earth. Even so, elucidating the impact of viruses on microbial communities and associated ecosystem processes often requires identification of unambiguous host-virus linkages-an undeniable challenge in many ecosystems. Subsurface fractured shales present a unique opportunity to first make these strong linkages via spacers in CRISPR-Cas arrays and subsequently reveal complex long-term host-virus dynamics. Here, we sampled two replicated sets of fractured shale wells for nearly 800 days, resulting in 78 metagenomes from temporal sampling of six wells in the Denver-Julesburg Basin (Colorado, USA). At the community level, there was strong evidence for CRISPR-Cas defense systems being used through time and likely in response to viral interactions. Within our host genomes, represented by 202 unique MAGs, we also saw that CRISPR-Cas systems were widely encoded. Together, spacers from host CRISPR loci facilitated 2,110 CRISPR-based viral linkages across 90 host MAGs spanning 25 phyla. We observed less redundancy in host-viral linkages and fewer spacers associated with hosts from the older, more established wells, possibly reflecting enrichment of more beneficial spacers through time. Leveraging temporal patterns of host-virus linkages across differing well ages, we report how host-virus co-existence dynamics develop and converge through time, possibly reflecting selection for viruses that can evade host CRISPR-Cas systems. Together, our findings shed light on the complexities of host-virus interactions as well as long-term dynamics of CRISPR-Cas defense among diverse microbial populations.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Mass spectrometry imaging of natural carbonyl products directly from agar-based microbial interactions using 4-APEBA derivatization

Aliphatic carboxylic acids, aldehydes, and ketones play diverse roles in microbial adaptation to their microenvironment, from excretion as toxins to adaptive metabolites for membrane fluidity. However, the spatial distribution of these molecules throughout biofilms and how microbes in these environments exchange these molecules remain elusive for many of these bioactive species due to inefficient molecular imaging strategies. Herein, we apply on-tissue chemical derivatization (OTCD) using 4-(2-((4-bromophenethyl)dimethylammonio)ethoxy)benzenaminium dibromide (4-APEBA) on a co-culture of a soil bacterium (Bacillus subtilis NCIB 3610) and fungus (Fusarium sp. DS 682) grown on agar as our model system. Using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), we spatially resolved more than 300 different metabolites containing carbonyl groups within this model system. Various spatial patterns are observable in these species, which indicate possible extracellular or intercellular processes of the metabolites and their up- or downregulation during microbial interaction. The unique chemistry of our approach allowed us to bring additional confidence in accurate carbonyl identification, especially when multiple isomeric candidates were possible, and this provided the ability to generate hypotheses about the potential role of some aliphatic carbonyls in this B. subtilis/Fusarium sp. interaction. The results shown here demonstrate the utility of 4-ABEBA-based OTCD MALDI-MSI in probing interkingdom interactions directly from microbial co-cultures, and these methods will enable future microbial interaction studies with expanded metabolic coverage.

4-APEBA↗

DOE OSTI2021

Influence of shrub willow buffers strategically integrated in an Illinois corn-soybean field on soil health and microbial community composition

We report soil serves many important ecological functions and is an integral part of our existence as a society. However, concerns for soil health are growing globally, in part due to the negative impacts of agricultural management on soil resources. The production of perennial bioenergy crops on marginal land in row-crop production systems is one solution that could improve land-use efficiency and address the sustainability of cropland management. Because the relationship between crop management and the environment is complex, more research is needed to evaluate the potential benefits perennial bioenergy crop production has on soil health, as well as other ecosystem services. In this study, shrub willow buffers were strategically integrated into a corn-soybean cropping system with the main objective of reducing nitrate-N leaching from grain crop production while producing biomass for bioenergy. Two buffer systems (defined by landscape positions) were included for comparison, one on marginal land with exposure to nitrate-N leaching from upslope grain (southern plots) and one on fertile soils with less nitrate-N leaching potential (northern plots). Evaluation of soil (chemistry, bulk density, microbial community) and shrub willow vegetation properties ( fine roots, leaf litter decomposition, and nutrient uptake dynamics), showed that landscape position plays an important role in (1) the dynamics of soil chemical properties, (2) shrub willow's influence and productivity, and (3) the provision of additional ecosystem services such as reductions in nitrous oxide emissions and nitrate-N leaching. In addition, the combination of crop type and landscape position (N-grain, N-willow, S-grain, and S-willow) influenced the species composition of the soil microbial community, resulting in unique and identifiable communities. These results highlight the potential application of shrub willow buffers for ecosystem service provision and support of ecosystem processes; however, understanding the relationship between the microbial community, crop type, and landscape is important for understanding the sustainability of the design.

54 ENVIRONMENTAL SCIENCES↗

NASA NTRS2004

Microbial Characterization of Free Floating Condensate Aboard the Mir Space Station

Three samples of humidity condensate that had accumulated behind panels aboard the Russian space station Mir were collected and returned to earth for analysis. As these floating masses of liquid come into contact with the astronauts and the engineering systems, they have the potential to affect both crew health and systems performance. Using a combination of culturing techniques, a wide variety of organisms were isolated included Escherichia coli, Serratia marcescens, and a presumed Legionella species. In addition, microscopic analysis indicated the presence of protozoa, dust mites, and spirochetes. These findings suggest the need for more comprehensive microbial analysis of the environment through the use of new methodologies to allow a more thorough risk assessment of spacecraft. Copyright 2004 Springer-Verlag.

Spacecraft↗

DOE OSTI2020

A Microbe Associated with Sleep Revealed by a Novel Systems Genetic Analysis of the Microbiome in Collaborative Cross Mice

The microbiome influences health and disease through complex networks of host genetics, genomics, microbes, and environment. Identifying the mechanisms of these interactions has remained challenging. Systems genetics in laboratory mice (Mus musculus) enables data-driven discovery of biological network components and mechanisms of host–microbial interactions underlying disease phenotypes. To examine the interplay among the whole host genome, transcriptome, and microbiome, we mapped QTL and correlated the abundance of cecal messenger RNA, luminal microflora, physiology, and behavior in a highly diverse Collaborative Cross breeding population. One such relationship, regulated by a variant on chromosome 7, was the association of Odoribacter (Bacteroidales) abundance and sleep phenotypes. In a test of this association in the BKS.Cg-Dock7 m +/+ Lepr db /J mouse model of obesity and diabetes, known to have abnormal sleep and colonization by Odoribacter, treatment with antibiotics altered sleep in a genotype-dependent fashion. The many other relationships extracted from this study can be used to interrogate other diseases, microbes, and mechanisms.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Metabolome patterns identify active dechlorination in bioaugmentation consortium SDC-9™

Ultra-high performance liquid chromatography–high-resolution mass spectrometry (UPHLC–HRMS) is used to discover and monitor single or sets of biomarkers informing about metabolic processes of interest. The technique can detect 1000’s of molecules (i.e., metabolites) in a single instrument run and provide a measurement of the global metabolome, which could be a fingerprint of activity. Despite the power of this approach, technical challenges have hindered the effective use of metabolomics to interrogate microbial communities implicated in the removal of priority contaminants. Herein, our efforts to circumvent these challenges and apply this emerging systems biology technique to microbiomes relevant for contaminant biodegradation will be discussed. Chlorinated ethenes impact many contaminated sites, and detoxification can be achieved by organohalide-respiring bacteria, a process currently assessed by quantitative gene-centric tools (e.g., quantitative PCR). This laboratory study monitored the metabolome of the SDC-9™ bioaugmentation consortium during cis-1,2-dichloroethene (cDCE) conversion to vinyl chloride (VC) and nontoxic ethene. Untargeted metabolomics using an UHPLC-Orbitrap mass spectrometer and performed on SDC-9™ cultures at different stages of the reductive dechlorination process detected ~10,000 spectral features per sample arising from water-soluble molecules with both known and unknown structures. Multivariate statistical techniques including partial least squares-discriminate analysis (PLSDA) identified patterns of measurable spectral features (peak patterns) that correlated with dechlorination (in)activity, and ANOVA analyses identified 18 potential biomarkers for this process. Statistical clustering of samples with these 18 features identified dechlorination activity more reliably than clustering of samples based only on chlorinated ethene concentration and Dhc 16S rRNA gene abundance data, highlighting the potential value of metabolomic workflows as an innovative site assessment and bioremediation monitoring tool.

environmental monitoring↗

NASA NTRS1993

Origins of species: acquired genomes and individuality

Entire genomes with their accompanying protein synthetic systems are transferred throughout the biosphere primarily as bacteria and protists which become symbionts as they irreversibly integrate into pre-existing organisms to form more complex individuals. Individualization is stabilized by simultaneous transmission of once-separate heterologous genetic systems. The origin of new species is hypothesized to correlate with the acquisition, integration and subsequent inheritance of such acquired microbial genomes. These processes were recognized by Mereschkovsky ("Symbiogenesis" in Russian, 1909) and by Wallin ("Symbionticism", see p. 181, this issue).

NASA Discipline Exobiology↗

DOE OSTI2026

Interactions between molecular-scale biogeochemical processes and hyporheic exchange for understanding coupled Fe-S-C cycling in iron-rich riparian wetlands (Final Technical Report)

Riparian wetlands are dynamic interfaces that exert strong control over water quality, contaminant mobility, and greenhouse gas emissions. These systems are characterized by hyporheic exchange between oxic surface water and anoxic groundwater, which generates steep redox gradients and promotes spatially and temporally variable microbial activity. Despite growing recognition of tightly coupled iron (Fe), sulfur (S), and carbon (C) cycling in these environments, the mechanisms governing these interactions—particularly under low-sulfate freshwater conditions—remain poorly constrained. This project developed a mechanistic understanding of how hydrologic variability and microbial processes interact to control Fe–S–C cycling in iron-rich riparian wetlands. Using a multi-scale and multi-method approach integrating field observations, geochemical and spectroscopic analyses, metagenomics, and reactive transport modeling, we demonstrate that “cryptic” sulfur cycling—rapid sulfur transformations involving intermediate-valence species—plays a dominant and previously underrecognized role in freshwater wetlands. These processes persist despite low sulfate concentrations and significantly influence iron reduction, carbon mineralization, and methane dynamics. The results show that cryptic sulfur cycling enhances dissolved Fe 2+ production, regulates methane concentrations, and is strongly controlled by climate-driven hydrologic fluxes. By linking hydroclimate, subsurface flow, and biogeochemical reactions, this work provides a predictive framework for understanding how wetland systems respond to environmental change, with direct implications for water quality and carbon cycling.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2023

Engineered Escherichia coli platforms for tyrosine-derivative production from phenylalanine using phenylalanine hydroxylase and tetrahydrobiopterin-regeneration system

Aromatic compounds derived from tyrosine are important and diverse chemicals that have industrial and commercial applications. Although these aromatic compounds can be obtained by extraction from natural producers, their growth is slow, and their content is low. To overcome these problems, many of them have been chemically synthesized from petroleum-based feedstocks. However, because of the environmental burden and depleting availability of feedstock, microbial cell factories are attracting much attention as sustainable and environmentally friendly processes. To facilitate development of microbial cell factories for producing tyrosine derivatives, we developed simple and convenient tyrosine-producing Escherichia coli platforms with a bacterial phenylalanine hydroxylase, which converted phenylalanine to tyrosine with tetrahydromonapterin as a cofactor, using a synthetic biology approach. By introducing a tetrahydrobiopterin-regeneration system, the tyrosine titer of the plasmid-based engineered strain was 4.63 g/L in a medium supplemented with 5.00 g/L phenylalanine with a test tube. The strains were successfully used to produce industrially attractive compounds, such as tyrosol with a yield of 1.58 g/L by installing a tyrosol-producing module consisting of genes encoding tyrosine decarboxylase and tyramine oxidase on a plasmid. Gene integration into E. coli chromosomes has an advantage over the use of plasmids because it increases genetic stability without antibiotic feeding to the culture media and enables more flexible pathway engineering by accepting more plasmids with artificial pathway genes. Therefore, we constructed a plasmid-free tyrosine-producing platform by integrating five modules, comprising genes encoding the phenylalanine hydroxylase and tetrahydrobiopterin-regeneration system, into the chromosome. The platform strain could produce 1.04 g/L of 3,4-dihydroxyphenylalanine, a drug medicine, by installing a gene encoding tyrosine hydroxylase and the tetrahydrobiopterin-regeneration system on a plasmid. Moreover, by installing the tyrosol-producing module, tyrosol was produced with a yield of 1.28 g/L. We developed novel E. coli platforms for producing tyrosine from phenylalanine at multi-gram-per-liter levels in test-tube cultivation. The platforms allowed development and evaluation of microbial cell factories installing various designed tyrosine-derivative biosynthetic pathways at multi-grams-per-liter levels in test tubes.

09 BIOMASS FUELS↗

NASA NTRS2007

Effect Of Spaceflight On Microbial Gene Expression And Virulence: Preliminary Results From Microbe Payload Flown On-Board STS-115

Human presence in space, whether permanent or temporary, is accompanied by the presence of microbes. However, the extent of microbial changes in response to spaceflight conditions and the corresponding changes to infectious disease risk is unclear. Previous studies have indicated that spaceflight weakens the immune system in humans and animals. In addition, preflight and in-flight monitoring of the International Space Station (ISS) and other spacecraft indicates the presence of opportunistic pathogens and the potential of obligate pathogens. Altered antibiotic resistance of microbes in flight has also been shown. As astronauts and cosmonauts live for longer periods in a closed environment, especially one using recycled water and air, there is an increased risk to crewmembers of infectious disease events occurring in-flight. Therefore, understanding how the space environment affects microorganisms and their disease potential is critically important for spaceflight missions and requires further study. The goal of this flight experiment, operationally called MICROBE, is to utilize three model microbial pathogens, Salmonella typhimurium, Pseudomonas aeruginosa, and Candida albicans to examine the global effects of spaceflight on microbial gene expression and virulence attributes. Specifically, the aims are (1) to perform microarray-mediated gene expression profiling of S. typhimurium, P. aeruginosa, and C. albicans, in response to spaceflight in comparison to ground controls and (2) to determine the effect of spaceflight on the virulence potential of these microorganisms immediately following their return from spaceflight using murine models. The model microorganisms were selected as they have been isolated from preflight or in-flight monitoring, represent different degrees of pathogenic behavior, are well characterized, and have sequenced genomes with available microarrays. In particular, extensive studies of S. typhimurium by the Principal Investigator, Dr. Nickerson, using ground-based analog systems demonstrate important changes in the genotypic, phenotypic, and virulence characteristics of this pathogen resulting from exposure to a flight-like environment (i.e. modeled microgravity).

Wilson, J. W.↗

DOE OSTI2023

Microbial metabolic dependency and its impacts on the soil carbon cycle (Final Technical Report, DE-SC0016364)

The terrestrial biosphere contains a large fraction of global carbon (C) and nearly 70% of the organic C in terrestrial systems is found in soils. Global changes in atmospheric CO 2 , temperature, precipitation, and ecosystem N inputs, will impact primary production and carbon inputs to soils, but it remains difficult to predict soil responses to these environmental conditions. Our ability to predict C-cycle responses to global change remains limited by unexplained variability within the terrestrial C-cycle. Our difficulty in predicting responses of the soil C-cycle to environmental change suggests a need for a greater understanding of the biotic mechanisms that govern soil C-transformations. Changes in microbial community structure and function have been proposed to impact soil processes, but mechanistic connections between microbial activity and C-cycle transformations remain poorly described. We assembled an interdisciplinary team to conduct cross-scale examination of microbial contributions to C-cycle dynamics and C-fate in soils. We investigated microbial contributions to soil C-transformations using a suite of complementary isotopic techniques including DNA-Stable Isotope Probing (DNA-SIP), metagenomic-SIP, metabolomic-SIP, proteomic-SIP, and microspatial analysis of soil organic C. These techniques were deployed in experiments that tested fundamental assumptions that underlie terrestrial C-cycle models. We characterized the functional characteristics of soil microorganisms responsible for major C-transformations in soil, evaluated the metabolic and ecological interactions that underlie soil C-cycle dynamics, and evaluated the degree to which these interactions impact rates of C mineralization and stabilization in soils.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2025

A small number of point mutations confer formate tolerance in Shewanella oneidensis

ABSTRACT Microbial electrosynthesis (MES) is a sustainable approach to chemical production from CO 2 and clean electricity. However, limitations in electron transfer efficiency and gaps in understanding of electron transfer pathways in MES systems prevent full realization of this technology. Shewanella oneidensis could serve as an MES biocatalyst because it has a well-studied, efficient transmembrane electron transfer pathway. A key first step in MES in this organism could be CO 2 reduction to formate. However, we report that wild-type S. oneidensis does not tolerate high levels of formate. In this work, we created and characterized formate-tolerant strains of S. oneidensis for further engineering and future use in MES systems through adaptive laboratory evolution. Two different point mutations in a gene encoding a predicted sodium-dependent bicarbonate transporter and a DUF2721-containing protein separately confer formate tolerance to S. oneidensis . The mutations were further evaluated to understand their role in improving formate tolerance. We also show that the wild-type and mutant versions of the putative sodium-dependent bicarbonate transporter improve formate tolerance of Zymomonas mobilis , indicating the potential of transferring this formate tolerance phenotype to other organisms. IMPORTANCE Shewanella oneidensis is a bacterium with a well-studied, efficient extracellular electron transfer pathway. This capability could make this organism a suitable host for microbial electrosynthesis using CO 2 or formate as feedstocks. However, we report here that formate is toxic to S. oneidensis , limiting the potential for its use in these systems. In this work, we evolve several strains of S. oneidensis that have improved formate tolerance, and we investigate some mutations that confer this phenotype. The phenotype is confirmed to be attributed to several single point mutations by transferring the wild-type and mutant versions of each gene to the wild-type strain. Finally, the formate tolerance mechanism of one variant is studied using structural modeling and expression in another host. This study, therefore, presents a simple method for conferring formate tolerance to bacterial hosts.

Cross, Megan C. Gruenberg (ORCID:0000000291589900)↗

DOE OSTI2020

Analogous wheat root rhizosphere microbial successions in field and greenhouse trials in the presence of biocontrol agents Paenibacillus peoriae SP9 and Streptomyces fulvissimus FU14

Abstract Two Pythium ‐infested soils were used to compare the wheat root and rhizosphere soil microbial communities from plants grown in the field or in greenhouse trials and their stability in the presence of biocontrol agents. Bacteria showed the highest diversity at early stages of wheat growth in both field and greenhouse trials, while fungal diversity increased later on, at 12 weeks of the crop cycle. The microbial communities were stable in roots and rhizosphere samples across both soil types used in this study. Such stability was also observed irrespective of the cultivation system (field or greenhouse) or addition of biocontrol coatings to wheat seeds to control Pythium disease (in this study soil infected with Pythium sp. clade F was tested). In greenhouse plant roots, Archaeorhizomyces , Debaryomyces , Delftia , and unclassified Pseudeurotiaceae were significantly reduced when compared to plant roots obtained from the field trials. Some operational taxonomic units (OTUs) represented genetic determinants clearly transmitted vertically by seed endophytes (specific OTUs were found in plant roots) and the plant microbiota was enriched over time by OTUs from the rhizosphere soil. This study provided key information regarding the microbial communities associated with wheat roots and rhizosphere soils at different stages of plant growth and the role that Paenibacillus and Streptomyces strains play as biocontrol agents in supporting plant growth in infested soils.

Araujo, Ricardo↗

NASA NTRS2016

EVA-Compatible Microbial Swab Tool

When we send humans to search for life on Mars, we'll need to know what we brought with us versus what may already be there. To ensure our crewed spacecraft meet planetary protection requirements—and to protect our science from human contamination—we'll need to know whether micro-organisms are leaking/venting from our ships and spacesuits. This is easily done by swabbing external vents and suit surfaces for analysis, but requires a specialized tool for the job. Engineers at the National Aeronautics and Space Administration (NASA) recently developed an Extravehicular Activity (EVA)-compatible swab tool that can be used to sample current space suits and life support systems. Data collected now will influence Mars life support and EVA hardware early in the planning process, before design changes become difficult and expensive.NASA’s EVA swab tool pairs a Space Shuttle-era tool handle with a commercially available swab tip mounted into a custom-designed end effector. A glove-compatible release mechanism allows the handle to quickly switch between swab tips, much like a shaving razor handle can snap onto a disposable blade cartridge. Swab tips are stowed inside individual sterile containers, each fitted with a microbial filter that allows the container to equalize atmospheric pressure, but prevents cabin contaminants from rushing into the container when passing from the EVA environment into a pressurized cabin. A bank of containers arrayed inside a tool caddy allows up to six individual samples to be collected during a given spacewalk.NASA plans to use the tool in 2016 to collect samples from various spacesuits during ground testing to determine what (if any) human-borne microbial contamination leaks from the suit under simulated thermal vacuum conditions. Next, the tool will be used on board the International Space Station to assess the types of microbial contaminants found on external environmental control and life support system vents. Data will support advanced EVA and life support system maturation studies, helping to answer questions such as “how close can an EVA-suited crew member approach an area of scientific interest without compromising the science?”

Rucker, Michelle A.↗

NASA NTRS2008

Latent Herpes Viruses Reactivation in Astronauts

Space flight has many adverse effects on human physiology. Changes in multiple systems, including the cardiovascular, musculoskeletal, neurovestibular, endocrine, and immune systems have occurred (12, 32, 38, 39). Alterations in drug pharmacokinetics and pharmacodynamics (12), nutritional needs (31), renal stone formation (40), and microbial flora (2) have also been reported. Evidence suggests that the magnitude of some changes may increase with time in space. A variety of changes in immunity have been reported during both short (.16 days) and long (>30 days) space missions. However, it is difficult to determine the medical significance of these immunological changes in astronauts. Astronauts are in excellent health and in superb physical condition. Illnesses in astronauts during space flight are not common, are generally mild, and rarely affect mission objectives. In an attempt to clarify this issue, we identified the latent herpes viruses as medically important indicators of the effects of space flight on immunity. This chapter demonstrates that space flight leads to asymptomatic reactivation of latent herpes viruses, and proposes that this results from marked changes in neuroendocrine function and immunity caused by the inherent stressfullness of human space flight. Astronauts experience uniquely stressful environments during space flight. Potential stressors include confinement in an unfamiliar, crowded environment, isolation, separation from family, anxiety, fear, sleep deprivation, psychosocial issues, physical exertion, noise, variable acceleration forces, increased radiation, and others. Many of these are intermittent and variable in duration and intensity, but variable gravity forces (including transitions from launch acceleration to microgravity and from microgravity to planetary gravity) and variable radiation levels are part of each mission and contribute to a stressful environment that cannot be duplicated on Earth. Radiation outside the Earth's magnetosphere is particularly worrisome because it includes ionizing radiation from cosmic galactic radiation. Increased stress levels appear even before flight, presumably from the rigors of preflight training and the anticipation of the mission (12, 32, 38, 39). Space flight causes significant changes in human immune function (32), but the means by which these changes come about have been difficult to discern. Consistent indicators of stress associated with space flight include increased production of stress hormones, and changes in cells of the immune system. These changes include elevated white blood cell (WBC) and neutrophil counts at landing (15, 16, 35, 37). Activation of generalized stress responses before, during, and after space flight probably affects the function of the immune system. Space flight has been shown to decrease many aspects of immune function, including natural killer (NK) cell activity, interferon production, the blastogenic response of leukocytes to mitogens, cell-mediated immunity, neutrophil function and monocyte function (5, 16, 18, 21, 35-37).

Mehta, Satish K.↗

DOE OSTI2023

Metabolically diverse microorganisms mediate methylmercury formation under nitrate-reducing conditions in a dynamic hydroelectric reservoir

Brownlee Reservoir is a mercury (Hg)-impaired hydroelectric reservoir that exhibits dynamic hydrological and geochemical conditions and is located within the Hells Canyon Complex in Idaho, USA. Methylmercury (MeHg) contamination in fish is a concern in the reservoir. While MeHg production has historically been attributed to sulfate-reducing bacteria and methanogenic archaea, microorganisms carrying the hgcA gene are taxonomically and metabolically diverse and the major biogeochemical cycles driving mercury (Hg) methylation are not well understood. In this study, Hg speciation and redox-active compounds were measured throughout Brownlee Reservoir across the stratified period in four consecutive years (2016–2019) to identify the location where and redox conditions under which MeHg is produced. Metagenomic sequencing was performed on a subset of samples to characterize the microbial community with hgcA and identify possible links between biogeochemical cycles and MeHg production. Biogeochemical profiles suggested in situ water column Hg methylation was the major source of MeHg. These profiles, combined with genome-resolved metagenomics focused on hgcA-carrying microbes, indicated that MeHg production occurs in this system under nitrate- or manganese-reducing conditions, which were previously thought to preclude Hg-methylation. Using this multidisciplinary approach, we identified the cascading effects of interannual variability in hydrology on the redox status, microbial metabolic strategies, abundance and metabolic diversity of Hg methylators, and ultimately MeHg concentrations throughout the reservoir. This work expands the known conditions conducive to producing MeHg and suggests that the Hg-methylation mitigation efforts by nitrate or manganese amendment may be unsuccessful in some locations.

54 ENVIRONMENTAL SCIENCES↗