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All sourcesNASA NTRSDOE OSTI
NASA NTRS2004

Microbial structural diversity estimated by dilution-extinction of phenotypic traits and T-RFLP analysis along a land-use intensification gradient

The present work tested whether the relationship between functional traits and inoculum density reflected structural diversity in bacterial communities from a land-use intensification gradient applying a mathematical model. Terminal restriction fragment length polymorphism (T-RFLP) analysis was also performed to provide an independent assessment of species richness. Successive 10-fold dilutions of a soil suspension were inoculated onto Biolog GN(R) microplates. Soil bacterial density was determined by total cell and plate counts. The relationship between phenotypic traits and inoculum density fit the model, allowing the estimation of maximal phenotypic potential (Rmax) and inoculum density (KI) at which Rmax will be half-reduced. Though Rmax decreased with time elapsed since clearing of native vegetation, KI remained high in two of the disturbed sites. The genetic pool of bacterial community did not experience a significant reduction, but the active fraction responding in the Biolog assay was adversely affected, suggesting a reduction in the functional potential. c2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

NASA Center KSC↗
NASA NTRS2012

Genetic Inventory Task Final Report

Contaminant terrestrial microbiota could profoundly impact the scientific integrity of extraterrestrial life-detection experiments. It is therefore important to know what organisms persist on spacecraft surfaces so that their presence can be eliminated or discriminated from authentic extraterrestrial biosignatures. Although there is a growing understanding of the biodiversity associated with spacecraft and cleanroom surfaces, it remains challenging to assess the risk of these microbes confounding life-detection or sample-return experiments. A key challenge is to provide a comprehensive inventory of microbes present on spacecraft surfaces. To assess the phylogenetic breadth of microorganisms on spacecraft and associated surfaces, the Genetic Inventory team used three technologies: conventional cloning techniques, PhyloChip DNA microarrays, and 454 tag-encoded pyrosequencing, together with a methodology to systematically collect, process, and archive nucleic acids. These three analysis methods yielded considerably different results: Traditional approaches provided the least comprehensive assessment of microbial diversity, while PhyloChip and pyrosequencing illuminated more diverse microbial populations. The overall results stress the importance of selecting sample collection and processing approaches based on the desired target and required level of detection. The DNA archive generated in this study can be made available to future researchers as genetic-inventory-oriented technologies further mature.

microbial diversity↗
NASA NTRSDate not supplied

Bionutrients-1: Utilizing Genomics and Transcriptomics to Assess the Reliability of Microorganisms for In Situ Nutrient Production on Long Duration Missions

The resupply of current long-duration crewed missions to the ISS relies on ground-launched supplies. As NASA looks toward Mars, ground-based resupply will no longer be an option. Critical nutrients, including vitamin C, vitamin K, folate, and thiamin, degrade during long-term storage, and regular consumption of these nutrients is essential for astronaut health. Another challenge of current food systems is the difficulty of consuming sufficient calories when subsisting on the limited flavors of freeze-dried food, which can lead to weight loss. The inclusion of microorganism-based food systems could alleviate both concerns. For example, the fermentation of rehydrated milk into yogurt with microorganisms genetically incorporating genes to produce critical vitamins would allow for both in situ production of nutrients and a fresh food product with additional flavor profiles. In comparison to plant food production, microorganisms require less flight infrastructure. The BioNutrients-1 mission is demonstrating viability of microbial fermentation food production in microgravity and testing the reliability of this approach for long-duration missions lacking resupply. While the BioNutrients-1 mission includes the collection of multiple phenotypic measurements, this status update will focus on the processing of samples for genomics and transcriptomics analyses as well as the planned analysis pipelines. First, the BioNutrients-1 mission seeks to identify microorganisms capable of surviving long-duration storage at ambient temperatures while maintaining genetic fidelity. To achieve this, nine commonly employed microbial species were stored at ambient temperatures in Stasis Packs on the ISS for five years. The viability and mutation rates will be measured at multiple time points for both flown and ground control samples. From an omics perspective, the changes in the bulk rates of point mutations and genetic rearrangements across the Stasis Pack species during the five years of storage will be determined, providing valuable insights into the potential of these microorganisms for long-duration space missions. Second, the BioNutrients-1 mission is characterizing the impact of microgravity on fermentation. Two strains of the yeast Saccharomyces cerevisiae, each encoding antioxidants (β-carotene or zeaxanthin) were flown to ISS for storage and fermentation within simplified bioreactors (Production Packs). The impact of microgravity on the expression of the antioxidant production genes and general metabolic genes will be determined using RNA sequencing. Ultimately, the transcriptome data will be compared to phenotypic measurements, such as the antioxidant yield, end-state biomass, and the production of EtOH, to determine the impacts of microgravity and long-term storage on microbial fermentation. The findings from this research will be instrumental in understanding the challenges and opportunities of microorganism-based food systems in space missions.

BioNutrients↗
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↗
NASA NTRS2016

PowerCells Payload on EuCROPIS - Measuring Synthetic Biology in Space

NASA’s PowerCell payload on the DLR (Deutsches Zentrum fur Luft- und Raumfahrt, i.e. German Aerospace Center) Eu:CROPIS satellite will compare the effect of multiple simulated gravity regimes on basic processes required for synthetic biology in space including growth, protein production, and genetic transformation of the bacterium Bacillus subtilis. In addition, it will pioneer the use of a cyanobacterially-produced feedstock for microbial growth in space, a concept we call “PowerCell.” The PowerCell experiment system will be integrated using the Spaceflight Secondary Payload System with the German Space Agency's (DLR's) Euglena and Combined Regenerative Organic-food Production In Space (Eu:CROPIS) satellite, to be launched during the summer of 2017. In order to simulate the gravitational gradient of different celestial bodies, the Eu:CROPIS satellite will establish artificial microgravity, lunar, and Martian gravity levels prior to conducting each set of biological experiments, with experimental results compared to ground controls. Experiments will be carried out in microfluidics cards with experimental progress measured through absorbance as detected by the LED-based optical system. Here we describe the ground studies that led to these experiments, along with a description of the flight hardware and its performance. The results of this mission will provide foundational data for the use and production of genetically engineered organisms for extraterrestrial missions.

PowerCell experiment system↗
NASA NTRS2013

Elucidating Microbial Adaptation Dynamics via Autonomous Exposure and Sampling

The adaptation of micro-organisms to their environments is a complex process of interaction between the pressures of the environment and of competition. Reducing this multifactorial process to environmental exposure in the laboratory is a common tool for elucidating individual mechanisms of evolution, such as mutation rates. Although such studies inform fundamental questions about the way adaptation and even speciation occur, they are often limited by labor-intensive manual techniques. Current methods for controlled study of microbial adaptation limit the length of time, the depth of collected data, and the breadth of applied environmental conditions. Small idiosyncrasies in manual techniques can have large effects on outcomes; for example, there are significant variations in induced radiation resistances following similar repeated exposure protocols. We describe here a project under development to allow rapid cycling of multiple types of microbial environmental exposure. The system allows continuous autonomous monitoring and data collection of both single species and sampled communities, independently and concurrently providing multiple types of controlled environmental pressure (temperature, radiation, chemical presence or absence, and so on) to a microbial community in dynamic response to the ecosystem's current status. When combined with DNA sequencing and extraction, such a controlled environment can cast light on microbial functional development, population dynamics, inter- and intra-species competition, and microbe-environment interaction. The project's goal is to allow rapid, repeatable iteration of studies of both natural and artificial microbial adaptation. As an example, the same system can be used both to increase the pH of a wet soil aliquot over time while periodically sampling it for genetic activity analysis, or to repeatedly expose a culture of bacteria to the presence of a toxic metal, automatically adjusting the level of toxicity based on the number or growth rate of surviving cells. We are on our second prototype iteration, with demonstrated functions of microbial growth monitoring and dynamic exposure to UV-C radiation and temperature. We plan to add functionality for general chemical presence or absence by Nov. 2013. By making the project low-cost and open-source, we hope to encourage others to use it as a basis for future development of a common microbial environmental adaptation testbed.

Microbiology↗
NASA NTRS2005

Effects of artificial defoliation of pines on the structure and physiology of the soil fungal community of a mixed pine-spruce forest

Loss of photosynthetic area can affect soil microbial communities by altering the availability of fixed carbon. We used denaturing gradient gel electrophoresis (DGGE) and Biolog filamentous-fungus plates to determine the effects of artificial defoliation of pines in a mixed pine-spruce forest on the composition of the fungal community in a forest soil. As measured by DGGE, two fungal species were affected significantly by the defoliation of pines (P < 0.001); the frequency of members of the ectomycorrhizal fungus genus Cenococcum decreased significantly, while the frequency of organisms of an unidentified soil fungus increased. The decrease in the amount of Cenococcum organisms may have occurred because of the formation of extensive hyphal networks by species of this genus, which require more of the carbon fixed by their host, or because this fungus is dependent upon quantitative differences in spruce root exudates. The defoliation of pines did not affect the overall composition of the soil fungal community or fungal-species richness (number of species per core). Biolog filamentous-fungus plate assays indicated a significant increase (P < 0.001) in the number of carbon substrates utilized by the soil fungi and the rate at which these substrates were used, which could indicate an increase in fungal-species richness. Thus, either small changes in the soil fungal community give rise to significant increases in physiological capabilities or PCR bias limits the reliability of the DGGE results. These data indicate that combined genetic and physiological assessments of the soil fungal community are needed to accurately assess the effect of disturbance on indigenous microbial systems.

Pinus/growth & development/physiology↗
NASA NTRSDate not supplied

Strategies for Determining Safety of Fermented Foods Produced in Space

The BioNutrients experiments examine the feasibility of using microbially produced foods – yogurt, kefir, fermented food products– to supplement the current prepackaged food system to provide freshly synthesized nutrients and genetically engineered protein therapeutics to support crew health for sustainable and long duration crewed exploration. Safety of the products must be established as direct consumption of such products is a risk due to potential contamination with pathogenic organisms. Identifying pathogen contamination is particularly challenging as the fermenting cultures can mask the presence of contaminating organisms. Here we consider current methods of detecting coliforms, aerobic colonies, non-lactic acid bacteria, molds (yeasts), and specific pathogens including Staphylococcus aureus and salmonella. We also consider alternate and additional testing including sequencing and quantitative PCR. We investigate the use of the E-Nose developed by NASA Ames Research Center. This portable device senses volatile organic compounds in real time and can be trained to recognize “good” uncontaminated products from contaminated ones as a means of predicting food safety. Finally, we consider methods of in-flight pasteurization to increase safety and decrease overall microbial load of these products. This presentation will summarize testing status and outline the strategy that will be employed in the BioNutrients-3 ISS fermented foods experiment.

Natalie Ball↗
NASA NTRSDate not supplied

Strategies for Determining Safety of Fermented Foods Produced in Space

The BioNutrients experiments examine the feasibility of using microbially produced foods – yogurt, kefir, fermented food products– to supplement the current prepackaged food system to provide freshly synthesized nutrients and genetically engineered protein therapeutics to support crew health for sustainable and long duration crewed exploration. Safety of the products must be established as direct consumption of such products is a risk due to potential contamination with pathogenic organisms. Identifying pathogen contamination is particularly challenging as the fermenting cultures can mask the presence of contaminating organisms. Here we consider current methods of detecting coliforms, aerobic colonies, non-lactic acid bacteria, molds (yeasts), and specific pathogens including Staphylococcus aureus and salmonella. We also consider alternate and additional testing including sequencing and quantitative PCR. We investigate the use of the E-Nose developed by NASA Ames Research Center. This portable device senses volatile organic compounds in real time and can be trained to recognize “good” uncontaminated products from contaminated ones as a means of predicting food safety. Finally, we consider methods of in-flight pasteurization to increase safety and decrease overall microbial load of these products. This presentation will summarize testing status and outline the strategy that will be employed in the BioNutrients-3 ISS fermented foods experiment.

Food↗
NASA NTRS2002

Solar Radiation as Driving Force In Early Evolution

Ultraviolet radiation (UVR) has provided an evolutionary challenge to life on Earth in that it is both an agent of mutation and as well as a selective force. Today surface fluxes of UVR vary diurnally, seasonally, etc. Still, the UVR flux was probably substantially higher during the early phases of evolution, suggesting that its role in evolution was even more prominent during this time. In this presentation, the creative role of UVR in evolution is discussed, specifically in connection with the role that UVR may have played in the evolution of early microbial ecosystems. The presentation will include discussions of the direct influence of UVR on such processes as photosynthesis and genetic damage, as well as the indirect influence of UVR as mediated through the production of reactive oxygen species. These biological effects of UVR will be viewed against the backdrop of the physical nature of the early Earth, surely a very different place then than now.

Rothschild, Lynn J.↗
NASA NTRS2010

Utilization of Alternate Chirality Enantiomers in Microbial Communities

Our previous study of chirality led to interesting findings for some anaerobic extremophiles: the ability to metabolize substrates with alternate chirality enantiomers of amino acids and sugars. We have subsequently found that not just separate microbial species or strains but entire microbial communities have this ability. The functional division within a microbial community on proteo- and sugarlytic links was also reflected in a microbial diet with L-sugars and D-amino acids. Several questions are addressed in this paper. Why and when was this feature developed in a microbial world? Was it a secondary de novo adaptation in a bacterial world? Or is this a piece of genetic information that has been left in modern genomes as an atavism? Is it limited exclusively to prokaryotes, or does this ability also occur in eukaryotes? In this article, we have used a broader approach to study this phenomenon using anaerobic extremophilic strains from our laboratory collection. A series of experiments were performed on physiologically different groups of extremophilic anaerobes (pure and enrichment cultures). The following characteristics were studied: 1) the ability to grow on alternate chirality enantiomers -- L-sugars and D- amino acids; 2) Growth-inhibitory effect of alternate chirality enantiomers; 3) Stickland reaction with alternate chirality amino acids. The results of this research are presented in this paper.

Pikuta, Elena V.↗
NASA NTRS2011

Marine Microbial Mats and the Search for Evidence of Life in Deep Time and Space

Cyanobacterial mats in extensive seawater evaporation ponds at Guerrero Negro, Baja California, Mexico, have been excellent subjects for microbial ecology research. The studies reviewed here have documented the steep and rapidly changing environmental gradients experienced by mat microorganisms and the very high rates of biogeochemical processes that they maintained. Recent genetic studies have revealed an enormous diversity of bacteria as well as the spatial distribution of Bacteria, Archaea and Eukarya. These findings, together with emerging insights into the intimate interactions between these diverse populations, have contributed substantially to our understanding of the origins, environmental impacts, and biosignatures of photosynthetic microbial mats. The biosignatures (preservable cells, sedimentary fabrics, organic compounds, minerals, stable isotope patterns, etc.) potentially can serve as indicators of past life on early Earth. They also can inform our search for evidence of any life on Mars. Mars exploration has revealed evidence of evaporite deposits and thermal spring deposits; similar deposits on Earth once hosted ancient microbial mat ecosystems.

Des Marais, David J.↗
NASA NTRS2017

Microbiological Horticultural Internship Final Abstract

GMO dwarf plum (Prunus domestica) is being evaluated as a candidate food crop for long duration space flight missions. A project was undertaken to develop a protocol for transferring selected genetic lines of GMO plum (previously maintained in pots and propagated by cuttings at NASA's Kennedy Space Center in Florida) into in vitro tissue culture. In vitro culture may reduce the space, materials, and labor required to maintain the current lines of GMO plum and better preserve them for future study. Fresh plant material from three selected GMO plum lines (NASA-5, NASA-10, and NASA-11) and a non-modified control line (Control-5) were processed aseptically into in vitro culture on four separate occasions. The impact of multiple treatments on the successful growth of GMO plum tissue in vitro were tested: Parent explant tissue type (leaf petioles, stem nodes containing buds and internodes without buds), tissue sterilization method [soaking in 10 bleach only (5 min for petioles or 10 min for nodesinternodes), or soaking in 70 EtOH (30 sec) followed by 10 bleach (5 min for petioles and 10 min for nodesinternodes)], and media type [three Murashige and Skoog-based medias (SGM, SRM, and SRM+2,4-D) and one recipe containing woody plant media (WPM)]. 22.2 of the plates containing tissue sterilized with bleach alone developed microbial contamination after two weeks, while only 11.8 of plates containing tissue sterilized sequentially with EtOH and bleach developed contamination. Node bud tissue from all four genetic lines of plum produced leafy plantlets on SGM and SRM media after 4-6 weeks. The most numerous and well-developed plantlets were present on SGM. Upon reaching suitable size, plantlets were transferred to larger media containers for further growth. Some node bud growth occurred on SRM+2,4-D and WPM 2.5 weeks after plating, however as of yet no pieces on SRM+2,4-D have adequate development for transferring. Tissue pieces from NASA-5 plated on WPM are developing leaves and will be ready for transferring soon. Petioles and internode tissue lacking bud meristem failed to produce any plantlets on any plates, however petioles developed large masses of undifferentiated callus tissue on SRM+2,4-D media. These callused pieces were then transferred to SRM+TDZ media, which resulted in even larger callus growth but no differentiation. All four selected plum lines were successfully transitioned into in vitro culture. Nodes from NASA-5 and NASA-10 lines produced the most numerous and well-developed leafy plantlets in vitro, while those from NASA-11 and Control-5 were generally smaller, slower growing and less numerous. The best method overall was to use young stem node tissue with buds, surface sterilize the pieces sequentially with 70 EtOH and 10 bleach, and then plate them onto SGM media. Future areas of study will include introducing additional genetic lines of GMO plum into in vitro culture, attempting to induce shoot growth in petiole callus tissue, testing methods (such as cold storage) that extend the time interval between transferring explants into new media, and testing viability of plantlets transferred from in vitro culture back to traditional pot culture.

bioregenerative life support↗
NASA NTRSDate not supplied

PowerCell aboard the EuCROPIS satellite - Results from the first synthetic biology experiment in space

Synthetic biology is the design and construction of new biological parts and systems not found in nature, a technology that we believe will be a game-changing technology for space exploration. The PowerCell concept is an ecology based on photosynthetic microbes taking advantage of in situ materials and energy to generate, on-demand, useful products (food, fuel, cloths, drugs, etc.) that satisfy specific needs of long-term human presence away from Earth. This first PowerCell experiment is testing this concept as well as several key mechanisms of engineering biology in a 48-well plate microfluidics system. NASA's PowerCell experiment was loaded in the fluidics system at NASA Ames in May 2016, and subsequently integrated on the DLR's compact satellite Eu:CROPIS (Euglena Combined Regenerative Organic food Production In Space), as a secondary payload. Launched into Earth’s orbit from the Vandenberg Air Force Base, California, USA, in December 3, 2019, the Eu:CROPIS platform has provided an artificial gravity through rotational acceleration, simulating gravity ranges compatible to those found on Mars and on the Moon, as well as microgravity in the absence of acceleration. Each set of biological experiments performed at Earth's orbit was compared to ground controls. Over 400 days' worth of mission data showed that all hardware and software worked as planned, and microbial growth was observed in all four gravity regimes, including growth on cyanobacterial cell extract in three out of four gravity regimes. The genetic transformation experiment was successful in microgravity and provided valuable long-term reagent stability data, including antibiotic efficacy. The demonstrated ability of hardy bacterial spores to recover from a long-term stasis period will be fundamental for deep human space missions. Lessons learned from hardware design to experimental operations are leading to improved designs and capabilities for future missions.

Ivan Glaucio Paulino Lima↗
NASA NTRS2018

Enabling Experimental Evolution: Multi-Parameter Sensor System Integration into a Culture/Stressor Biofluidics System

Experimental evolution (EE) exposes microbial communities to ecological stressors, simulating dynamics up to near-extinction events. Combined with comparative sequencing and other molecular tools, such data can inform the genetic and other biological mechanisms underlying extremophile adaptation, and other observed effects. Automating this type of experiment using biofluidics can mitigate many traditional obstacles, including delays in assay results and environment adjustment and the need for many replicates. A first-generation device for automating EE procedures, the Automated Adaptive Directed Evolution Chamber (AADEC), was developed at NASA Ames. UV-C radiation was the stressor, an LED-photodiode array measured optical density, magnetic agitation and peristaltic pump systems ensured nutrient availability, and Arduino microcontrollers provided control. Escherichia coli in LB kanamycin media was used for testing and performance verification. A manual laboratory procedure with timed exposure to UV-C was performed to typify tolerance acquisition. Approximately a 106 factor increase in survival ratio was recorded over multiple iterations. Currently, a second-generation device is being developed integrating more real-time sensors: redox potential (ORP), indicating available/consumed metabolic energy; dissolved oxygen (DO), indicating aerobic/anaerobic growth; pH, indicating metabolic products; and electrical conductivity (EC), another indicator of metabolic products. The EC sensor system was constructed and calibrated in-house and matched commercial sensors in the required range. A Raspberry Pi computer automated the electrical system, allowing real-time data acquisition. The fluidics card was made of CNC-milled polycarbonate for biocompatibility. Each sensor parameter can also be used as a selection pressure alone or in combination with others to create extreme microbial environments. As a proof of concept, this work demonstrated sensor operation in one pair of growth-sensor chambers. It can be expanded to a multi-chamber system to enable inter-culture comparisons and multi-population studies. The prior Arduino system will be ported to the RPi system. Future stressors to be added include thermal, reactive oxygen species, and varying nutrient availability.

Govinda Raj, Chinmayee↗
NASA NTRSDate not supplied

BioNutrients-3: Precision Fermentation, Pasteurization, and Pathogen Detection - Towards Safe Fermentation and Production of Nutrients in Space

The BioNutrients (BN) project is developing a microbial manufacturing approach to supplement the NASA food system to address known nutrient degradation associated with long-term storage. BN uses synthetic biology to deliver high-value nutrients and therapeutics through genetic engineering of microbes and production of fermented food products like yogurt and kefir, in a fully dehydrated system. On-demand production of nutrients for human consumption requires rigorous safety protocols to ensure contaminants are not introduced during the fermentation process. The third iteration of the BN flight project, BN-3, further develops the BN project by investigating strategies for pathogen detection and pasteurization in microgravity. BN-3 will test the limits of pathogen detection using whole genome sequencing, standard microbial assays and the NASA Ames E-Nose, a volatile carbon nano tube-based sensor array, to detect unwanted microbes. BN-3 also expands on the number of nutrients produced in a single bioreactor to enhance efficiency of the system by combining production of B vitamins in conjunction with the carotenoids, beta-carotene and zeaxanthin. Serial production of food products through yogurt passaging, new updates to the fluorinated ethylene propylene bags to allow crew access via a straw, as well as use of a food safe pH indicator dye to indicate readiness of the food product, will further enhance the useability of this system. This presentation will provide status of the BN-3 flight project with the aim of advancing in-space biomanufacturing for on-demand microbially based food production for future space exploration.

Biomanufacturing↗
NASA NTRS1997

Biological potential of extraterrestrial materials. 2. Microbial and plant responses to nutrients in the Murchison carbonaceous meteorite

Meteoritic materials are investigated as potential early planetary nutrients. Aqueous extracts of the Murchison C2 carbonaceous meteorite are utilized as a sole carbon source by microorganisms, as demonstrated by the genetically modified Pseudomonas fluorescence equipped with the lux gene. Nutrient effects are observed also with the soil microorganisms Nocardia asteroides and Arthrobacter pascens that reach populations up to 5 x 10(7) CFU/ml in meteorite extracts, similar to populations in terrestrial soil extracts. Plant tissue cultures of Asparagus officinalis and Solanum tuberosum (potato) exhibit enhanced pigmentation and some enhanced growth when meteorite extracts are added to partial nutrient media, but inhibited growth when added to full nutrient solution. The meteorite extracts lead to large increases in S, Ca, Mg, and Fe plant tissue contents as shown by X-ray fluorescence, while P, K, and Cl contents show mixed effects. In both microbiological and plant tissue experiments, the nutrient and inhibitory effects appear to be best balanced for growth at about 1:20 (extracted solid : H2O) ratios. The results suggest that solutions in cavities in meteorites can provide efficient concentrated biogenic and early nutrient environments, including high phosphate levels, which may be the limiting nutrient. The results also suggest that carbonaceous asteroid resources can sustain soil microbial activity and provide essential macronutrients for future space-based ecosystems.

Non-NASA Center↗