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NASA NTRSDate not supplied

Plasma Sanitization of Radish Seeds for International Space Station Growth Experiments

Plant growth in space has long been a topic of key interest to enable sustained human presence. Seeds are common sources for microbial growth and must be sanitized before being sent to space. Current disinfection methods use toxic chemicals that require careful handling and dosing or are ineffective against certain seed types. Low temperature plasma provides an alternative to traditional methods. Plasma produces reactive species and ultraviolet light that reduce contaminants on seeds. NASA’s Kennedy Space Center has explored the treatment of radish seeds with plasma for sanitization and germination. Radio Frequency (RF) and Alternating Current (AC) plasma sources provided continuous treatment of radish seeds in a low-pressure and atmospheric environment, respectively. The seeds were analyzed to compare the presence of bacteria, fungi, and the germination rate. Comparisons between direct and indirect treatments, power supplies, treatment times, and treatment pressures showed the importance of proper dosing of reactive species. Higher powers and longer treatment times have shown larger reductions in microbial contaminants. However, extended direct exposure to high powered plasma can sometimes lead to a reduction in germination rates. Overall, plasma has shown promise as a viable alternative to chemical disinfection techniques. *Work supported by NASA's Kennedy Space Center Engineering and Safety Operations Independent Research and Technology Development.

K W Engeling↗

NASA NTRS2020

Bayesian Framework For Bioburden Density Calculations To Perform Planetary Protection Probabilistic Risk Assessment

The planetary protection discipline aims to minimize the microbial contamination on spacecraft to prevent the inadvertent contamination of other planetary bodies, known as forward planetary protection (PP). Planetary protection probabilistic risk assessment (PRA) relies on two core methodologies-the contamination probability event tree analysis and statistical parameter estimation. Planetary protection engineers combine several techniques to estimate the bioburden present on spacecraft components. A direct assay to enumerate CFU (colony forming units) is the preferred methodology, but given a similar processing environment the bioburden present on certain components is inferred using: (1) a NASA defined bioburden estimate based upon the biological cleanliness of the manufacturing/assembly environment or (2) sampled data from a similar spacecraft component. The paper presents an empirical Bayesian framework to systematically treat bioburden estimation and its uncertainties on different levels starting with measurement procedures to combining different components to subsystems and whole spacecraft. It is shown that the Bayesian approach can effectively handle estimations and their uncertainties at different levels and produce a reliable estimate for bioburden to be used to evaluate the probability of contamination.

Seuylemezian, Arman↗

NASA NTRSDate not supplied

Precision Fermentation: Relieving the Crabtree Effect in S. cerevisiae through Genetic Engineering

Nutrient degradation in the current NASA pre-packaged food system poses significant challenges to crew health in missions beyond low earth orbit as certain vitamins and nutrients have been shown to degrade during extended storage, and resupply and fresh food items cannot be readily provided. BioNutrients(BN) uses synthetic biology in an effort to supplement the NASA food system by delivering in situ production of nutrients and therapeutics produced by genetically engineered microbes. One yeast S. cerevisiae strain utilized in BN is genetically modified to produce two types of antioxidants: zeaxanthin and β-carotene. Zeaxanthin is a non-provitamin A carotenoid and has been implicated in eye health and vision performance, and β-carotene is a provitamin A carotenoid. Microbial production of high-value nutrients can aid our understanding and address the issue of nutrient degradation and loss in the NASA food system. Although S. cerevisiae is an excellent candidate for a long shelf life, on-demand nutrient production system, its space applications are limited by the Crabtree effect where respiration is shut down in favor of fermentative metabolism when glucose is present in high concentrations, causing the release of unwanted ethanol. Current and likely future life support systems are negatively impacted by ethanol and ethanol release is strictly limited.To improve feasibility of using S. cerevisiae, we explored ways to reduce the Crabtree effect, thereby limiting its ethanol production. Specifically, we used CRISPR-cas9 to genetically modify various genes in this pathway

E Zaroff↗

NASA NTRSDate not supplied

Exploring rock-regolith interfaces in Jezero crater with Mars 2020 SHERLOC

The Perseverance rover successfully landed in Jezero crater, Mars in February 2021 at the Octavia E. Butler landing site and began its mission to explore and sample an ancient crater lake basin. Principal goals of the Mars 2020 mission include characterizing the geology of Mars and seeking signs of ancient microbial life via the spacecraft cameras and spectroscopic instruments onboard. The Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals (SHERLOC) instrument is a deep UV Raman spectrometer that utilizes a 248.6nm pulsed laser. Part of SHERLOC is a color camera known as the Wide Angle Topographic Sensor for Operations and eNgineering (WATSON). The SHERLOC suite provides coordinated, spectroscopic and imaging capabilities at high spatial resolution, to detect minerals and organic molecules in microtextural context. By pairing high spatial resolution (~100 μm) resonance Raman and native fluorescence spectroscopy with microscopic imaging in a novel spacecraft capability, SHERLOC enables texture-specific molecular composition measurements of rock and regolith targets on Mars. Coordinated rock-regolith observations illuminate unique insights into weathering processes and thereby to primary properties of rocks in Jezero crater. This work describes the potential of rock-regolith interfaces to preserve unique records of geological processes in Jezero crater and can powerfully supplement observations of the more general rock record on Mars. Linking observations of local rock texture with associated regolith reveals important lithologic information based on the interrelationship between differential weathering behavior and mineralogy, grain size, and cement chemistry. Preliminary observations indicate that the polygonally fractured lithotype common near the Octavia E. Butler landing site may weather by granular disintegration and/or surface creep, a relation that can be uniquely observed at the rock-regolith interface. SHERLOC -specific observations of microtextural and elemental composition transitions presented here trace rock-regolith boundaries at multiple indurated surfaces adjacent to regolith. At these locales, grain-scale based examinations suggest chemical weathering could be related to a variably distributed coating or rind on dark rock targets that may be mafic in composition. Granule deposits overlying widely distributed fine-grained material are also observable. Mineral identifications of each phase are presented, with cross-scale comparisons to the remote insights gained by the SuperCam instrument.

Emily L. Cardarelli↗

NASA NTRS2012

Can Terrestrial Microbes Grow on Mars?

The theme for AbSciCon 2012 is "Exploring Life: Past and Present, Near and Far." The conference will address our current understanding of life - from processes at the molecular level to those which operate at planetary scales. Studying these aspects of life on Earth provides an essential platform from which to examine the potential for life on other worlds, both within our solar system and beyond. Mars exhibits a variety of extreme environments characterized by high UV and ionizing radiation flux, low pressure anoxic atmosphere, scarce or absent liquid water, extreme low temperatures, etc. The ability of terrestrial microorganisms to survive and adapt to the Mars environment has profound implications for astrobiology, planetary protection, and Mars life detection missions. At the NASA Ames Synthetic Biology Initiative, we believe that synthetic biology has the potential to revolutionize human space exploration. As such, the initiative is dedicated to applying the tools and techniques of synthetic biology to space exploration and astrobiology. Biological solutions will be invaluable for space exploration because they are not resource intensive, and they are versatile and self-renewing. An understanding of how to work with DNA in an unfavorable environment is paramount to utilizing biological tools on space missions. Furthermore, the ability to adjust life to the parameters of Mars is vital both to discovering what life on Mars might look like, and to using biological tools under such conditions. As a first step, we need an energy-efficient, low cost means of transporting, storing, and protecting genomic DNA, DNA parts, and whole microbial strains. Our goal is to develop and demonstrate viable and superior alternatives to standard DNA storage methods, which can be optimized to the conditions of space exploration, using synthetic biology as a tool. This includes protocols and kit designs for easy and repeatable DNA and strain recovery from protective storage conditions. We are constructing newly engineered genetic parts for different valuable host organisms, designed to increased long-term survival and functional retention. These methods should be applied for DNA and strain storage and transportation. In parallel, we seek inspiration from natural organisms that have developed means for survival in extreme environmental conditions. We are utilizing novel techniques for analysis of lipid biomarkers in the Antarctic Dry Valleys in order to identify resident microbes in the Antarctic soil and permafrost, as well as biomarker fossils of organisms that survived in the valleys in ages past. Through the identification of these life forms, we hope to understand and draw on new biological tools and strategies for synthetic biological applications on Mars.

Rothschild, Lynn↗

NASA NTRS2011

Microbiological Lessons Learned from the Space Shuttle

After 30 years of being the centerpiece of NASA s human spacecraft, the Space Shuttle will retire. This highly successful program provided many valuable lessons for the International Space Station (ISS) and future spacecraft. Major microbiological risks to crewmembers include food, water, air, surfaces, payloads, animals, other crewmembers, and ground support personnel. Adverse effects of microorganisms are varied and can jeopardize crew health and safety, spacecraft systems, and mission objectives. Engineering practices and operational procedures can minimize the negative effects of microorganisms. To minimize problems associated with microorganisms, appropriate steps must begin in the design phase of new spacecraft or space habitats. Spacecraft design must include requirements to control accumulation of water including humidity, leaks, and condensate on surfaces. Materials used in habitable volumes must not contribute to microbial growth. Use of appropriate materials and the implementation of robust housekeeping that utilizes periodic cleaning and disinfection will prevent high levels of microbial growth on surfaces. Air filtration can ensure low levels of bioaerosols and particulates in the breathing air. The use of physical and chemical steps to disinfect drinking water coupled with filtration can provide safe drinking water. Thorough preflight examination of flight crews, consumables, and the environment can greatly reduce pathogens in spacecraft. The advances in knowledge of living and working onboard the Space Shuttle formed the foundation for environmental microbiology requirements and operations for the International Space Station (ISS) and future spacecraft. Research conducted during the Space Shuttle Program resulted in an improved understanding of the effects of spaceflight on human physiology, microbial properties, and specifically the host-microbe interactions. Host-microbe interactions are substantially affected by spaceflight. Astronaut immune functions were found to be altered. Selected microorganisms were found to become more virulent during spaceflight. The increased knowledge gained on the Space Shuttle resulted in further studies of the host-microbe interactions on the ISS to determine if countermeasures were necessary. Lessons learned from the Space Shuttle Program were integrated into the ISS resulting in the safest space habitat to date.

Pierson, Duane L.↗

NASA NTRS2016

Establishing and Monitoring an Aseptic Workspace for Building the MOMA Mass Spectrometer

Mars Organic Molecule Analyzer (MOMA) is an instrument suite on the European Space Agency (ESA) ExoMars 2020 Rover, and the Mass Spectrometer (MOMA-MS) is being built at Goddard Space Flight Center (GSFC). MOMA-MS is a life-detection instrument and thus falls in the most stringent category of Planetary Protection (PP) biological cleanliness requirements. Less than 0.03 spore/m2 are allowed in the instrument sample path. In order to meet these PP requirements, MOMA-MS must be built and maintained in a low bioburden environment. The MOMA-MS project at GSFC maintains three clean rooms with varying levels of bioburden control. The Aseptic Assembly Clean room has the highest level of control, applying three different bioburden reducing methods: 70% Isopropyl Alcohol (IPA), 7.5% Hydrogen Peroxide, and Ultra-Violet C (UVC) light. The three methods are used in rotation and each kills microorganisms by a different mechanism, reducing the likelihood of microorganisms developing resistance to all three. The Integration and Mars Chamber Clean rooms use less biocidal cleaning, with the option to deploy extra techniques as necessary. To support the monitoring of clean rooms and verification that MOMA-MS hardware meets PP requirements, a new Planetary Protection lab was established that currently has the capabilities of standard growth assays for spore or vegetative bacteria, rapid bioburden analysis that detects Adenosine Triphosphate (ATP), plus autoclave and Dry Heat microbial Reduction (DHMR) verification. The clean rooms are monitored for vegetative microorganisms and by rapid ATP assay, and a clear difference in bioburden is observed between the aseptic and other clean room.

Contamination Control Engineering↗

NASA NTRS2015

Extravehicular Activity (EVA) 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 surfaces for analysis, but there was no US EVA tool for that job. NASA engineers developed an EVA-compatible swab tool that can be used to collect data on current hardware, which will influence eventual Mars life support and EVA hardware designs.

Rucker, Michelle↗

NASA NTRSDate not supplied

BioNutrients: Microbial on-demand production of short shelf-life micronutrients in space

Long duration deep space missions pose a significant challenge when considering adequate nutrition for human spaceflight. Many micronutrients supplied in pre-packaged foods for low Earth orbit flights have a short shelf life and will degrade over multi-year missions. One strategy to mitigate the risk of micronutrient malnutrition is to grow fresh foods to supplement a pre-packaged diet. The BioNutrients project aims to develop systems for growing edible microorganisms that are genetically engineered to produce short shelf life micronutrients. BioNutrients-1 (BN-1) is a five-year spaceflight project currently underway on the International Space Station (ISS) that tests long-term storage and production of two carotenoids, β-carotene and zeaxanthin, in genetically engineered yeast strains. BN-1 on-orbit and ground control activities indicate that the nutrient production system will be stable for at least two years in ambient storage conditions. BN-1 is also testing the effects of long-term ISS storage on viability and gene expression using “stasis packs” containing a series of microorganisms useful in food and biomanufacturing processes. Stasis pack analysis is distinguishing microorganisms with full viability after two years of storage from other species that lose viability rapidly. In parallel, BioNutrients-2 (BN-2) is being developed as a six-month flight experiment that will expand the range of foods produced on ISS to include yogurt, kefir, and probiotic yeast. BN-2 will improve upon hardware design by transitioning from a hard-shell polycarbonate bioreactor to a flexible fluorinated ethylene propylene (FEP) bag. This change reduces the mass and volume required to store production packs. The long-term goal of BioNutrients is to develop the capability for rapid and safe delivery of multiple micronutrients in single-use production packs. We expect synthetic biology approaches similar to BioNutrients will play an important role for astronaut health in future NASA missions to the Moon and Mars.

Synthetic Biology↗

NASA NTRSDate not supplied

3D Printed Materials Characterization for Rapid Prototyping and Plant Growth

Space Biology experimentation but their effects during plant growth and sanitation have not been documented. With further testing we can determine how certain 3D printed materials affect our plants. The way we tested germination included laying two pieces of germination paper down with the printed petri dish insert of the various material and 14 seeds of Outrageous lettuce to fit in the holes of the insert. We kept an even moisture in all parts by wrapping the petri dishes in two layers of parafilm tape. This project tests over 15 different materials to document performance during plant growth scenarios. This research will not only help space biology but also citizens who use these materials to understand the potential of leaching and ill effects. 3D printing is a recent generally accessed technology with new materials coming out every day. It is important to understand how materials could impact the way the plants grow, and the safety of the food grown. As we continue testing these materials, we will also test engineering properties like tension and elasticity. We have a microbiology team working on this project as well to test biofilm formation. From the germination tests we have completed we have seen little impact on germination rate of these seeds. However, some parts absorb moisture. An ideal material is one that doesn’t leach chemicals or microplastics, can withstand high tension and compression, and doesn’t promote microbial growth. This work is funded by NASA. M. Gandhi was funded by MAIANSE.

Mily Gandhi↗

NASA NTRS1990

Space Station Environmental Health System water quality monitoring

One of the unique aspects of the Space Station is that it will be a totally encapsulated environment and the air and water supplies will be reclaimed for reuse. The Environmental Health System, a subsystem of CHeCS (Crew Health Care System), must monitor the air and water on board the Space Station Freedom to verify that the quality is adequate for crew safety. Specifically, the Water Quality Subsystem will analyze the potable and hygiene water supplies regularly for organic, inorganic, particulate, and microbial contamination. The equipment selected to perform these analyses will be commercially available instruments which will be converted for use on board the Space Station Freedom. Therefore, the commercial hardware will be analyzed to identify the gravity dependent functions and modified to eliminate them. The selection, analysis, and conversion of the off-the-shelf equipment for monitoring the Space Station reclaimed water creates a challenging project for the Water Quality engineers and scientists.

Vincze, Johanna E.↗

NASA NTRS2019

Bionutrients-1, On-Demand Production of Nutrients in Space

Future long-duration missions face significant challenges maintaining crew health. A critical area is supplying adequate nutrition, as certain vitamins and nutrients in supplied foods and supplements demonstrate substantial degradation during extended storage. To address this issue, we are developing and flight-testing a platform technology that demonstrates in situ microbial production of targeted nutrients over extended mission durations. This 5-year experiment, known as BioNutrients-1, was started on the International Space Station in May 2019. It involves two components: an on-orbit hydration and production experiment; and the development of space-compatible, key bio-manufacturing microorganisms. On-orbit testing utilizes a small “production pack” system that encloses sterile edible growth substrate and desiccated Saccharomyces cerevisiae strains genetically engineered to produce the nutrients beta-carotene or zeaxanthin. On hydration and mixing of the production pack, the organisms revive and grow until limited by the depletion of growth media, hypothetically leading to consistent amounts of biomass and nutrients. In eventual mission applications, the packet contents would be heat treated to inactivate the microorganisms prior to consumption. For these flight experiments, the packet will not be heat treated, but will instead be frozen for return to Earth for analyses. In addition to the production pack trials, 14 different microorganisms/treatments were also delivered to ISS for long-duration storage. These samples will be intermittently returned to Earth and analyzed to determine survival rates and genomics. For this presentation, initial data from returned samples and ground controls will be discussed.

Hindupur, Aditya↗

NASA NTRS2019

BioNutrients-1: On-Demand Production of Nutrients in Space

Future long-duration missions face significant challenges maintaining crew health. A critical area is supplying adequate nutrition, as certain vitamins and nutrients in supplied foods and supplements demonstrate substantial degradation during extended storage. To address this issue, we are developing and flight-testing a platform technology that demonstrates in situ microbial production of targeted nutrients over extended mission durations. This 5-year experiment, known as BioNutrients-1, was started on the International Space Station in May 2019. It involves two components: an on-orbit hydration and production experiment; and the development of space-compatible, key bio-manufacturing microorganisms. On-orbit testing utilizes a small "production pack" system that encloses sterile edible growth substrate and desiccated Saccharomyces cerevisiae strains genetically engineered to produce the nutrients beta-carotene or zeaxanthin. On hydration and mixing of the production pack, the organisms revive and grow until limited by the depletion of growth media, hypothetically leading to consistent amounts of biomass and nutrients. In eventual mission applications, the packet contents would be heat treated to inactivate the microorganisms prior to consumption. For these flight experiments, the packet will not be heat treated, but will instead be frozen for return to Earth for analyses. In addition to the production pack trials, 14 different microorganisms/treatments were also delivered to ISS for long-duration storage. These samples will be intermittently returned to Earth and analyzed to determine survival rates and genomics. For this presentation, initial data from returned samples and ground controls will be discussed.

Hindupur, Aditya↗

NASA NTRS2020

EDL Simulation Results for the Mars 2020 Landing Site Safety Assessment

The Mars 2020 rover is NASA’s next flagship mission, set to explore Mars in search of scientific evidence of past microbial life. Importantly, the rover will also, for the first time, have the ability to collect and cache rock and soil samples for retrieval and return to laboratories here on Earth. A key step in the development of the Mars 2020 mission is the selection of a suitable landing site with the largest likelihood of meeting scientific goals. This decision is a complex and critical one that requires close interaction between the scientific and engineering communities. The chosen landing site must be both scientifically interesting — providing the project with the greatest possible chance of gathering credible and defendable scientific evidence — and also safe enough to attempt a landing in the first place. Thus, arguably one of the most important undertakings of the Entry, Descent, and Landing (EDL) team, is to effectively enumerate, quantify, and communicate the landing risks to all of the stakeholders. The culmination of this effort is the Landing Site Safety Assessment, which is a review commissioned by the project, presided over by the EDL Standing Review Board, and attended by management and science stakeholders, in which the EDL team communicates their assessment of the associated landing risks and the statistical probability of a successful landing at each of the final candidate landing sites. This paper summarizes the results of high-fidelity computer simulations of the Mars 2020 EDL sequence used in this assessment. From an EDL performance perspective, all four candidates offer similar level of robustness, which is in-family with Mars Science Laboratory (MSL). However, two new features of the Mars 2020 EDL sequence – range trigger and Terrain-Relative Navigation (TRN) – dramatically enhance the capability of the EDL system to safely land at landing sites with much more rugged terrain than ever before considered. This has allowed the landing site selection for Mars 2020 to proceed in a manner that has been unprecedentedly weighted more heavily toward scientific interest and less heavily on engineering constraints. With TRN, the overall probability of success is predicted to be approximately 99% for all of the candidates.

David Way↗

NASA NTRS1977

Engineering and simulation of life science Spacelab experiments

Approaches to the planning and realization of Spacelab life sciences experiments, which may involve as many as 16 Space Shuttle missions and 100 tests, are discussed. In particular, a Spacelab simulation program, designed to evaluate problems associated with the use of live animal specimens, the constraints imposed by zero gravity on equipment operation, training of investigators and data management, is described. The simulated facility approximates the hardware and support systems of a current European Space Agency Spacelab model. Preparations necessary for the experimental program, such as crew activity plans, payload documentation and inflight experimental procedures are developed; health problems of the crew, including human/animal microbial contamination, are also assessed.

Bush, B.↗

NASA NTRS2016

FOST 2 Upgrade with Hollow-Fiber CTA FO Module and Generation of Osmotic Agent for Microorganism Growth Studies

FOST 2 is an integrated membrane system that incorporates a forward osmosis subsystem and a reverse osmosis subsystem working in series. It has been designed as a post treatment system to process the effluent from the Membrane Aerated Biological Reactor developed at NASA Johnson Space Center and Texas Tech University. Its function is to remove dissolved solids residual such as ammonia and suspended solids, as well as to provide a physical barrier to microbial and viral contamination. A tubular CTA membrane module from HTI and a flat-sheet lipid-base membrane module from Porifera were integrated and tested on FOST 2 in the past, using both a bioreactor's effluent and greywater as the feed solution. This paper documents the performance of FOST 2 after its upgrade with a hollow-fiber CTA membrane module from Toyobo, treating real black-water to generate the osmotic agent solution necessary to conduct growth studies of genetically engineered microorganism for the Synthetic Biological Membrane project.

Water treatment↗

NASA NTRS1986

Design concepts for bioreactors in space

Microbial food sources are becoming viable and more efficient alternatives to conventional food sources especially in the context of Closed Ecological Life Support Systems (CELSS) in space habitats. Since bioreactor designs for terrestrial operation will not readily apply to conditions of microgravity, there is an urgent need to learn about the differences. These differences cannot be easily estimated due to the complex nature of the mass transport and mixing mechanisms in fermenters. Therefore, a systematic and expeditious experimental program must be undertaken to obtain the engineering data necessary to lay down the foundations of designing bioreactors for microgravity. Two bioreactor design concepts presented represent two dissimilar approaches to grappling with the absence of gravity in space habitats and deserve to be tested for adoption as important components of the life support function aboard spacecrafts, space stations and other extra-terrestrial habitats.

Seshan, P. K.↗

NASA NTRS2007

Living Together in Space: The International Space Station Internal Active Thermal Control System Issues and Solutions-Sustaining Engineering Activities at the Marshall Space Flight Center From 1998 to 2005

On board the International Space Station, heat generated by the crew and equipment is removed by the internal active thermal control system to maintain a comfortable working environment and prevent equipment overheating. Test facilities simulating the internal active thermal control system (IATCS) were constructed at the Marshall Space Flight Center as part of the sustaining engineering activities to address concerns related to operational issues, equipment capability, and reliability. A full-scale functional simulator of the Destiny lab module IATCS was constructed and activated prior to launch of Destiny in 2001. This facility simulates the flow and thermal characteristics of the flight system and has a similar control interface. A subscale simulator was built, and activated in 2000, with special attention to materials and proportions of wetted surfaces to address issues related to changes in fluid chemistry, material corrosion, and microbial activity. The flight issues that have arisen and the tests performed using the simulator facilities are discussed in detail. In addition, other test facilities at the MSFC have been used to perform specific tests related to IATCS issues. Future testing is discussed as well as potential modifications to the simulators to enhance their utility.

Wieland, P. O.↗