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While traveling to deep space is difficult for many reasons, food is a crucial one. Round-trip Mars mission scenarios last 3 years, demanding food with a shelf-life of 5 years; this means that feeding human crew sustainably for long-duration missions beyond low Earth orbit (LEO) will ultimately lead to a paradigm shift away from the current Earth-based food production system, which depends upon storing and transporting prepackaged foods, and toward bio-regenerative production of food in space. Pharmaceuticals and nutritional supplements face similar challenges. Moreover, the methods we currently use to detect dangerous microbes in food require sample return to Earth, a situation not viable for deep-space missions. While the science behind generating foods and bioproducts is covered by other white papers, in this paper we discuss a crucial gap uniting all of them: how to ensure that such products are free of unwanted microbial contamination and safe for crew to consume. Because Earth-based food safety systems cannot be directly applied in space, safety assurance is currently a critical bottleneck in the space production of food and other bioproducts. Future sustainable deep-space missions will require NASA to devote more resources in the coming decade to understanding the biological and physical science principles underlying microbial food safety in space, and to developing efficient, reliable methods in this area.

The Space Crop Production (SCP) at the NASA Kennedy Space Center (KSC) is aimed at achieving nutrient supplementation and moving towards caloric independence from Earth by growing crops for astronauts. However, growing crops outside of Earth’s orbit creates many new challenges. The KCS SCP Project Scientists have developed a method of organizing these challenges, called the Gaps List. This Gaps List is a dynamic, taxonomically arranged list of missing knowledge and technologies needed to reach production goals. The purpose of this evaluation was to assess the effectiveness and current research coverage of the Gaps List by using it to document the KSC SCP Program research. In evaluating the research, it was determined that several needs within the SCP Program that are not being met, and many deficiencies within the Gaps List were encountered. Gaps involving hardware were largely unrecognized by the KSC researchers and the funding sources that would typically support them. The KSC SCP Project will require more engineering and physical sciences support to close those gaps. The Gaps List was found to be out of date and in need of rearrangement. A recommendation from this review was that the Gaps List be reassessed at regular intervals to ensure all gaps and relevant research topics are included. In order to better understand the gaps within the list, any assessment needs to include a measure of priority, dependency and breadth. These measures may be included within or along side the current taxonomical arrangement of the Gaps List.

Space crop production systems are being developed to grow fresh produce in-situ to supplement the astronauts’ diet, but the required ventilation rates for crops in different gravity environments remains poorly understood. The reduction or lack of buoyancy-driven convection in reduced gravity environments leads to impaired gas exchange (CO2 absorption, water transpiration and O2 release) at the leaf surface if no extra ventilation is provided, and this could lead to a reduction in biomass production in the long run. To better characterize the influence of different airspeeds on photosynthesis and be able to model this in low gravity, a chamber was designed to interface with a LI-6800 portable photosynthesis system. This paper details the design of this chamber, specifically made to measure whole-plant and small canopy gas exchange at different airspeeds. The fans provide turbulent mixing in the chamber to ensure that it behaves like a continuous stirred tank reactor (CSTR)and that the residence time distribution (RTD) is the same for any fan speed; the computational fluid dynamic (CFD) model of the gas domain (the air in the chamber) hence uses a k-omega turbulence model. An airflow map of the chamber was created using anemometer measurements for the different airspeeds tested, and this was used together with the CFD simulation results to relate the experimentally measured fan outputs to actual airspeeds on top of an artificial plant. Environmental parameters (air temperature, relative humidity, CO2level) are controlled by the LI-6800. This work was funded by NASA Space Biology through the NASA postdoctoral program / USRA.

Growing fresh, nutritious, palatable produce for crew consumption during spaceflight may provide health-promoting, bioavailable nutrients and enhance the astronaut dietary experience as we move toward longer-duration missions. Tending plants may also serve as a countermeasure for crew psychological stresses associated with spaceflight. However, requirements to support consistent growth of a variety of high quality, nutritious crops under spaceflight environmental conditions remain unclear. This study explores the potential to grow crops for consumption on the International Space Station (ISS) using the Veggie vegetable-production system. VEG-04A and B were two flight tests conducted in 2019 with the leafy green crop mizuna mustard. Mizuna was grown in two Veggie chambers simultaneously, with the chambers set to different red-to-blue light formulations; one Veggie was programmed as "red-rich" and the second as "blue-rich." Light quality is known to impact plant growth, nutrition, microbiology, and organoleptic characteristics on Earth, and the Veggie flight tests examined how these impacts might differ in microgravity. VEG-04A, a 35-day growth test with a single harvest, was initiated in June and harvested in July 2019. At harvest, the astronauts froze half of the edible plant tissue to return to Earth and weighed the remaining half using the Mass Measurement Device (MMD). Weighed samples were then cleaned with produce-sanitizing wipes, and consenting crew members participated in organoleptic evaluation of the fresh produce. The remaining sanitized produce was available for crew consumption as desired. Frozen flight samples were returned at the end of August for microbial and chemical analyses to assess food safety and nutritional quality. No pathogens were detected on VEG-04A flight or ground control samples. On average, bacterial and fungal counts were significantly lower on ground control samples than flight samples. VEG-04B, a 56-day test with multiple harvests from the same plants, assessed sustained productivity. VEG-04B was initiated in October 2019 with three harvests at four, six, and eight weeks after initiation. Challenges with the watering program occurred early during VEG-04A, and several plants failed to survive in both the flight and ground control operations. Thus, prior to VEG-04B, an extra test was conducted to tailor water timing and volumes. This test determined that mizuna grew best if the wicks inside the plant pillow were allowed to dry after plants germinated, reducing persistent water around the stem. The wicks changed from being a conduit for water out of the plant pillow to being a conduit for air into the root zone. This test allowed a fine tuning of methods for VEG-04B. It is our hope that these tests on ISS will help mitigate the risk of an inadequate food supply for long-duration missions by adding fresh vegetables to the crew diet. This research was co-funded by the Human Research Program and Space Biology (MTL#1075) in the ILSRA 2015 NRA call.

In-situ food production is a necessary step for human exploration of the solar system and requires a deep understanding of plant growth in reduced gravity environments. In particular, the lack of buoyancy-driven convection changes the gas exchange at the leaf surface, which decreases photosynthesis and transpiration rates, and ultimately biomass production. To understand the intricate relations between physical, chemical, and biochemical processes, the following methodology combines the development of a mechanistic model of plant growth in reduced gravity environments, computational fluid dynamics (CFD) simulations, and experiments in different time frames.The model presented here is a coupled mass and energy balance using the single round leaf assumption, including gravity as an entry parameter, and the leaf surface temperature as an output variable. Measures of the leaf surface temperature using infra-red cameras allow for a computation of the transpiration rate. This approach was followed to design a parabolic flight experiment, which performed 7 flights, and enabled data collection for model validation in different gravity and ventilation settings on a short time frame. Current measures of carbon assimilation and transpiration rate at the leaf and canopy level using an infra-red gas analyzer (Li-6800) in 1g lab conditions on several species will enable a validation on longer time frames and further calibration of the model. CFD studies both on the parabolic flight and on the lab experimental set-up allow the precise assessment of ventilation above the canopy and plants' leaves.Ultimately, this work will provide recommendations for the design of future plant growth hardware, especially on the lowest adequate ventilation for optimal plant growth in reduced gravity environments, as well as assessing biomass and oxygen production rates on planetary surfaces and space stations. This work was funded by CNES, CNRS, Clermont Auvergne Metropole, and NASA Space Biology through NASA postdoctoral program / USRA.

Space crop production will be important in future long duration exploration missions to supplement the packaged diet with fresh bioactive nutrients. Plant care and the addition of fresh veggies to the diet may also have a role in astronaut well-being. Pick-and-eat salad crops are the best candidates for this near-term supplementation since they require minimal processing or preparation to add to meals. While light quality can strongly influence plant responses on Earth, the impacts of light quality on plant growth and composition in spaceflight remain unclear. The VEG-04 experiment uses two Veggie plant growth chambers on the International Space Station to simultaneously test different red: blue light ratios on the growth of Mizuna mustard, a leafy green salad crop. In addition to plant health and yield, the composition of key nutrients is assessed. Astronauts conduct on-board organoleptic evaluation of the fresh produce. Microbial food safety of returned produce is examined, and a Hazard Analysis Critical Control Point (HACCP) plan has been developed for this crop. VEG-04 consists of two experiments, one lasting 28 days with a single harvest, and the second lasting 56 days, with three cut-and-come-again harvests. These different scenarios provide an opportunity to test two production concepts, examine different fertilizers, monitor microbial changes over time for this crop, and assess potential impacts of interacting with plants on crew behavioral health and performance in spaceflight operations. In ground testing, plant growth was not significantly different across the different light treatments, however nutrient composition did differ significantly. Flight test results will be compared with ground data. This research was co-funded by NASA's Human Research Program and Space Biology in the ILSRA 2015 NRA call.

The Veggie facility on ISS was used to demonstrate the suitability of the leafy greens, red romaine 'Outredgeous' lettuce, Waldman's green lettuce, and mizuna in a mixed crop configuration.

In a microgravity setting, such as the environment aboard the International Space Station (ISS), an ideal plant water delivery system is one that can grow edible crops with minimal resource consumption and minimal risk to crew members. There are also concerns associated with the ability to control fluid escape and biofilm formation resulting in potential dangers to systems, crops, or crewmembers. To identify an appropriate system, candidate systems were assembled and operated under simulated ISS environmental conditions (T,CO2,and RH) with red romaine lettuce (Lactuca sativa cultivar 'Outredgeous') as a model crop. Fluid reservoirs and randomly selected planting sites were sampled every seven days until maturity at which point edible plant biomass and root samples were also taken. Heterotrophic bacteria and fungi growth patterns throughout each planting cycle were determined by plate counts on appropriate agar media. The candidate systems were compared to a classic hydroponics system as a control and harvested crops were compared to controls as well as Veggie-grown and market produce. Plants harvested from candidate systems yielded lower average heterotrophic bacteria and fungi per gram of plant mass levels when compared to market and Veggie samples as well as those from the control system. Additional studies to evaluate the system sanitation regimen as well as testing additional crops should be considered to aid in the selection of an ideal system.

Plant associated microbiomes, the rhizosphere and phyllosphere, are composed of communities of bacteria and fungi that may be mutualistic or pathogenic. These communities have the potential to influence plant health and development and can affect plant growth. Crop plants are being investigated as a fresh and safe supplement to astronauts’ diet and it is critical to understand and characterize these microbial communities. Multi-species crops, Mizuna mustard (Brassica rapa var japonica), ‘Outredgeous’ red romaine lettuce (Lactuca sativa), and Waldman’s Green lettuce (Lactuca sativa) were grown in two Veggie units on the International Space Station (ISS) for three grow outs in various combinations of plant types. Upon harvest, plant and pillow samples were frozen and returned to Earth for analysis. Bacterial and fungal community analyses for plant leaf and root, as well as pillow components, wick and media, were completed using next generation sequencing with the goal of surveying the composition of the entire community and identifying any potential pathogens. Bacteria were identified using the 16S rRNA gene whereas, fungi were identified with the internal transcribed spacer (ITS). The community composition for these three crops was compared between crop types and between plant tissue types. It is vital to mission success for the short term and long term to add nutritious, safe to eat vegetables providing a supplement to the crew members’ dietary requirements as well as to develop planning for deep space missions as we reach for the moon and on to Mars. Veggie technology validation tests were supported by NASA’s Space Biology Program.

This presentation will be an introduction and overview of space crop production needs, goals, and challenges in the areas of robotics and automation for the workshop Aug. 6-7, 2019 at Kennedy Space Center. This presentation will be used to start the workshop and set the direction.

Cultivation of crops in controlled environmental agricultural systems may limit microbial colonization and reduce diversity of the microbial communities. Practices like seed and growth medium sanitization may further impact microbial communities in the mature plant and the plant’s capacity to limit the growth of pathogens through competition. As humans expand their travels to space, understanding plant growth, health, and development in closed environments will be critical to the success of producing a safe, supplemental food source for astronauts. To determine the persistence of a potential human pathogen in plant growth and development, sanitized and unsanitized seeds from, mizuna (Brassica rapa var japonica) and red romaine lettuce (Lactuca sativa cultivar ‘Outredgeous’), were inoculated with Escherichia coli, ATCC 21445, germinated under simulated International Space Station (ISS) environmental conditions and harvested every 7 days until maturity. The persistence of E. coli in the rhizosphere was determined by plating on selective media, real time PCR (Polymerase Chain Reaction) and community sequencing of the rhizosphere communities. E. coli was detected in the crops’ roots and leaves for several weeks post germination. At day 28, plants from sanitized seeds had significantly higher counts of E. coli on the roots than those from unsanitized seeds. E. coli was also detected on a few uninoculated plants indicating airborne cross contamination among plants in the same growth chamber and suggesting an influence of the natural microbiome on human pathogen survival and persistence in leafy greens. Sequencing analysis revealed variations in composition and diversity between the communities. Understanding the microbial community of the rhizospheric microbiome is only the first step in determining the relationships between plants. Additional studies to include genotypic and phenotypic variations in the plants should be considered to determine if the natural microbes in the rhizosphere may contribute to the health and therefore, safety of the edible plants.

New technologies will be needed as mankind moves towards exploration of cislunar space, the Moon and Mars. Although many advances in our understanding of the effects of spaceflight on plant growth have been achieved in the last 40 years, spaceflight plant growth systems have been primarily designed to support space biology studies. Recently, the need for a sustainable and robust food system for future missions beyond Low Earth Orbit (LEO) has identified gaps in current technologies for food production. The goal is to develop safe and sustainable food production systems with reduced resupply mass and crew time compared to current systems.

NASA's mission for manned long- duration space exploration drives the research for crop selection to provide a nutritious and safe supplement to an astronaut's diet. Understanding plant growth, health, and the associated microbial communities in closed environments will be critical to the success of this mission. Cultivation of crops in closed controlled environment agricultural systems may limit microbial colonization and reduce diversity of the microbial communities. Furthermore, practices like seed and growth medium sanitization may impact microbial communities in the mature plant and the capacity to limit the growth of food borne pathogens through competition.

This is our annual "station report" of activities related to controlled environment research to the North Central Education Research Activity (NCERA-101) committee. The committee is sponsored the USDA National Institute for Food and Agriculture (NIFA). Kennedy Space Center has participated in this committee for over 30 years.

Plant growth research for food production at Kennedy Space Center looks at how future residents of Mars and the Moon will enjoy the sight, smell, taste, and nutrition of plants. Overall, the goal is to provide a sustainable source of healthy food, on long-duration space flights, so astronauts can get the nutrition they need and produce food. The sustainable production of food will aid in the efforts of closed life support. Plants have a vital application for bio regenerative life support as demands for food and oxygen can be provided through photosynthesis, while the carbon dioxide from human respiration is removed. Transpiration is also used in life support processes as waste water that can be recycled through plant systems with the resultant humidity then condensed as clean water. Selected crops will provide the nutrient requirements needed for long duration space flight. Currently, projects in food production are investigating how plants grow in Martian regolith simulant, new crops testing with tomato and pepper cultivars, acquiring real-time photosynthetic data on crops, assessing plant growth in porous concrete media, and the use of plasma for surface decontamination of seeds.

Supplemental safe food production has been an essential goal of NASA to meet the nutritional needs of astronauts on the International Space Station (ISS) as well as for future long duration missions to the moon and beyond. Food crops grown in space experience different environmental conditions than plants grown on Earth (i.e. microgravity and spaceflight physical sciences impacts). To test the growth methods and effects of the space environment, red romaine lettuce Lactuca sativa cv. 'Outredgeous', was grown in Veggie plant growth chambers on the ISS. Microbiological food safety of the plants grown on the ISS was determined by heterotrophic plate counts to assess total microbial load for bacteria and fungi as well as screening for specific pathogens and isolate identification. Molecular characterization was completed using Next Generation Sequencing (NGS) to provide valuable information on the taxonomic composition and community structure of the plant microbiome. Chemical analyses of plant tissue were conducted to understand spaceflight-induced changes in key elements in the space diet, phenolics, anthocyanin levels, and Oxygen radical absorbance capacity (ORAC), a measure of antioxidant capacity. Three growth tests of red romaine lettuce were completed on ISS, VEG-01A, VEG-01B, and VEG-03A. Plants were harvested using two harvest methods, either a single terminal harvest (after 33 days) or cut-and-come-again repetitive harvesting (64 days total growth). Ground controls were grown simultaneously with a delay to accommodate condition monitoring and replication. A comparison of the plant tissue returned to Earth showed leaves from the second grow-out had significantly higher bacterial counts than the preceding or subsequent growth test or any of the ground controls. Fungal counts were significantly higher on the final cut-and-come-again harvest of the third grow out. None of the potential foodborne pathogens that were screened for were detected. Bacterial and fungal isolate identification and community characterization indicated similar diversity between VEG-01A and VEG-01B growth tests, however, there appeared to be subtle differences in diversity and distribution among the three growth tests. Chemical analysis of plant tissue revealed significant variation in a few elemental data, but variation in levels of phenolics, anthocyanins, and ORAC was not significantly different. This study indicated that leafy vegetable crops could safely provide an edible supplement to astronauts' diet, and our analysis provided baseline data for continual operation of the Veggie plant growth units on ISS. This research was funded by NASA's space biology program.