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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.

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.

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.

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.

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