Crop yield and light/energy efficiency in a closed ecological system: Laboratory Biosphere experiments with wheat and sweet potato.

Two crop growth experiments in the soil-based closed ecological facility, Laboratory Biosphere, were conducted from 2003 to 2004 with candidate space life support crops. Apogee wheat (Utah State University variety) was grown, planted at two densities, 400 and 800 seeds m-2. The lighting regime for the wheat crop was 16 h of light-8 h dark at a total light intensity of around 840 micromoles m-2 s-1 and 48.4 mol m-2 d-1 over 84 days. Average biomass was 1395 g m-2, 16.0 g m-2 d-1 and average seed production was 689 g m-2 and 7.9 g m-2 d-1. The less densely planted side was more productive than the denser planting, with 1634 g m-2 and 18.8 g m-2 d-1 of biomass vs. 1156 g m-2 and 13.3 g m-2 d-1; and a seed harvest of 812.3 g m-2 and 9.3 g m-2 d-1 vs. 566.5 g m-2 and 6.5 g m-2 d-1. Harvest index was 0.49 for the wheat crop. The experiment with sweet potato used TU-82-155 a compact variety developed at Tuskegee University. Light during the sweet potato experiment, on a 18 h on/6 h dark cycle, totaled 5568 total moles of light per square meter in 126 days for the sweet potatoes, or an average of 44.2 mol m-2 d-1. Temperature regime was 28 +/- 3 degrees C day/22 +/- 4 degrees C night. Sweet potato tuber yield was 39.7 kg wet weight, or an average of 7.4 kg m-2, and 7.7 kg dry weight of tubers since dry weight was about 18.6% wet weight. Average per day production was 58.7 g m-2 d-1 wet weight and 11.3 g m-2 d-1. For the wheat, average light efficiency was 0.34 g biomass per mole, and 0.17 g seed per mole. The best area of wheat had an efficiency of light utilization of 0.51 g biomass per mole and 0.22 g seed per mole. For the sweet potato crop, light efficiency per tuber wet weight was 1.33 g mol-1 and 0.34 g dry weight of tuber per mole of light. The best area of tuber production had 1.77 g mol-1 wet weight and 0.34 g mol-1 of light dry weight. The Laboratory Biosphere experiment's light efficiency was somewhat higher than the USU field results but somewhat below greenhouse trials at comparable light levels, and the best portion of the crop at 0.22 g mol-1 was in-between those values. Sweet potato production was overall close to 50% higher than trials using hydroponic methods with TU-82-155 at NASA JSC. Compared to projected yields for the Mars on Earth life support system, these wheat yields were about 15% higher, and the sweet potato yields averaged over 80% higher.     

Carbon dioxide dynamics of combined crops of wheat,cowpea, pinto beans in the Laboratory Biosphere closed ecological system

A mixed crop consisting of cowpeas, pinto beans and Apogee ultra-dwarf wheat was grown in the Laboratory Biosphere, a 40 m³ closed life system equipped with 12,000 W of high pressure sodium lamps over planting beds with 5.37 m² of soil. Similar to earlier reported experiments, the concentration of carbon dioxide initially increased to 7860 ppm at 10 days after planting due to soil respiration plus CO₂ contributed from researchers breathing while in the chamber for brief periods before plant growth became substantial. Carbon dioxide concentrations then fell rapidly as plant growth increased up to 29 days after planting and subsequently was maintained mostly in the range of about 200–3000 ppm (with a few excursions) by CO₂ injections to feed plant growth. Numerous analyses of rate of change of CO₂ concentration at many different concentrations and at many different days after planting reveal a strong dependence of fixation rates on CO₂ concentration. In the middle period of growth (days 31–61), fixation rates doubled for CO₂ at 450 ppm compared to 270 ppm, doubled again at 1000 ppm and increased a further 50% at 2000 ppm. High productivity from these crops and the increase of fixation rates with elevated CO₂ concentration supports the concept that enhanced CO₂ can be a useful strategy for remote life support systems. The data suggests avenues of investigation to understand the response of plant communities to increasing CO₂ concentrations in the Earth’s atmosphere. Carbon balance accounting and evapotranspiration rates are included.     

Atmospheric dynamics and bioregenerative technologies in a soil-based ecological life support system: initial results from Biosphere 2.

Biosphere 2 is the first man-made, soil-based, bioregenerative life support system to be developed and tested. The utilization and amendment of local space resources, e.g. martian soil or lunar regolith, for agricultural and other purposes will be necessary if we are to minimize the requirement for Earth materials in the creation of long-term off-planet bases and habitations. Several of the roles soil plays in Biosphere 2 are 1) for air purification 2) as a key component in created wetland systems to recycle human and animal wastes and 3) as nutrient base for a sustainable agricultural cropping program. Initial results from the Biosphere 2 closure experiment are presented. These include the accelerated cycling rates due to small reservoir sizes, strong diurnal and seasonal fluxes in atmospheric CO2, an unexpected and continuing decline in atmospheric oxygen, overall maintenance of low levels of trace gases, recycling of waste waters through biological regeneration systems, and operation of an agriculture designed to provide diverse and nutritionally adequate diets for the crew members.     

Technical review of the Laboratory Biosphere closed ecological system facility.

Laboratory Biosphere is a 40 m3 closed life system that commenced operation in May 2002. Light is from 12,000 W of high pressure sodium lamps over planting beds with 5.37 m2 of soil. Water is 100% recycled by collecting condensate from the temperature and humidity control system and mixing with leachate collected from under the planting beds. Atmospheric leakage was estimated during the first closure experiment to be 0.5-1% per day in general plus about 1% for each usage of the airlock door. The first trial run of 94 days was with a soybean crop grown from seeds (May 17, 2002) to harvest (August 14, 2002) plus 5 days of post-harvest closure. The focus of this initial trial was system testing to confirm functionality and identify any necessary modifications or improvements. This paper describes the organizational and physical features of the Laboratory Biosphere.     

Excess nutrients in hydroponic solutions alter nutrient content of rice, wheat, and potato.

Environment has significant effects on the nutrient content of field-grown crop plants. Little is known, however, about compositional changes caused by controlled environments in which plants receive only artificial radiation and soilless, hydroponic culture. This knowledge is essential for developing a safe, nutritious diet in a Controlled Ecological Life-Support System (CELSS). Three crops that are candidates for inclusion in a CELSS (rice, wheat, and white potato) were grown both in the field and in controlled environments where the hydroponic nutrient solution, photosynthetic photon flux (PPF), and CO2 level were manipulated to achieve rapid growth rates. Plants were harvested at maturity, separated into discrete parts, and dried prior to analysis. Plant materials were analyzed for proximate composition (protein, fat, ash, and carbohydrate), total nitrogen (N), nitrate, minerals, and amino-acid composition. The effect of environment on nutrient content varied by crop and plant part. Total N and nonprotein N (NPN) contents of plant biomass generally increased under controlled-environment conditions compared to field conditions, especially for leafy plant parts and roots. Nitrate levels were increased in hydroponically-grown vegetative tissues, but nitrate was excluded from grains and tubers. Mineral content changes in plant tissue included increased phosphorus and decreased levels of certain micronutrient elements under controlled-environment conditions. These findings suggest that cultivar selection, genetic manipulation, and environmental control could be important to obtain highly nutritious biomass in a CELSS.     

Effect of bacterial population density on germination wheat seeds and dynamics of simple artificial ecosystems.

Effect of the size of rhizospheric bacterial populations on germination of seeds and development of simple terrestrial "wheat plants--rhizospheric microorganisms--artificial soil" and "wheat plants-artificial soil" systems has been studied. Experiments demonstrated that within specify ranges in the inoculate, the rhizospheric bacteria are capable of increasing the yield of germinated seeds and stimulate the growth of plantlets. Germination of seeds inoculated with bacteria was either stimulated, or inhibited or remained at control levels depending on the amount of bacteria. Plant biomass growth and total photoassimilation has been found to depend on the amount of bacteria on the plant roots: the higher the amount of bacteria on plant roots, the smaller is the biomass of plants but the total photoassimilation is, higher. Thus, depending on the amount of bacteria on the roots of plants the system either increases the biomass of plants or increases the total photoassimilation, i.e. "pumps" carbon through itself involving bacteria. Grant numbers: N99-04-96017, N15.     

The legacy of Biosphere 2 for the study of biospherics and closed ecological systems.

The unprecedented challenges of creating Biosphere 2, the world's first laboratory for biospherics, the study of global ecology and long-term closed ecological system dynamics, led to breakthrough developments in many fields, and a deeper understanding of the opportunities and difficulties of material closure. This paper will review accomplishments and challenges, citing some of the key research findings and publications that have resulted from the experiments in Biosphere 2. Engineering accomplishments included development of a technique for variable volume to deal with pressure differences between the facility and outside environment, developing methods of atmospheric leak detection and sealing, while achieving new standards of closure, with an annual atmospheric leakrate of less than 10%, or less than 300 ppm per day. This degree of closure permitted detailed tracking of carbon dioxide, oxygen, and trace gases such as nitrous oxide and ethylene over the seasonal variability of two years. Full closure also necessitated developing new approaches and technologies for complete air, water, and wastewater recycle and reuse within the facility. The development of a soil-based highly productive agricultural system was a first in closed ecological systems, and much was learned about managing a wide variety of crops using non-chemical means of pest and disease control. Closed ecological systems have different temporal biogeochemical cycling and ranges of atmospheric components because of their smaller reservoirs of air, water and soil, and higher concentration of biomass, and Biosphere 2 provided detailed examination and modeling of these accelerated cycles over a period of closure which measured in years. Medical research inside Biosphere 2 included the effects on humans of lowered oxygen: the discovery that human productivity can be maintained with good health with lowered atmospheric oxygen levels could lead to major economies on the design of space stations and planetary/lunar settlements. The improved health resulting from the calorie-restricted but nutrient dense Biosphere 2 diet was the first such scientifically controlled experiment with humans. The success of Biosphere 2 in creating a diversity of terrestrial and marine environments, from rainforest to coral reef, allowed detailed studies with comprehensive measurements such that the dynamics of these complex biomic systems are now better understood. The coral reef ecosystem, the largest artificial reef ever built, catalyzed methods of study now being applied to planetary coral reef systems. Restoration ecology advanced through the creation and study of the dynamics of adaptation and self-organization of the biomes in Biosphere 2. The international interest that Biosphere 2 generated has given new impetus to the public recognition of the sciences of biospheres (biospherics), biomes and closed ecological life systems. The facility, although no longer a materially-closed ecological system, is being used as an educational facility by Columbia University as an introduction to the study of the biosphere and complex system ecology and for carbon dioxide impacts utilizing the complex ecosystems created in Biosphere '.The many lessons learned from Biosphere 2 are being used by its key team of creators in their design and operation of a laboratory-sized closed ecological system, the Laboratory Biosphere, in operation as of March 2002, and for the design of a Mars on Earth(TM) prototype life support system for manned missions to Mars and Mars surface habitats. Biosphere 2 is an important foundation for future advances in biospherics and closed ecological system research.     

Methods for measurement and control of leakage in CELSS and their application and performance in the Biosphere 2 facility.

Atmospheric leakage between a CELSS and its surround is driven by the differential pressure between the two. In an earth-based CELSS, both negative and positive differential pressures of atmosphere are created as the resultant of three influences: thermal expansion/contraction, transition of water between liquid and vapor phases, and external barometric pressure variations. The resultant may typically be on the order of 5000 pascals. By providing a flexible expansion chamber, the differential pressure range can be reduced two, or even three, orders of magnitude, which correspondingly reduces the leakage. The expansion chamber itself can also be used to measure the leak rate. Independent confirmation is possible by measurement of the progressive dilution of a trace gas. These methods as employed at the Biosphere 2 facility have resulted in an estimated atmospheric leak rate of less than 10 percent per year.     

Determining the potential productivity of food crops in controlled environments.

The quest to determine the maximum potential productivity of food crops is greatly benefitted by crop growth models. Many models have been developed to analyze and predict crop growth in the field, but it is difficult to predict biological responses to stress conditions. Crop growth models for the optimal environments of a Controlled Environment Life Support System (CELSS) can be highly predictive. This paper discusses the application of a crop growth model to CELSS; the model is used to evaluate factors limiting growth. The model separately evaluates the following four physiological processes: absorption of PPF by photosynthetic tissue, carbon fixation (photosynthesis), carbon use (respiration), and carbon partitioning (harvest index). These constituent processes determine potentially achievable productivity. An analysis of each process suggests that low harvest index is the factor most limiting to yield. PPF absorption by plant canopies and respiration efficiency are also of major importance. Research concerning productivity in a CELSS should emphasize: 1) the development of gas exchange techniques to continuously monitor plant growth rates and 2) environmental techniques to reduce plant height in communities.     

Development and growth of potato tubers in microgravity.

A potato explant consisting of a leaf, its axillary bud, and a small segment of stem will develop a tuber in 10-14 days when grown on earth. The tubers develop from the axillary buds and accumulate starch derived from sugars produced through photosynthesis and/or mobilized from leaf tissue. Potato explants were harvested and maintained in the Astroculture (TM) unit, a plant growth chamber designed for spaceflight. The unit provides an environment with controlled temperature, humidity, CO2 level, light intensity, and a nutrient delivery system. The hardware was loaded onto the space shuttle Columbia 24 hours prior to the launch of the STS-73 mission. Explant leaf tissue appeared turgid and green for the first 11 days of flight, but then became chlorotic and eventually necrotic by the end of the mission. The same events occurred to ground control explants with approximately the same timing. At the end of the 16-day mission, tubers were present on each explant. The size and shape of the space-grown tubers were similar to the ground-control tubers. The arrangement of cells in the tuber interior and at the exterior in the periderm was similar in both environments. Starch and protein were present in the tubers grown in space and on the ground. The range in starch grain size was similar in tubers from both environments, but the distribution of grains into size classes differed somewhat, with the space-grown tubers having more small grains than the ground control tubers. Proteinaceous crystals were found in tubers formed in each condition.     

Food production and nutrition in Biosphere 2: results from the first mission September 1991 to September 1993.

The initial test of the Biosphere 2 agricultural system was to provide a nutritionally adequate diet for eight crew members during a two year closure experiment, 1991-1993. The overall results of that trial are presented in this paper. The 2000 m2 cropping area provided about 80 percent of overall nutritional needs during the two years. Adaptation of the crew to the diet which averaged 2200 calories, 73 g. of protein and 32 g. of fat per person over the course of the two years. The diet was primarily vegetarian, with only small amounts of milk, meat and eggs from the system's domestic animals. The crew experienced 10-20 percent weight loss, most of which occurred in the first six months of the closure reflecting adaptation to the diet and lower caloric intake during that period. Since Biosphere 2 is a tightly sealed system, non-toxic methods of pest and disease control were employed and inedible plant material, domestic animal wastes and human waste-water were processed and nutrients returned to the soil. Crop pests and diseases, especially broad mites and rootknot nematode, reduced yields, and forced the use of alternative crops. Outstanding crops included rice, sweet potato, beets, banana, and papaya. The African pygmy goats were the most productive of the domestic animals. Overall, the agriculture and food processing required some 45% of the crew time.     

Laboratory investigations of the survivability of bacteria in hypervelocity impacts.

It is now well established that material naturally moves around the Solar System, even from planetary surface to planetary surface. Accordingly, the idea that life is distributed throughout space and did not necessarily originate on the Earth but migrated here from elsewhere (Panspermia) is increasingly deemed worthy of consideration. If life arrived at the Earth from space, its relative speed will typically be of order many km s-1, and the resulting collision with the Earth and its atmosphere will be in the hypervelocity regime. A mechanism for the bacteria to survive such an impact is required. Therefore a programme of hypervelocity impacts in the laboratory at (4.5 +/- 0.6) km s-1 was carried out using bacteria (Rhodococcus) laden projectiles. After impacts on a variety of target materials (rock, glass and metal) attempts were made to culture Rhodococcus from the surface of the resulting craters and also from the target material ejected during crater formation. Control shots with clean projectiles yielded no evidence for Rhodococcus growth from any crater surface or ejecta. When projectiles doped with Rhodococcus were used no impact crater surface yielded colonies of Rhodococcus. However, for four shots of bacteria into rock (two on chalk and two on granite) the ejecta was afterwards found to give colonies of Rhodococcus. This was not true for shots onto glass. In addition, shots into aerogel (density 96 kg m-3) were also carried out (two with clean projectiles and two with projectiles with Rhodococcus). This crudely simulated aero-capture in a planetary atmosphere. No evidence for Rhodococcus growth was found from the projectiles captured in the aerogel from any of the four shots.     

Consideration in selecting crops for the human-rated life support system: a Linear Programming model.

A Linear Programming model has been constructed which aids in selecting appropriate crops for CELSS (Controlled Environment Life Support System) food production. A team of Controlled Environment Agriculture (CEA) faculty, staff, graduate students and invited experts representing more than a dozen disciplines, provided a wide range of expertise in developing the model and the crop production program. The model incorporates nutritional content and controlled-environment based production yields of carefully chosen crops into a framework where a crop mix can be constructed to suit the astronauts' needs. The crew's nutritional requirements can be adequately satisfied with only a few crops (assuming vitamin mineral supplements are provided) but this will not be satisfactory from a culinary standpoint. This model is flexible enough that taste and variety driven food choices can be built into the model.     

Induced abnormality in Mir- and Earth grown Super Dwarf wheat.

Super-dwarf wheat grown on the Mir space station using the Svet "Greenhouse" exhibited morphological, metabolic and reproductive abnormalities compared with Earth-grown wheat. Of prominent importance were the abnormalities associated with reproductive ontogeny and the total absence of seed formation on Mir. Changes in the apical meristem associated with transition from the vegetative phase to floral initiation and development of the reproductive spike were all typical of 'Super-Dwarf' wheat up to the point of anthesis. Observation of ruptured anthers from the Mir-grown plants revealed what appeared to be normally developed pollen. These pollen gains, however, contained only one nuclei, while normal viable pollen is tri-nucleate. A potentially important difference in the flight experiment, compared with ground reference studies, was the presence of a high level of atmospheric ethylene (1,200 ppb). Ground studies conducted by exposing 'Super-Dwarf' wheat to ethylene just prior to anthesis resulted in manifestation of the same abnormalities observed in the space flight samples.     

CO2 crop growth enhancement and toxicity in wheat and rice.

The effects of elevated CO2 on plant growth are reviewed and the implications for crop yields in regenerative systems are discussed. There is considerable theoretical and experimental evidence indicating that the beneficial effects of CO2 are saturated at about 0.12% CO2 in air. However, CO2 can easily rise above 1% of the total gas in a closed system, and we have thus studied continuous exposure to CO2 levels as high as 2%. Elevating CO2 from 340 to 1200 micromoles mol-1 can increase the seed yield of wheat and rice by 30 to 40%; unfortunately, further CO2 elevation to 2500 micromoles mol-1 (0.25%) has consistently reduced yield by 25% compared to plants grown at 1200 micromoles mol-1; fortunately, there was only an additional 10% decrease in yield as the CO2 level was further elevated to 2% (20,000 micromoles mol-1). Yield increases in both rice and wheat were primarily the result of increased number of heads per m2, with minor effects on seed number per head and seed size. Yield increases were greatest in the highest photosynthetic photon flux. We used photosynthetic gas exchange to analyze CO2 effects on radiation interception, canopy quantum yield, and canopy carbon use efficiency. We were surprised to find that radiation interception during early growth was not improved by elevated CO2. As expected, CO2 increased quantum yield, but there was also a small increase in carbon use efficiency. Super-optimal CO2 levels did not reduce vegetative growth, but decreased seed set and thus yield. The reduced seed set is not visually apparent until final yield is measured. The physiological mechanism underlying CO2 toxicity is not yet known, but elevated CO2 levels (0.1 to 1% CO2) increase ethylene synthesis in some plants and ethylene is a potent inhibitor of seed set in wheat.     

Yield comparisons and unique characteristics of the dwarf wheat cultivar 'USU-Apogee'.

Extremely short, high yielding cultivars of all crop plants are needed to optimize the food production of bioregenerative life support systems in space. In the early 1980's, we examined over a thousand wheat genotypes from the world germplasm collection in search of genotypes with appropriate characteristics for food production in space. Here we report the results of 12 years of hybridization and selection for the perfect wheat cultivar. 'USU-Apogee' is a full-dwarf hard red spring wheat (Triticum aestivum L.) cultivar developed for high yields in controlled environments. USU-Apogee was developed by the Utah Agricultural Experiment Station in cooperation with the National Aeronautics and Space Administration and released in April 1996. USU-Apogee is a shorter, higher yielding alternative to 'Yecora Rojo' and 'Veery-10', the short field genotypes previously selected for use in controlled environments. The yield advantage of USU-Apogee is 10 to 30% over these other cultivars, depending on environmental conditions. USU-Apogee (45-50 cm tall, depending on temperature) is 10 to 15 cm shorter than Yecora Rojo and 1 to 4 cm shorter than Veery-10. USU-Apogee was also selected for resistance to the calcium-induced leaf tip chlorosis that occurs in controlled-environments. Breeder seed of USU-Apogee will be maintained by the Crop Physiology Laboratory and seed is available for testing on request.     

High phosphorylase activity is correlated with increased potato minituber formation and starch content during extended clinorotation.

The major purpose of these experiments were to investigate growth of potato storage organs and starch synthesis in minitubers at slow horizontal clinorotation (2 rpm), which partly mimics microgravity, and a secondary goal was to study the activity and localization of phosphorylase (EC 2.4.1.1) in storage parenchyma under these conditions. Miniplants of Solanum tuberosum L. (cv Adreta) were grown in culture for 30 days for both the vertical control and the horizontal clinorotation. During long-term clinorotation, an acceleration of minituber formation, and an increase of amyloplast number and size in storage parenchyma cells, as well as increased starch content, was observed in the minitubers. The differences among cytochemical reaction intensity, activity of phosphorylase, and carbohydrate content in storage parenchyma cells of minitubers grown in a horizontal clinostat were established by electron-cytochemical and biochemical methods. It is shown that high phosphorylase activity is correlated with increased starch content during extended clinorotation. The results demonstrate the increase in carbohydrate metabolism and possible accelerated growth of storage organs under the influence of microgravity, as mimicked by clinorotation; therefore, clinorotation can be used as a basis for future studies on mechanisms of starch synthesis under microgravity.     

Population dynamics of transgenic microorganisms in the different microecosystem conditions.

The role of key environmental factors in adaptation of spore-forming and non-spore-forming transgenic microorganisms (TM) have been studied in model ecosystems. Model TM Escherichia coli Z905 (bearing plasmid genes of bacterial luminescence Ap (r) Lux+) has been found to have a higher adaptation potential than TM Bacillus subtilis 2335/105 (bearing genes of human alpha 2-interferon Km (r) Inf+), planned for employment as a living vaccine under varying environmental conditions. Effects of abiotic factors on migration of natural and recombinant plasmids between microorganisms under model ecosystem conditions has been estimated. The transgenic microorganisms with low copy number survived better under introduction conditions in the microcosms studied. This trend has been shown to be independent of the microcosm type and its complexity. Grant numbers: 99-04-96017, 25, 00-07-9011.     

Super-optimal CO2 reduces wheat yield in growth chamber and greenhouse environments.

Seven growth chamber trials (six replicate trials using 0.035, 0.12, and 0.25% CO2 in air and one trial using 0.12, 0.80, and 2.0% CO2 in air) and three replicate greenhouse trials (0.035, 0.10, 0.18, 0.26, 0.50, and 1.0% CO2 in air) compare the effects of super-optimal CO2 on the seed yield, harvest index, and vegetative growth rate of wheat (Triticum aestivum L. cvs. USU-Apogee and Veery-10). Plants in the growth chamber trials were grown hydroponically under fluorescent lamps, while the greenhouse trials were grown under sunlight and high pressure sodium lamps and in soilless media. Plants in the greenhouse trials responded similarly to those in the growth chamber trials; maximum yields occurred near 0.10 and 0.12% CO2 and decreased significantly thereafter. This research indicates that the toxic effects of elevated CO2 are not specific to only one environment and has important implications for the design of bio-regenerative life support systems in space, and for the future of terrestrial agriculture.     

Spirulina acceptability trials in rats. A study for the "MELISSA" life-support system.

Groups of five rats were fed for sixteen weeks a slightly deficient diet, supplemented with 0-40% of a dried preparation of the blue-green alga Spirulina as a protein source. Control groups were fed a normal rat diet. No significant differences between groups were found in food intake, growth rate or carbon dioxide production. All animals remained apparently healthy, and had similar organ weights. The study suggests that Spirulina may be used as a protein source in rat diets.