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Directed plant cell-wall accumulation of iron: Embedding co-catalyst for efficient biomass conversion

Plant lignocellulosic biomass is an abundant, renewable feedstock for the production of biobased fuels and chemicals. Previously, we showed that iron can act as a co-catalyst to improve the deconstruction of lignocellulosic biomass. However, directly adding iron catalysts into biomass prior to pretreatment is diffusion limited, and increases the cost of biorefinery operations. Recently, we developed a new strategy for expressing iron-storage protein ferritin intracellularly to accumulate iron as a catalyst for the downstream deconstruction of lignocellulosic biomass. In this study, we extend this approach by fusing the heterologous ferritin gene with a signal peptide for secretion into Arabidopsis cell walls (referred to here as FerEX). The transgenic Arabidopsis plants. FerEX. accumulated iron under both normal and iron-fertilized growth conditions; under the latter (iron-fertilized) condition, FerEX transgenic plants showed an increase in plant height and dry weight by 12 and 18 %, respectively, compared with the empty vector control plants. The SDS- and native-PAGE separation of cell-wall protein extracts followed by Western blot analyses confirmed the extracellular expression of ferritin in FerEX plants. Meanwhile, Perls' Prussian blue staining and X-ray fluorescence microscopy (XFM) maps revealed iron depositions in both the secondary and compound middle lamellae cell-wall layers, as well as in some of the corner compound middle lamella in FerEX. Remarkably, their harvested biomasses showed enhanced pretreatability and digestibility, releasing, respectively, 21 % more glucose and 34 % more xylose than the empty vector control plants. These values are significantly higher than those of our recently obtained ferritin intracellularly expressed plants. This study demonstrated that extracellular expression of ferritin in Arabidopsis can produce plants with increased growth and iron accumulation, and reduced thermal and enzymatic recalcitrance. Here, the results are attributed to the intimate colocation of the iron co-catalyst and the cellulose and hemicellulose within the plant cell-wall region, supporting the genetic modification strategy for incorporating conversion catalysts into energy crops prior to harvesting or processing at the biorefinery.

09 BIOMASS FUELS↗

Spatial arrangement of canopy structure and land-use history alter the effect that herbivores have on plant growth

The spatial arrangement of past and present human activities could affect the strength of species interactions through changes to environmental conditions. To better understand how land-use history might mediate the effect of insect herbivory on the growth of four herbaceous plant species at the edges between woodlands and open savannas, we coupled large-scale manipulations of canopy structure by thinning overstory trees to create savanna habitat in non-agricultural and post-agricultural longleaf pine woodlands with local-scale herbivore exclosures at two distances from habitat edges. Light availability, grasshopper abundance, and herbaceous plant cover all increased in patches that received overstory tree thinning. Land-use history altered the effect that edges created by the overstory thinning treatments had on grasshopper herbivory for one plant species (Carphephorus bellidifolius). The edge between land-use histories also altered herbivore effects on three plant species in thinned patches. These edge effects were driven in part by changes in availability of alternative resources for grasshoppers, shady edges, or grasshopper abundance. Canopy manipulation also had important effects on herbivory that were independent of the habitat edge: grasshopper herbivory reduced plant growth of the smallest species (C. bellidifolius) regardless of canopy manipulation, whereas herbivory on two larger species (Solidago nemoralis and So. odora) reduced plant growth mainly in patches with an intact overstory canopy. Collectively, this work suggests that the species-specific effects of grasshopper herbivory on plant growth were driven by both plant traits (e.g., size or growth rates) and landscape-mediated changes to grasshopper abundance, light availability, or availability of alternative resources for grasshoppers created by land-use history and canopy structure. Furthermore, because the heterogeneity in herbivory revealed by our large-scale experiment is likely linked to plant traits, our results provide a means for understanding the profound, but often idiosyncratic, effects of land-use legacies and edges on plant populations and communities.

60 APPLIED LIFE SCIENCES↗

Simulating Solar Power Plant Variability: A Review of Current Methods

It is important to be able to accurately simulate the variability of solar PV power plants for grid integration studies. We aim to inform integration studies of the ease of implementation and application-specific accuracy of current PV power plant output simulation methods. This report reviews methods for producing simulated high-resolution (sub-hour or even sub-minute) PV power plant output profiles for variability studies and describes their implementation. Two steps are involved in the simulations: estimation of average irradiance over the footprint of a PV plant and conversion of average irradiance to plant power output. Six models are described for simulating plant-average irradiance based on inputs of ground-measured irradiance, satellite-derived irradiance, or proxy plant measurements. The steps for converting plant-average irradiance to plant power output are detailed to understand the contributions to plant variability. A forthcoming report will quantify the accuracy of each method using application-specific validation metrics.

Lave, Matthew Samuel↗

Methods to Automate Assessing the Outage Risk and Technical Specification Compliance for Planned Work and Current Plant Conditions. Light Water Reactor Sustainability Program report

This research effort is a part of the Light Water Reactor Sustainability (LWRS) Program, which is a research and development (R&D) program sponsored by the Department of Energy. The LWRS Program is performed in close collaboration with industry R&D programs that provide the technical foundations for licensing and managing the long term, safe, and economical operation of current nuclear power plants (NPPs). The LWRS Program serves to help the United States (U.S.) nuclear industry adopt new technologies and engineering solutions that facilitate the continued safe operation of these NPPs and the extension of their current operating licenses. The Outage Risk Management Improvement Pilot Project seeks to improve the management of nuclear power plant outages. This is done through the development of tools to assist in evaluating pending activities against requirements to detect undesired interactions. Significant efforts are expended to manage the nuclear risk of an outage. The utilities conduct pre outage risk assessments, based on a very detailed review of the outage schedule, to identify where combinations of outage work and out of service equipment would result in degraded conditions with respect to nuclear safety or regulatory compliance. Probabilistic risk assessment studies are conducted to quantify incremental core damage frequency as a result of the outage activities and system unavailability. These studies are usually presented to site and fleet management, the site plant operational review committee, and the NPP’s independent Nuclear Safety Review Board for concurrence that the outage is planned safely and that reasonable measures have been taken to reduce the added risk of conducting the outage. During the outage, the plant configuration is monitored continuously to ensure that it conforms to the approved safety plan. Deviations must be assessed and approved by management committees and, in some cases, the plant operational review committee. In virtually all outage meetings and job briefings, the current nuclear safety status of the plant is communicated, including information on the specific equipment necessary to meet the requirements of the nuclear safety plan. In addition, the operations and outage organizations implement several layers of physical and administrative barriers to prevent unintended interaction with the systems and equipment credited for nuclear safety. In spite of all these efforts, nuclear safety challenges still occur too frequently during outages. While some of these are due to failures of equipment credited for safety, the majority occur because of human error. These typically involve some form of interaction between work activities and plant configuration changes. Some of them are very subtle and are extremely challenging to detect in advance. Nevertheless, they are not acceptable and represent clear opportunities to improve nuclear safety during outages. This particular report will provide a demonstration of the capabilities in the development of a predictive software platform for improving configuration control and accessing the plant risks associated with outage work protocols and illustrate how the software assists in strategies to minimize these interactions. In addition to maintaining nuclear safety, there are numerous administrative requirements that must be complied with during an outage. This report will discuss the use of logic modelling to monitor planned work against technical specification requirements for given plant conditions and to integrate these requirements with shutdown risk models, clearance order requirements and other plant condition requirements.

22 GENERAL STUDIES OF NUCLEAR REACTORS↗

Failure Analysis for Molten Salt Thermal Energy Storage Tanks for In-Service CSP Plants

Thermal Energy Storage (TES) is a fundamental component in concentrating solar power (CSP) plants to increase the plant's dispatchability, capacity factor, while reducing the levelized cost of electricity. In central receivers CSP plants, nitrate molten salts have been used for several years for operation temperatures of up to 565 degrees C. Despite many efforts to advance nitrate salt to higher operation temperatures (even considering a replacement with molten chloride salts) to achieve higher energy conversion efficiencies, the 565 degrees C temperature is currently considered the state-of-the art. Although molten salt tanks have been broadly deployed in commercial CSP plants worldwide, several failures have been reported in these tanks after a few months or years of operation, causing significant economic loss and mistrust in CSP technologies. Most of these failures are associated with the infancy of the technology and multiple issues related to tank design, fabrication, commissioning, and aggressive operation. A technical standard dedicated to the design and fabrication of molten nitrate TES tanks does not exist today. Current in-service molten salt tanks have been generally designed based on the American Petroleum Institute's (API) 650 and ASME Section II standards. The API 650 code provides guidelines for dimensions and fabrication for oil storage tanks up to 260 degrees C. The ASME standard provides allowable stress values for various materials at a range of temperatures and conditions. Both standards seem to be limited for molten salt TES tanks where high temperatures, thermal cycling, and transient conditions are expected. In 2020, NREL released the Concentrating Solar Power Best Practices Study (NREL/TP-5500-75763) that summarized multiple issues in CSP plants, along with potential alternatives and recommendations to address those issues based on information collected from participants representing about 80% of operating CSP plants in the world. One of the recommendations from this study was the development of accurate and validated models to evaluate the plant's transient operation, capable of capturing the effect of short-term clouds and operator response, while being flexible in being adapted to various spatial and temporal resource data. The "Failure Analysis for Molten Salt Thermal Energy Tanks for In-Service CSP Plants" project was inspired on this recommendation and was focused on (1) the development and validation of a physics-based model for a representative, commercial-scale molten salt tank, (2) performing simulations to evaluate the behavior of the tank as a function of typical plant operation conditions, (3) understanding tank failures mechanisms, (4) determining the residual stress and distortion in the tank floor after welding fabrication and evaluating their impact in the stresses developed in the tank during operation, (5) assessing the impact of key operation parameters on the temperature and stress distribution, (6) conduct a preliminary evaluation of design features to reduce stress and improve tank's reliability, and (7) estimate the tank's service life based on the stress developed under diverse operation scenarios. From the analysis conducted in the project and presented in this report, it was found that maximum stresses surpassing the yield strength point of the stainless steel (SS) 347H are developed on the tank floor near the perimeter. These large stresses are strongly influenced by the initial residual stresses and distortion of the tank floor after welding fabrication. During operation, large stresses are developed in the tank floor at high operation temperatures with large salt inventory levels during transient operation. High stresses are also related to elevated temperature gradients in the tank floor that could be attributed to insufficient mixing within the salt inflow and the salt inventory. Based on the analysis, creep is the predominant failure mechanism. However, the large stress levels could favor the plastic deformation into buckles, and crack formation due to stress relaxation cracking during cycle operation. A lifetime below 3 years was estimated for the typical plant operation conditions and a specific initial residual stress and deformation distribution of the tank floor. The estimated life agrees with the service time to failure reported in several commercial molten salt tanks. Desing and operation guidelines can be extracted from the analysis presented in this report, which could be adopted by tank manufacturers and CSP operators to advance toward an ultimate solution for tank failures by reducing residual and operational stresses to achieve a tank service life of more than 30 years. Addressing failures in molten salt TES tanks is fundamental for the CSP industry's survivability, but it is also important for other industrial and power generation applications using this technology, including nuclear and concentrating solar thermal.

14 SOLAR ENERGY↗

Invasive Plant Species Management Plan for Los Alamos National Laboratory (Rev.3)

Native species are plants and animals that continually occupy a natural range without direct or indirect introduction and/or care by humans. They are adapted to the environmental conditions and processes of the ecosystem in which they reside. A species introduced into a novel ecosystem can either exploit that ecosystem and thrive or be unable to survive in that ecosystem (Hobbs et al. 2006). Alien or non-native species are species that are intentionally or accidentally introduced into a novel ecosystem and are capable of living and propagating within the physical parameters of that ecosystem. Invasive species is a species that is non-native (or alien) to the ecosystem under consideration and a noxious species are those whose introduction causes or is likely to cause economic or environmental harm. The term invasive species is applicable to plants and animals alike; however, this invasive species management plan currently focuses on invasive vegetation. Invasive plant species are usually capable of rapid colonization of disturbed ground, such as after a change in wildfire regime intensity and frequency (Reilly et al. 2020) or anthropogenic ground disturbance (Burke and Grime 1996; Hobbs and Huenneke 1992). Climate change facilitates the spread and establishment of many alien species and creates new opportunities for them to become invasive (Turbelin and Catford 2021). As climate change impacts increase in the coming decades, we may see in an increase in invasive species establishment. See a list of definitions of terms pertinent to this document in Appendix A: Definitions. Los Alamos National Laboratory (LANL) hosts populations of non-native and invasive species all typical of the northern New Mexico region (Martin 2004; NMDA 2020). By implementing an invasive species management plan, LANL will have readily accessible management strategies for invasive species that are found on-site. The benefits of managing invasive species include a decrease in wildland fire risks, an increase in soil productivity, an increase in (productive or beneficial) wildlife habitat, an increase in water quantity and quality, and the restoration of impacted areas (Burke and Grime 1996; Hobbs and Huenneke 1992; D’Antonio and Hobbie 2005; MacDougall et al. 2013; Reilly et al. 2020). The aforementioned benefits from invasive species management directly enable the LANL mission by ensuring compliance requirements are met and site-wide programs, such as the Vulnerability Assessment and Resilience Plan, are supported for a mutually beneficial outcome. An example for the LANL site specifically, controlling non-native annual plant species, for example, could reduce the costs associated with stabilizing soils during the stormwater pollution prevention compliance process. Roadway and utility right-of-way areas are another place where an integrated vegetation management strategy would promote low growing perennial plants in a way that is mutually beneficial to habitat and the institution through lowered maintenance costs. There is also an economic benefit to managing invasive species. In one nationwide study, invasive plants had an estimated impact cost of $190.45 billion (Fantel-Lepczyk et al. 2022). Investing in preventative measures and surveillance could help to offset future control and management costs of invasive species that have the potential to become established. The State of New Mexico has developed plant species lists and recommendations through the New Mexico Department of Agriculture’s Noxious Weeds Management Act, Article 7D (NM Statute § 76-7D-4 2021); however, effective invasive species management must rely on local knowledge of the site and region. Los Alamos County (LAC) has already compiled a target invasive plant species list and species-specific management objectives (Martin 2004). Therefore, the New Mexico Noxious Weed List and the LAC invasive plant species list, as well as management objectives from those documents, are integrated into LANL’s invasive plant species management plan.

54 ENVIRONMENTAL SCIENCES↗

Hanford Site Rare Plant Monitoring Report for Calendar Year 2022

Rare plants are monitored on the Hanford Site as distribution and abundance data of current and historic rare plant populations are necessary to make well-informed environmental and ecological resource management decisions. The Central Hanford Rare Plant Management Plan (DOE/RL-2021-35) guides rare plant monitoring methodology in the context of resource management at the Hanford Site. In 2022, a rare plant monitoring effort occurred on the portion of the Hanford Site (referred to herein as Central Hanford) managed by the U.S. Department of Energy, Richland Operations Office (DOE-RL; Figure 1). The goal of this effort was to revisit known occurrences of upland rare plant populations on Central Hanford to improve baseline information on locations of rare plant populations on the Hanford Site. Rare upland plants were thoroughly surveyed in 2021 but unusually low precipitation that year limited germination of many upland annual plants. Surveys in 2022 focused on rare upland annual species after the Hanford Site received higher than average spring precipitation that was expected to increase germination. Rainfall from April 2022 to June 2022 was 76% higher than average; however, these months were also colder than average. Winter rainfall from January 2022 to March 2022 was 84% lower than average. The high spring precipitation in 2022 appears to have triggered the germination of some populations of rare upland annuals but the dry winter and cold spring may have inhibited others. Population stability, viability, and threats were evaluated and will be used to inform future management of rare plant species.

54 ENVIRONMENTAL SCIENCES↗

The Co-Firing of Pine Biomass and Waste Coal in 100 and 600 MW Power Plants: A Sustainable Approach to Reduce GHG Emissions

Climate change is a global issue that has gained much attention recently. Co-firing biomass with coal/waste coal reduces the electricity sector’s GHG emissions sustainably. This study uses commercial software to model waste coal and biomass co-firing in 100 MW and 600 MW power plants. The objective is to assess the effects of fluid types (subcritical and supercritical), plant capacities (100 MW and 600 MW), boiler types (pulverized coal and circulating fluidized bed boilers), biomass and waste coal co-firing ratios (0:100, 20:80, 40:60, 60:40, 80:20, and 100:0), and carbon capture and storage efficiencies (0%, 90%, 95%, and 97%) on performance parameters such as net plant efficiency, heat rate, net plant CO2 and SO2, and particulate matter emissions. The feedstocks selected for this investigation include anthracite waste coal and loblolly pine biomass. As the biomass fraction increases from 0% to 100%, co-fired power plants net efficiency increases by 3–8%. Supercritical plants had a 6% higher net plant efficiency than the subcritical plants. The study found that the biomass’s high heating value decreased the fuel flow rate and reduced plant CO2 emissions by 10–16%. With 100% biomass power plant feed and 90% carbon capture and storage efficiency, CO2 emissions drop by 83% and SO2 and PM emissions drop to zero.

Bhoi, Prakashbhai R. (ORCID:0000000333104888)↗

Data from: Crop-associated virus infection in a native perennial grass: reduction in plant fitness and dynamic patterns of virus detection

To understand the eco-evolutionary significance of plant viruses in nature, we must (i) quantify the effects of infection on plant fitness and (ii) recognize that native plants are increasingly exposed to crop-associated viruses. Studies of perennials are particularly needed: most of our knowledge of plant-virus interactions is from annuals, yet long-lived species dominate landscapes. Here we used aster models for life-history analysis and longitudinal measures of plant virus status to evaluate multi-year consequences of crop virus infection in a native perennial. We used Barley yellow dwarf virus acquired from wheat to inoculate seedlings of Panicum virgatum L. (switchgrass), a North American prairie grass. We grew inoculated and mock-inoculated individuals of two ecotypes for 3 years in the field. We measured plant size, infection status and fitness components. Aster modelling provided integrated multi-year measures of fitness. Crop virus inoculation reduced multi-year native plant fitness by 30% over 2 years despite generally asymptomatic infection and evidence of resistance. This reduction was greater than predicted from individual fitness components or most size measures. Ecotypes differed in response, with the lowland ecotype experiencing higher apparent recovery from infection. Virus treatment in the upland ecotype delayed flowering phenology and reduced seed filling. Synthesis. Our use of field experimentation, surveys of plant infection status and aster modelling demonstrates a rigorous and broadly applicable approach for quantifying the effects of viruses and other microbes on multi-year plant fitness. We found that a crop virus had negative multi-year effects on native plant fitness even after infection was no longer detected. Viruses may have substantial effects on native vegetation with domestication of landscapes and agricultural expansion.

59 BASIC BIOLOGICAL SCIENCES↗

The Role of Gravity on the Reproduction of Arabidopsis Plants

The presence of gravity as a necessary environmental factor for higher plants to complete their life cycle was examined. Arabidopsis thalliana (L.) Heynh. Columbia strain plants were grown continuously for three generations in a simulated micro-g environment as induced by horizontal clinostats. Growth, development and reproduction were followed. The Arabidopsis plants were selected for three generations on clinostats because: (1) a short life cycle of around 35 days; (2) the cells of third generation plants would in theory be free of gravity imprint; and (3) a third generation plant would therefore more than likely grow and respond like a plant growing in a micro-g environment. It is found that gravity is not a required environmental factor for higher plants to complete their life cycle, at least as tested by a horizontal clinostat. Clinostatting does not prevent the completion of the plant life cycle. However, clinostatting does appear to slow down the reproductive process of Arabidopsis plants. Whether higher plants can continue to reproduce for many generations in a true micro-g environment of space can only be determined by long duration experiments in space.

Hoshizaki, T.↗

P-MASS and P-GBA: Two new hardware developments for growing plants in space

Plant growth, and especially plant performance experiments in microgravity are limited by the currently available plant growth facilities (low light levels, inadequate nutrient delivery and atmosphere conditioning systems, insufficient science instrumentation, infrequent flight opportunities). In addition, mission durations of 10 to 14 days aboard the NSTS Space Shuttle allow for only brief periods of microgravity exposure with respect to the life cycle of a plant. Based on seed germination experiments, using the Generic BioProcessing Apparatus hardware (GBA), two new payloads have been designed specifically for plant growth. These payloads provide new opportunities for plant gravitational and space biology research and emphasize the investigation of plant performance (photosynthesis, biomass accumulations) in microgravity. The Plant-Module for Autonomous Space Support (P-MASS) was designed to utilize microgravity exposure times in excess of 30 days on the first flight of the recoverable COMET satellite (Commercial Experiment Transporter). The Plant-Generic Bioprocessing Apparatus (P-GBA), is designed for the National Space Transportation System (NSTS) Space Shuttle middeck and the SPACEHAB environment. The P-GBA is an evolution from the GBA hardware and P-MASS (plant chamber and instrumentation). The available light levels of both payloads more than double currently available capabilities.

Hoehn, Alexander↗

An Axenic Plant Culture System for Optimal Growth in Long-Term Studies: Design and Maintenance

The symbiotic co-evolution of plants and microbes leads to difficulties in understanding which of the two components is responsible for a given environmental response. Plant-microbe studies greatly benefit from the ability to grow plants in axenic (sterile) culture. Several studies have used axenic plant culture systems, but experimental procedures are often poorly documented, the plant growth environment is not optimal, and axenic conditions are not rigorously verified. We developed a unique axenic system using inert components that promotes plant health and can be kept sterile for at least 70 d. Crested wheatgrass (Agropyron cristatum cv. DII) plants were grown in sand within flow-through glass columns that were positively pressured with filtered air. Plant health was optimized by regulating temperature, light level, CO2 concentration, humidity, and nutrients. The design incorporates several novel aspects, such as pretreatment of the sand with Fe, graduated sand layers to optimize the air-water balance of the root zone, and modification of a laminar flow hood to serve as a plant growth chamber. Adaptations of several sterile techniques were necessary for maintenance of axenic conditions. Axenic conditions were verified by plating and staining leachates as well as rhizoplane stain. This system was designed to study nutrient and water stress effects on root exudates, but is useful for assessing a broad range of plant-microbe-environment interactions. Based on total organic C analysis, 74% of exudates was recovered in the leachate, 6% was recovered in the bulk sand, and 17% was recovered in the rhizosphere sand. Carbon in the leachate after 70 d reached 255 micro-g/d. Fumaric, malic, malonic, oxalic, and succinic acids were measured as components of the root exudates.

Henry, Amelia↗

The Effect of Plant Cultivar, Growth Media, Harvest Method and Post Harvest Treatment on the Microbiology of Edible Crops

Systems for the growth of crops in closed environments are being developed and tested for potential use in space applications to provide a source of fresh food. Plant growth conditions, growth media composition and harvest methods can have an effect on the microbial population of the plant, and therefore should be considered along with the optimization of plant growth and harvest yields to ensure a safe and palatable food crop. This work examines the effect of plant cultivar, growth media, and harvest method on plant microbial populations. Twelve varieties of leafy greens and herbs were grown on a mixture of Fafard #2 and Arcillite in the pillow root containment system currently being considered for the VEGGIE plant growth unit developed by Orbitec. In addition, ,Sierra and Outredgeous lettuce varieties were grown in three different mixtures (Fafard #2, Ardllite, and Perlite/Vermiculite). The plants were analyzed for microbial density. Two harvest methods, "cut and come again" (CACA) and terminal harvest were also compared. In one set ofexpe'riments red leaf lettuce and mizuna were grown in pots in a Biomass Production System for education. Plants were harvested every two weeks by either method. Another set of experiments was performed using the rooting pillows to grow 5 varieties of leafy greens and cut harvesting at different intervals. Radishes were harvested and replanted at two-week intervals. Results indicate up to a 3 IOglO difference in microbial counts between some varieties of plants. Rooting medium resulted in an approximately 2 IOglO lower count in the lettuce grown in arscillite then those grown in the other mixtures. Harvest method and frequency had less impact on microbial counts only showing a significant increase in one variety of plant. Post harvest methods to decrease the bacterial counts on edible crops were investigated in these and other experiments. The effectiveness of PRO-SAN and UV-C radiation is compared.

Hummerick, Mary P.↗

Does Seed Sanitization Affect the Plant Rhizosphere Microbiome and Its Ability to Compete with the Human Associated Pathogen, E. coli on Salad Crops?

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.

Khodadad, Christina L. M.↗

The Plant Water Management Experiments: Hydroponics 3 & 4

As humans consider longer-duration missions in space, NASA has identified production of fresh vegetables aboard spacecraft as beneficial for crew nutrition, mental wellbeing, and enabling bioregenerative life support (i.e., air, water, and waste processing). Current low-g plant growth techniques have successfully grown a variety of leafy and flowering plants. However, unique microgravity fluidics challenges to maintain plant moisture levels persist which hamper overall system reliability. The Plant Water Management (PWM) experiments seek to demonstrate low-cost, low-mass, reusable plant growth systems that leverage recent advances in low-g capillary fluidics phenomena to provide routine, largely passive, water delivery to plants. This paper presents findings from a series of PWM-Hydroponics 3 & 4 experiments, which were collected during three ISS flight operations that occurred in March, May, and July of 2021. Open hydroponic capillary channel flows with synthetic evapo-transpiring plant models were used. Tests demonstrated flow stability for single and parallel channel flow configurations across a range of flow rates, plant types, and plant arrangements. Technology demonstrations of both passive aeration and bubble phase separation are reported. We provide details of the data reduction and archive. Insights from the successful flight demonstrations provide a foundation from which follow-on PWM-Hydroponics 5 & 6 experiments on ISS, potentially incorporate living plants, are being considered. Xx Summary of positives and negatives..

Marc B. Wasserman↗

A Simulated High CO 2 Spaceflight Environment Increases Plant Preference for Ammonium as a Nitrogen Source.

Future long-duration missions will require a sustainable supply of food to support human crews. The spaceflight cabin environment often contains very high concentrations of CO 2 due to release of CO 2 by astronauts that is not completely scrubbed from the cabin, and it is therefore crucial to understand plant responses to elevated CO 2 (eCO 2 ) environments. Much focus has been given to changes in plant photosynthetic and performance parameters in response to eCO 2 , but the effects of eCO 2 on nitrogen (N) uptake are poorly understood. Shoot nitrate reduction may be reduced at eCO 2 , likely due to less reductant available for nitrate reduction because of reduced photorespiration and increased carbon fixation 1. Relative growth rate can be reduced at eCO 2 when N is provided only as nitrate and can be unaffected when N is supplied as ammonium 1. However, N uptake in response to eCO 2 has, to our knowledge, not yet been studied. An increased ‘preference’ for plants to take up N as ammonium at eCO 2 could have important implications for growth in the space environment, where N is currently only supplied as nitrate. In this study, novel stable isotope approaches were used in conjunction with hydroponics and isotope ratio mass spectrometry to determine the effect of eCO 2 on N preference for ammonium or nitrate in spring barley and lettuce when both N forms are provided equally. Several varieties of spring barley displayed increased ammonium preference at eCO 2 (720 ppm) compared to ambient CO 2 (410 ppm), though this was not true for all varieties 2. In most cases, increases in ammonium preference were driven by increases in ammonium uptake at eCO 2 and not decreases in nitrate uptake. Current research is assessing whether similar responses are observed in the candidate space crop lettuce, at levels of CO 2 like those observed on ISS (3000 ppm), and these results will also be presented. This work will enable the development of optimized nutrient regimes for candidate crops in space environments and the selection of crop varieties adapted to eCO 2 environments. Plants adapted to ammonium nutrition may play a role in future plant-based bioregenerative life support systems with higher ammonium concentrations due to waste recycling 3. Moreover, this research will further our understanding of plant responses to the eCO 2 environment brought about by climate change, allowing the development of future-proof crops that will help to maintain food security. References: 1. Bloom (2015). The increasing importance of distinguishing among plant nitrogen sources. Current Opinion in Plant Biology 25, 10-16. 2. Fountain (2023). Understanding interactions of barley (Hordeum vulgare) with soil nitrogen cycling activity and links to plant nitrogen preference. Ph.D. Thesis, The University of Sheffield. 3. Schiefloe et al. (2023). From urine to food and oxygen: effects of high and low NH4+:NO3- ratio on lettuce cultivated in a gas-tight hydroponic facility. Frontiers in Plant Science 14:1229476.

nitrogen↗

DEVELOPMENT AND TESTING OF FAULT-DIAGNOSIS ALGORITHMS FOR REACTOR PLANT SYSTEMS

Argonne National Laboratory is further developing fault diagnosis algorithms for use by the operator of a nuclear plant to aid in improved monitoring of overall plant condition and performance. The objective is better management of plant upsets through more timely, informed decisions on control actions with the ultimate goal of improved plant safety, production, and cost management. Integration of these algorithms with visual aids for operators is taking place through a collaboration under the concept of an operator advisory system. This is a software entity whose purpose is to manage and distill the enormous amount of information an operator must process to understand the plant state, particularly in off-normal situations, and how the state trajectory will unfold in time. The fault diagnosis algorithms were exhaustively tested using computer simulations of twenty different faults introduced into the chemical and volume control system (CVCS) of a pressurized water reactor (PWR). The algorithms are unique in that each new application to a facility requires providing only the piping and instrumentation diagram (PID) and no other plant-specific information; a subject-matter expert is not needed to install and maintain each instance of an application. The testing approach followed accepted procedures for verifying and validating software. It was shown that the code satisfies its functional requirement which is to accept sensor information, identify process variable trends based on this sensor information, and then to return an accurate diagnosis based on chains of rules related to these trends. The validation and verification exercise made use of GPASS, a one-dimensional systems code, for simulating CVCS operation. Plant components were failed and the code generated the resulting plant response. Parametric studies with respect to the severity of the fault, the richness of the plant sensor set, and the accuracy of sensors were performed as part of the validation exercise. The background and overview of the software will be presented to give an overview of the approach. Following, the verification and validation effort using the GPASS code for simulation of plant transients including a sensitivity study on important parameters will be presented

Fault Diagnosis Algorithm↗

Microalgae Commodities from Coal-Fired Power Plant Flue Gas CO 2 (Draft Final Report, October 1, 2015 to June 30, 2018)

The Orlando Utilities Commission (OUC) operates an approximately 900-MWe coal fired-power plant at Stanton Energy Center (SEC), which was the site for this project to develop an integrated process for microalgae utilization of flue gas CO 2 for the production of biofuels and animal feeds. Microalgae have both potentially high yields (tons biomass/acre-year) and can be used for biofuels or high value animal feeds. The high biomass yields of microalgae cultures allows efficient use of land and water resources. They can also use wastewaters, recycle fertilizers, and have lower overall greenhouse gas emissions than conventional biofuels or animal feeds. However, many technical issues and technology gaps related to large-scale algae biomass production still must be resolved before such a process can be scaled-up. In this project, techno-economic analyses (TEAs) and life cycle assessments (LCAs) were carried out, projecting the costs and impacts of large-scale algae biomass cultivation for production of biofuels and animal feeds using OUC-SEC flue gas CO 2 . In addition, 3.5-m 2 (~ 40-ft 2 ) experimental raceway ponds were used for the cultivation of microalgae at OUC-SEC and also at the University of Florida. A large-scale algae farm was designed for a 1,300-acre site (~500-ha) located near the OUC-SEC plant, consisting of 400 hectares (1,000 acres) of paddle wheel mixed raceway ponds (each 4 ha), receiving flue gas piped approximately 3 km (2 miles) from the SEC plant. In the first year of this project (Budget Period 1, BP1), the TEA and LCA studies focused on the production of biogas produced from algae biomass and its use to replace coal at the power plant. However, due to the resulting very high cost of CO 2 mitigation, the process was modified to produce compressed natural gas (‘renewable natural gas’, RNG) for transportation fuel from the biogas, in conjunction with wastewater treatment by the algae cultures. During Budget Period 2 (BP2), the TEA and LCA studies focused on the production of animal feeds by the algae farm, using fresh water and agricultural fertilizers. In this case, CO 2 mitigation costs would be entirely covered if the algal biomass could be sold at between a two to three-fold premium above soybean feed, based on a higher nutritional value for the algal biomass. In support of the above analyses, microalgae cultivation experiments were carried out for over two years during this project in small 3.5-m 2 ponds at both SEC and the University of Florida. The most important results were the demonstration at SEC of the cultivation, on flue gas CO 2 , of filamentous algae biomass, which can be harvested at much lower cost than processes required for the colloidal algae species, as currently used by almost all algal mass culture biofuels and animal feeds projects. The yield of biogas production from the algal biomass was investigated by the University of Florida, and the value of the algal biomass as animal feed was projected based on biochemical analysis of the SEC ad UF biomasses. The conclusion from TEA/LCA carried out for the biogas case in BP1 was that a 400-ha algae system producing biomethane for co-firing at OUC-SEC could generate 27,500 MWh/yr of renewable power, offsetting 28,400 Mg CO 2eq /yr, which amounted to less than 1% of the CO 2 emitted by the SEC plant. (Mg = metric ton = 1.10 short tons). The overall cost was projected at $820/Mg of CO 2eq reduction, making this process unfeasible under any possible scenario. More favorable economics were projected when the biogas was converted to higher value transportation fuel, RNG, which also would benefit from renewable fuel (RINs) and possibly other credits. However, only by combining RNG production with municipal wastewater treatment in the algal ponds would CO 2 utilization result in no net cost or even be economically beneficial (i.e., generate income to the power plant). However, in that case the amount of CO 2 utilization would be even lower than that in the biogas case, because a large fraction of the carbon required would be supplied from the wastewater and CO 2 recovered during biogas upgrading to RNG. In brief, use of power plant CO 2 emissions for algal biofuels production was not favorable, even with optimistic assumptions about biomass productivity and biogas yields. For the animal feed case, developed during BP2, algal biomass would replace soybean feeds. The same scale 400-ha algal production plant was projected to produce 26,280 Mg per year of animal feed with a utilization of 45,300 Mg flue gas CO 2 taken up by the algal biomass. However, the process would result in a projected reduction of 112,500 Mg CO 2eq emissions per year, or roughly 1.9% of OUC-SEC emissions, mainly due to the large CO 2eq emissions from conventional soybean production. A major factor in this analysis were greenhouse gas emissions from land use changes. The cost of CO 2 utilization was projected at $116/Mg if the biomass sold for the same price as soybean meal, but if sold as a premium feed (e.g., based on high omega-3 fatty acid and carotenoid content), the power plant could actually charge for the CO 2 utilized. These economic projections are sensitive to assumptions underlying the TEA and LCA models, including biomass composition and the financial models used to project CAPEX and OPEX. Further development of an algal animal feed production using coal-fired power plant flue gas CO 2 is recommended based on the TEA and LCA, as well as experimental results. The required research and development is detailed in a technology gap analysis (Section 11), which focuses on the demonstrations of the cultivation of higher value filamentous algal biomass at OUC-SEC.

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