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NASA NTRS2007

Benchtop Detection of Proteins

A process, and a benchtop-scale apparatus for implementing the process, have been developed to detect proteins associated with specific microbes in water. The process and apparatus may also be useful for detection of proteins in other, more complex liquids. There may be numerous potential applications, including monitoring lakes and streams for contamination, testing of blood and other bodily fluids in medical laboratories, and testing for microbial contamination of liquids in restaurants and industrial food-processing facilities. A sample can be prepared and analyzed by use of this process and apparatus within minutes, whereas an equivalent analysis performed by use of other processes and equipment can often take hours to days. The process begins with the conjugation of near-infrared-fluorescent dyes to antibodies that are specific to a particular protein. Initially, the research has focused on using near-infrared dyes to detect antigens or associated proteins in solution, which has proven successful vs. microbial cells, and streamlining the technique in use for surface protein detection on microbes would theoretically render similar results. However, it is noted that additional work is needed to transition protein-based techniques to microbial cell detection. Consequently, multiple such dye/antibody pairs could be prepared to enable detection of multiple selected microbial species, using a different dye for each species. When excited by near-infrared light of a suitable wavelength, each dye fluoresces at a unique longer wavelength that differs from those of the other dyes, enabling discrimination among the various species. In initial tests, the dye/antibody pairs are mixed into a solution suspected of containing the selected proteins, causing the binding of the dye/antibody pairs to such suspect proteins that may be present. The solution is then run through a microcentrifuge that includes a membrane that acts as a filter in that it retains the dye/antibody/protein complexes while allowing any remaining unbound dye/antibody pairs to flow away. The retained dye/antibody/protein complexes are transferred to a cuvette, wherein they are irradiated with light from a miniature near-infrared laser delivered via a fiber-optic cable. The resulting fluorescence from the dye(s) is measured by use of a miniature spectrometer, the output of which is digitized, then analyzed by laptop computer. The software running in the computer identifies the protein species by the wavelengths of their spectral peaks and determines the amounts of the proteins, and thus, one day, microbes of the various species from the intensities of the peaks. The abovementioned removal of the unbound dye/antibody pairs during centrifugation prevents false positive readings. The process proves successful in detecting proteins in solution and thus can now be employed for use in microbe detection.

Scardelletti, Maximilian C.↗

NASA NTRS2021

Microbial Food Safety in Space Production Systems

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.

microbiology↗

DOE OSTI2024

Editorial: Water supply sustainability and challenges in Asian megadeltas under global change

Globally, riverine deltas comprise barely 1% of land area, but they support the livelihoods of more than 500 million people (Moorhouse et al., 2021). The Asian megadeltas (e.g., Indus, Bengal, Mekong, Red River, and Yangtze) and their riverine floodplains are the most dynamic economic and environmental systems in the world. These low-lying coastal landforms support surface water bodies and highly productive aquifer systems. However, water resources of the Asian megadeltas are extremely vulnerable to geogenic (e.g., arsenic) and anthropogenic (e.g., bacteria) contamination, increased water salinisation from rising sea levels (Figure 1), and chemicals (e.g., fertilizers, pesticides) from land-use practices (Mukherjee et al., 2024) – all of which are threatening the sustainability of water supply and food security in the region. This Research Topic collection features five original articles that explore water chemistry including arsenic and bacteriological contamination, water salinisation, water-energy-food nexus in irrigated agriculture, and groundwater depletion resulting from irrigation, rapid urbanization, and climate change. Although the Research Topic collection does not include case studies from all the Asian Megadeltas shown in Figure 1, the relevance of multi-hazard driven water risks to all deltaic and river floodplain environments around the world is well-illustrated.

54 ENVIRONMENTAL SCIENCES↗

DOE OSTI2024

Mercury bioaccumulation and Hepatozoon spp. infections in two syntopic watersnakes in South Carolina

Mercury (Hg) is a ubiquitous environmental contaminant known to bioaccumulate in biota and biomagnify in food webs. Parasites occur in nearly every ecosystem and often interact in complex ways with other stressors that their hosts experience. Hepatozoon spp. are intraerythrocytic parasites common in snakes. The Florida green watersnake (Nerodia floridana) and the banded watersnake (Nerodia fasciata) occur syntopically in certain aquatic habitats in the Southeastern United States. The purpose of this study was to investigate relationships among total mercury (THg) concentrations, body size, species,habitat type and prevalence and parasitemia of Hepatozoon spp. infections in snakes. In the present study, we sampled N. floridana and N. fasciata from former nuclear cooling reservoirs and isolated wetlands of the Savannah River Site in South Carolina. We used snake tail clips to quantify THg and collected blood samples for hemoparasite counts. Our results indicate a significant, positive relationship between THg and snake body size in N. floridana and N. fasciata in both habitats. Average THg was significantly higher for N. fasciata compared to N. floridana in bays (0.22 ± 0.02 and 0.08 ± 0.006 mg/kg, respectively; p < 0.01), but not in reservoirs (0.17 ± 0.02 and 0.17 ± 0.03 mg/kg, respectively; p = 0.29). Sex did not appear to be related to THg concentration or Hepatozoon spp. infections in either species. In conclusion, we found no association between Hg and Hepatozoon spp. prevalence or parasitemia; however, our results suggest that species and habitat type play a role in susceptibility to Hepatozoon spp. infection.

63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTAN↗

DOE OSTI2023

Molecular Transformation and Metabolic Insights of Microbial Electrolysis Treatment and Valorization of Post-hydrothermal Liquefaction Wastewater

Hydrothermal liquefaction (HTL) presents a promising approach for the conversion of wet waste into biocrude and biofuels. However, the post-hydrothermal liquefaction wastewater (PHW) poses significant challenges for treatment and valorization due to its high concentration and complex nature. In this study, we investigated the conversion pathways of major organic contaminants within the microbial electrolytic treatment of PHW from food waste HTL. To achieve this, we employed high-performance liquid chromatography (HPLC) and 2D nuclear magnetic resonance (NMR). Our findings demonstrate volatile fatty acids (VFAs) and monohydric and polyhydric alcohols were effectively transformed through the synergistic metabolism of fermentative and electroactive bacteria, which led to over 70% COD removal of the recalcitrant compounds and a record high H2 production rate (1.62 L L -1 d -1 ). We also employed the liquid-state 15N NMR on wastewater samples for the first time and revealed that the nitrogen-containing heteroaromatics were persistent to microbial electrochemical treatment. By integrating the chemical profiles with bioanode community profiles, we constructed a metabolic network that provides insights for enhancing treatment efficiency and facilitating resource recovery.

08 HYDROGEN↗

DOE OSTI2022

Genome-Wide Association Study Reveals Complex Genetic Architecture of Cadmium and Mercury Accumulation and Tolerance Traits in Medicago truncatula

Heavy metals are an increasing problem due to contamination from human sources that and can enter the food chain by being taken up by plants. Understanding the genetic basis of accumulation and tolerance in plants is important for reducing the uptake of toxic metals in crops and crop relatives, as well as for removing heavy metals from soils by means of phytoremediation. Following exposure of Medicago truncatula seedlings to cadmium (Cd) and mercury (Hg), we conducted a genome-wide association study using relative root growth (RRG) and leaf accumulation measurements. Cd and Hg accumulation and RRG had heritability ranging 0.44 – 0.72 indicating high genetic diversity for these traits. The Cd and Hg trait associations were broadly distributed throughout the genome, indicated the traits are polygenic and involve several quantitative loci. For all traits, candidate genes included several membrane associated ATP-binding cassette transporters, P-type ATPase transporters, oxidative stress response genes, and stress related UDP-glycosyltransferases. The P-type ATPase transporters and ATP-binding cassette protein-families have roles in vacuole transport of heavy metals, and our findings support their wide use in physiological plant responses to heavy metals and abiotic stresses. We also found associations between Cd RRG with the genes CAX3 and PDR3, two linked adjacent genes, and leaf accumulation of Hg associated with the genes NRAMP6 and CAX9. When plant genotypes with the most extreme phenotypes were compared, we found significant divergence in genomic regions using population genomics methods that contained metal transport and stress response gene ontologies. Several of these genomic regions show high linkage disequilibrium (LD) among candidate genes suggesting they have evolved together. Minor allele frequency (MAF) and effect size of the most significant SNPs was negatively correlated with large effect alleles being most rare. This is consistent with purifying selection against alleles that increase toxicity and abiotic stress. Conversely, the alleles with large affect that had higher frequencies that were associated with the exclusion of Cd and Hg. Overall, macroevolutionary conservation of heavy metal and stress response genes is important for improvement of forage crops by harnessing wild genetic variants in gene banks such as the Medicago HapMap collection.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS1997

NASA tests composters for space

For long term missions, composters may be an integral part of a life support system that provides edible food crops, extracts nutrients from plant biomass and removes contaminants from the recycling stream.

Plants/growth & development↗

NASA NTRS1992

Thin-Membrane Sensor With Biochemical Switch

Modular sensor electrochemically detects chemical or biological agent, indicating presence of agent via gate-membrane-crossing ion current triggered by chemical reaction between agent and recognition protein conjugated to channel blocker. Used in such laboratory, industrial, or field applications as detection of bacterial toxins in food, military chemical agents in air, and pesticides or other contaminants in environment. Also used in biological screening for hepatitis, acquired immune-deficiency syndrome, and like.

Case, George D.↗

NASA NTRS1998

Enhanced Detection of Vibrio Cholerae in Oyster Homogenate Based on Centrifugal Removal of Inhibitory Agents

The disease cholera, caused by Vibrio cholerae, has been associated with consumption of contaminated seafood, including raw oysters. Detection of V. cholerae in foods typically involves blending the oysters, diluting the homogenate in alkaline peptone water (APW), overnight enrichment, and isolation on selective agar. Unfortunately, the oyster homogenate must be diluted to large volumes because lower dilutions inhibit the growth of V. cholerae. The goals of this study were to develop an alternative to large dilutions and to evaluate the basis for the inhibition observed in lower dilutions of oyster homogenates. Centrifugation of oyster homogenates at 10,000 x g for 15 min, followed by enrichment of the resulting pellet in APW, was found to eliminate the inhibition of V. cholerae growth. Inhibition appears not to be due to competing microflora but to a component(s) released when V. cholerae grows in the presence of oyster homogenate. The inhibitory component(s) kills the V. cholerae after the cell concentration reaches > 10(exp 8) cells/mL, rather than initially preventing their growth. The pH also declines from 8.0 to 5.5 during this period; however, the pH decline by itself appears not to cause V. cholerae death. Seven strains of V. cholerae (01 and non-01) and two strains of V. vulnificus were susceptible to the inhibitory agent(s). However, other Vibrio and non-Vibrio species tested were not inhibited by the oyster homogenates. Based on digestion of oyster homogenates with pronase, trypsin and lipase, the inhibitory reaction involves a protein(s). In a preliminary trial with oyster homogenate seeded with 1 cfu/g of V. cholerae, the modified centrifugation technique detected a slightly higher percentage of samples at a 1:10 dilution than the standard FDA Bacteriological Analytical Method (BAM) detected in uncentrifuged oyster homogenate at a 1:100 dilution. V. cholerae in seeded samples could also be detected more frequently by the modified centrifugation method than by PCR at a 1:10 dilution.

Alexander, Donita↗

NASA NTRS2000

Microwave powered sterile access port

A device and method for elimination of contamination during transfer of materials either into or from bioreactors, food containers, or other microbially vulnerable systems. Using microwave power, thermal sterilizations of mating fixtures are achieved simply, reliably, and quickly by the volatilization of small quantities of water to produce superheated steam which contacts all exposed surfaces.

Sauer, Richard L.↗

DOE OSTI2024

Analysis of biokinetic parameters reveals patterns in mercury accumulation across aquatic species

Mercury (Hg) is a potent neurotoxicant and poses a risk to human health through the ingestion of Hg-contaminated fish. Mercury, especially in its organic form methylmercury (MeHg), biomagnifies up food chains such that even small aqueous concentrations of Hg can result in significant concentrations of total Hg in fish. Understanding the ecological and human health risks associated with Hg and MeHg exposure requires an understanding of the factors that affect its bioaccumulation in aquatic species. We compiled estimates of three biokinetic parameters: uptake rate (k u ), assimilation efficiency (AE), and efflux rate (k e ). These parameters describe contaminant uptake from aqueous (k u ) and dietary (AE) exposure and the rate of excretion (k e ). We found parameter values for 38 and 34 different species of fish and aquatic invertebrates, respectively, and collected 502 parameter values in total. Here, we used a machine learning technique to establish the relationships between experimental and physiological variables and these parameter values. We found differences in which variables were associated with biokinetic parameter values for fish and aquatic invertebrates. The form of Hg was the most impactful variable, influencing values of all parameters except k u for invertebrates, for which aqueous exposure time was the only significant predicator variable. The parameter k e were the only values significantly influenced by more than one variable, with water type (freshwater, brackish, or marine), organism weight, and form of Hg significantly impacting parameter values for fish and/or invertebrates. To our knowledge, this study represents the most extensive review of biokinetic parameters of Hg and MeHg accumulation in aquatic organisms. Environmental parameters found to significantly impact Hg and MeHg bioaccumulation in past studies were not identified as important in our analyses across aquatic ecosystems and species. Our dataset and analysis reveal novel patterns that may help us better understand and manage Hg bioaccumulation.

54 ENVIRONMENTAL SCIENCES↗

NASA NTRS2013

Microbial Monitoring of the International Space Station

Humans living and working in the harsh environment of space present many challenges for habitability engineers and microbiologists. Spacecraft must provide an internal environment in which physical (gas composition, pressure, temperature, and humidity), chemical, and biological environmental parameters are maintained at safe levels. Microorganisms are ubiquitous and will accompany all human-occupied spacecraft, but if biological contamination were to reach unacceptable levels, long-term human space flight would be impossible. Prevention of microbiological problems, therefore, must have a high priority. Historically, prevention of infectious disease in the crew has been the highest priority, but experience gained from the NASA-Mir program showed that microbial contamination of vehicle and life-support systems, such as biofouling of water and food, are of equal importance. The major sources of microbiological risk factors for astronauts include food, drinking water, air, surfaces, payloads, research animals, crew members, and personnel in close contact with the astronauts. In our efforts to eliminate or mitigate the negative effects of microorganisms in spacecraft, the National Aeronautics and Space Administration (NASA) implemented comprehensive microbial analyses of the major risk factors. This included the establishment of acceptability requirements for food, water, air, surfaces, and crew members. A robust monitoring program was then implemented to verify that the risks were within acceptable limits. Prevention of microbiological problems is preferred over mitigation of problems during flight, and preventive steps must begin very early in the design phase. Spacecraft development must include requirements to control free water from humidity, condensate, hygiene activities, and other releases. If water is available, microbes are likely to grow because sufficient nutrients are potentially available. Materials selected for the spacecraft must not promote or support microbial growth. Air filtration can dramatically reduce the number of airborne bacteria, fungi, and particulates in spacecraft breathing air. Waterborne bacteria can be reduced to acceptable levels by thermal inactivation of bacteria during water processing, along with a residual biocide, and filtration at the point of use can ensure safety. System design must include onboard capability to achieve recovery of the system from contamination. Robust housekeeping procedures that include periodic cleaning and disinfection will prevent high levels of microbial growth on surfaces. Food for consumption in space must be thoroughly tested for excessive microbial content and pathogens before launch. Thorough preflight examination of flight crews, consumables, payloads, and the environment can greatly reduce pathogens in spacecraft. Many of the lessons learned from the Space Shuttle and previous programs were applied in the early design phase of the International Space Station, resulting in the safest space habitat to date. This presentation describes the monitoring program for the International Space Station and will summarize results from preflight and on-orbit monitoring.

Pierson, Duane L.↗

DOE OSTI2024

Ionic liquid mediated one-step perpendicular assembly in block copolymer thin films for ultrafiltration membrane applications

The rapid rise of oil and gas, petrochemical, food processing, and pharmaceutical industries has added a complex mixture of contaminants like oil, particulates, metals, and other organic compounds to wastewater. Multipurpose membranes with precise pore structure can offer a solution to this problem and block copolymers (BCP) can be used to achieve such structures. Achieving vertical assembly of BCP domains provides a perfect template for developing uniform through film channels for separation and transport of material, besides being useful for the rectification of defects in photolithography patterns. Producing such morphologies may require extensive film/substrate processing and is not always feasible for scale-up. With this article, we demonstrate a facile solution casting method to induce vertical domain assembly in as-cast diblock copolymer thin films in the presence of an ionic liquid (IL) additive that preferentially segregates to one block and neutralizes interfacial interactions. We also show the tunability of domain sizes by controlling the additive concentration. These vertically aligned morphologies are important for the development of next-generation lithographic techniques and form excellent templates for ultrafiltration membranes with uniform pore sizes. In conclusion, this article demonstrates how IL additives can be used to obtain stable non-equilibrium morphologies in as-cast BCP films and the effect of BCP molecular mass, block volume fractions, and IL content on self-assembly.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS1973

Mercury: Aspects of its ecology and environmental toxicity

A study was conducted to determine the effects of mercury pollution on the environment. The possible sources of mercury contamination in sea water are identified. The effects of mercury on food sources, as represented by swordfish, are analyzed. The physiological effects of varying concentrations of mercury are reported. Emphasis is placed on the situation existing in the Hawaiian Islands.

Siegel, S. M.↗

NASA NTRS1989

Controlled Ecological Life Support System

The NASA CELLS program is based upon the integration of biological and physiochemical processes in order to produce a system that will produce food, a breathable atmosphere, and potable water from metabolic and other wastes. The CELSS concept is described and a schematic system diagram is provided. Central to the CELSS concept is the Plant Growth Chamber, where green plant photosynthesis produces food, and aids in the production of oxygen and water. Progress to date at the Breadboard Facility at the Kennedy Space Center is summarized. The Breadboard Facility will implement the basic techniques and processes required for a CELSS based on photosynthetic plant growth in a ground-based system of practical size and results will be extrapolated to predict the performance of a full-sized system. Current available technology and near-future forecasts for plant growth techniques (focusing on maximum productivity), food sources (to select optimal CELSS plants), and waste management and contaminant control are discussed.

Averner, Maurice M.↗

NASA NTRS1994

Advanced Life Support Systems: Opportunities for Technology Transfer

NASA's future missions to explore the solar system will be of long-duration possibly lasting years at a time. Human life support systems will have to operate with very high reliability for these long periods with essentially no resupply from Earth. Such life support systems will make extensive use of higher plants, microorganisms, and physicochemical processes for recycling air and water, processing wastes, and producing food. Development of regenerative life support systems will be a pivotal capability for NASA's future human missions. A fully functional closed loop human life support system currently does not exist and thus represents a major technical challenge for space exploration. Technologies where all life support consumables are recycled have many potential terrestrial applications as well. Potential applications include providing human habitation in hostile environments such as the polar regions or the desert in such a way as to minimize energy expenditures and to minimize negative impacts on those often ecologically-sensitive areas. Other potential applications include production of food and ornamental crops without damaging the environment from fertilizers that contaminate water supplies; removal of trace gas contaminants from tightly sealed, energy-efficient buildings (the so-called sick building syndrome); and even the potential of gaining insight into the dynamics of the Earth's biosphere such that we can better manage our global environment. Two specific advanced life support technologies being developed by NASA, with potential terrestrial application, are the zeoponic plant growth system and the Hybrid Regenerative Water Recovery System (HRWRS). The potential applications for these candidate dual use technologies are quite different as are the mechanisms for transfer. In the case of zeoponics, a variety of commercial applications has been suggested which represent potentially lucrative markets. Also, the patented nature of this product offers opportunities for licensing to commercial entities. In the case of the HRWRS, commercial markets with broad applications have not been identified but some terrestrial applications are being explored where this approach has advantages over other methods of waste water processing. Although these potential applications do not appear to have the same broad attraction from the standpoint of rapid commercialization, they represent niches where commercialization possibilities as well as social benefits could be realized.

Fields, B.↗

NASA NTRSDate not supplied

Space Crop Production

As astronauts venture farther from Earth, and for longer periods, food will become increasingly critical. Crop production can supplement a packaged diet to provide additional nutrients and variety for astronauts. Testing with the Veggie and Advanced Plant Habitat chambers on the International Space Station is allowing us to understand the impacts of gravity and spaceflight on crop growth and nutritional content, and the importance of plants to astronauts living and working away from our home planet. The space environment presents unique challenges to crop production, and technology and knowledge gaps have been identified. Key gaps include how to: • Identify and demonstrate effective options to provide both water and oxygen to the root zone in microgravity • Understand the deep space radiation impacts on seeds and plants • Investigate the relationship between microbiomes and food safety • Store and handle seeds to ensure they are viable, free of contaminants and long-lived • Identify / develop potential crops suitable for the space environment • Understand automation and human factors • Determine scalability for different concepts and architectures Filling these gaps will help enable future human exploration and move us toward Earth-independence.

Veggie↗

DOE OSTI2020

Compromise of Personal Protective Clothing from Liquid Exposure

Following critiques of multiple personal contamination events from entries into the Oak Ridge National Laboratory’s Spallation Neutron Source transfer bay, it was considered that the most likely causes for contamination were personal protective clothing doffing errors or that moisture (sweat) allowed contamination to wick through the protective clothing. To pursue this issue, radiological protection staff looked more closely, however, at the specific area of the clothing where contamination was highest; under enhanced lighting and photochromic manipulation, there appeared to have been some type of moisture in the area. Recognizing the possibility that moisture may have allowed for migration of contamination through the clothing, further experiments were undertaken to determine under which conditions this transport might have occurred. The objective for this work was to identify the susceptibility of different types of personal protective clothing to various liquids encountered in the workplace.Method: Several tests were performed to determine if perspiration had enabled migration of contamination, and to identify what other liquids might have affected contamination transport. Two layers of personal protective clothing were subjected to static conditions and dynamic conditions, to include active rubbing of the materials while wet. Food dye added to each of the liquids tested enabled visual indications of liquid breakthrough. Additional tests were conducted to see if solid contamination could be transported through the materials along with the liquids. As a result, it was found that all but one type of non-rubberized personal protective clothing in use at Oak Ridge National Laboratory were permanently compromised to some extent by the solvents used for decontamination. It was determined that most common cleaning agents immediately and permanently destroyed the hydrophobic nature of several of the splash-resistant protective clothing materials, allowing for radioactive contamination to penetrate through the material to the worker. Work around wet surfaces or performing wet decontamination will only be performed in protective clothing known to prevent transport of the wetting agent.

61 RADIATION PROTECTION AND DOSIMETRY↗