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Response of graywater recycling systems based on hydroponic plant growth to three classes of surfactants

Anionic (sodium laureth sulfate, SLES), amphoteric (cocamidopropyl betaine, CAPB) and nonionic (alcohol polyethoxylate, AE) surfactants were added to separate nutrient film technique (NFT) hydroponic systems containing dwarf wheat (Triticum aestivum cv. USU Apogee) in a series of 21 day trials. Surfactant was added either in a (1). temporally dynamic mode (1-3 g surfactant m(-2) growing area d(-1)) as effected by automatic addition of a 300 ppm surfactant solution to meet plant water demand, or (2). continuous mode (2 g surfactant m(-2) growing area d(-1)) as effected by slow addition (10 mLh(-1)) of a 2000 ppm surfactant solution beginning at 4d after planting. SLES showed rapid primary degradation in both experiments, with no accumulation 24 h after initial addition. CAPB and AE were degraded less rapidly, with 30-50% remaining 24 h after initial addition, but CAPB and AE levels were below detection limit for the remainder of the study. No reductions in vegetative growth of wheat were observed in response to SLES, but biomass was reduced 20-25% with CAPB and AE. Microbial communities associated with both the plant roots and wetted hardware surfaces actively degraded the surfactants, as determined by monitoring surfactant levels following pulse additions at day 20 (with plants) and day 21 (after plant removal). In order to test whether the biofilm communities could ameliorate phytotoxicity by providing a microbial community acclimated for CAPB and AE decay, the continuous exposure systems were planted with wheat seeds after crop removal at day 21. Acclimation resulted in faster primary degradation (>90% within 24h) and reduced phytotoxicity. Overall, the studies indicate that relatively small areas (3-5m(2)) of hydroponic plant systems can process per capita production of mixed surfactants (5-10 g x person(-1)d(-1)) with minimal effects on plant growth.

Hydroponics↗

Mobilifilum chasei: morphology and ecology of a spirochete from an intertidal stratified microbial mat community

Spirochetes were found in the lower anoxiphototrophic layer of a stratified microbial mat (North Pond, Laguna Figueroa, Baja California, Mexico). Ultra-structural analysis of thin sections of field samples revealed spirochetes approximately 0.25 micrometer in diameter with 10 or more periplasmic flagella, leading to the interpretation that these spirochetes bear 10 flagellar insertions on each end. Morphometric study showed these free-living spirochetes greatly resemble certain symbiotic ones, i.e., Borrelia and certain termite spirochetes, the transverse sections of which are presented here. The ultrastructure of this spirochete also resembles Hollandina and Diplocalyx (spirochetes symbiotic in arthropods) more than it does Spirochaeta, the well known genus of mud-dwelling spirochetes. The new spirochete was detected in mat material collected both in 1985 and in 1987. Unique morphology (i.e., conspicuous outer coat of inner membrane, large number of periplasmic flagella) and ecology prompt us to name a new free-living spirochete.

Non-NASA Center↗

Adaptations of Endolithic Communities to Abrupt Environmental Changes in A Hyper-Arid Desert

The adaptation mechanisms of microbial communities to natural perturbations remain relatively unexplored, particularly in extreme environments. The extremophilic communities of halite (NaCl) nodules from the hyper-arid core of the Atacama Desert are self-sustained and represent a unique opportunity to study functional adaptations and community dynamics with changing environmental conditions. We transplanted halite nodules to different sites in the desert and investigated how their taxonomic, cellular, and biochemical changes correlated with water availability, using environmental data modeling and metagenomic analyses. Salt-in strategists, mainly represented by haloarchaea, significantly increased in relative abundance at sites characterized by extreme dryness, multiple wet/dry cycles, and colder conditions. The functional analysis of metagenome-assembled genomes (MAGs) revealed site-specific enrichments in archaeal MAGs encoding for the uptake of various compatible solutes and for glycerol utilization. These findings suggest that opportunistic salt-in strategists took over the halite communities at the driest sites. They most likely benefited from metabolites newly released in the environment by the death of microorganisms least adapted to the new conditions. The observed changes were consistent with the need to maximize cellular bioenergetics when confronted with lower water availability and higher salinity, providing valuable information on microbial community adaptations and resilience to climate change.

Atacama↗

Distinguishing Indigenous from Contaminating Microorganisms in Rock Samples from a Deep Au Mine in South Africa

The concentration and distribution of microbial biomass within deep subsurface rock strata is not well known To date, most analyses are from water samples and a few cores. Hand samples, block samples and cores from an actively mined Carbon Leader ore zone at 3.2 kilometers depth were collected for microbial analyses. The Carbon Leader was comprised of quartz, S-bearing aromatic hydrocarbons, Fe(III) oxyhydroxides, sulfides, uraninite, Au and minor amounts of sulfate. The porosity of the ore was 1% and the maximum pore throat diameter was less than 0.1 microns; whereas, the porosity of the adjacent quartzite was .02 to .9% with a maximum pore throat diameter of 0.9 microns. Rhodamine dye, fluorescent microspheres, microbial enrichments, autoradiography, phospholipid fatty acid (PLEA) and 16S rDNA analyses were performed on these rock samples and the mining water. The date indicate that the levels of solute contamination less than 0.01% for pared rock samples. Despite this low level of contamination, PLEA, microbial enrichment, DNA and tracer analyses and calculations indicate that most of the viable microorganisms in the Carbon Leader represent gram negative aerobic heterotrophs and ammonia oxidizers that are phylogenetically identical or closely related to service water microorganisms. These microbial contaminants probably infiltrated the low permeability rock through mining-induced microfractures. Geochemical data also detected drilling water in a fault zone approx. 1 meter behind the rock face encountered during coring. The mining induced macrofractures that are common at these great depths act as pathways for the drilling water borne microorganisms into the lower temperature zone that extends several meters into rock strata from the rock face. Combined PLEA and T- RFLP analyses of the service water and Carbon Leader samples indicate that the concentration of indigenous microorganisms was less than 10(exp 2) cells/gram. Such a low concentrations result from the submicron pore throat diameters. PLFA. SO4-35 autoradiography and tracer analyses indicate that the bounding quartzite contains thermophilic sulfate reducing bacteria at 10(exp 3) cells/gram that are not attributable to drilling water contamination. The microorganisms may be surviving on sulfate generated by oxidation of sulfide by radiolytic reactions resulting from the high U concentration in the ore zone. The presence of up to 8,000 ppm of Fe(III) oxyhydroxides in the host rock will also act to recycle sulfide generated by the sulfate reducing bacteria into sulfate. The activity of these sulfate-reducing bacteria may be enhanced by mining induced fracturing which can propagate up to 40 meters into virgin rock where the temperatures are ca. 50 C, and decrepitate of sulfate rich fluid inclusions. In ultra deep mines, judicious application of tracers and multiple microbial characterization techniques can distinguish microbial contamination caused by the near field fracturing and drilling water migration from the indigenous microbial communities in rock strata. The importance of far field fracturing on indigenous microbial communities, however, remains unknown.

Onstott, T. C.↗

Extreme environments and exobiology

Ecological research on extreme environments can be applied to exobiological problems such as the question of life on Mars. If life forms (fossil or extant) are found on Mars, their study will help to solve fundamental questions about the nature of life on Earth. Extreme environments that are beyond the range of adaptability of their inhabitants are defined as "absolute extreme". Such environments can serve as terrestrial models for the last stages of life in the history of Mars, when the surface cooled down and atmosphere and water disappeared. The cryptoendolithic microbial community in porous rocks of the Ross Desert in Antarctica and the microbial mats at the bottom of frozen Antarctic lakes are such examples. The microbial communities of Siberian permafrost show that, in frozen but stable communities, long-term survival is possible. In the context of terraforming Mars, selected microorganisms isolated from absolute extreme environments are considered for use in creation of a biological carbon cycle.

NASA Discipline Exobiology↗

Biodiversity within hot spring microbial mat communities: molecular monitoring of enrichment cultures

We have begun to examine the basis for incongruence between hot spring microbial mat populations detected by cultivation or by 16S rRNA methods. We used denaturing gradient gel electrophoresis (DGGE) to monitor enrichments and isolates plated therefrom. At near extincting inoculum dilutions we observed Chloroflexus-like and cyanobacterial populations whose 16S rRNA sequences have been detected in the 'New Pit' Spring Chloroflexus mat and the Octopus Spring cyanobacterial mat. Cyanobacterial populations enriched from 44 to 54 degrees C and 56 to 63 degrees C samples at near habitat temperatures were similar to those previously detected in mat samples of comparable temperatures. However, a lower temperature enrichment from the higher temperature sample selected for the populations found in the lower temperature sample. Three Thermus populations detected by both DGGE and isolation exemplify even more how enrichment may bias our view of community structure. The most abundant population was adapted to the habitat temperature (50 degrees C), while populations adapted to 65 degrees C and 70 degrees C were 10(2)- and 10(4)-fold less abundant, respectively. However, enrichment at 70 degrees C favored the least abundant strain. Inoculum dilution and incubation at the habitat temperature favored the more numerically relevant populations. We enriched many other aerobic chemoorganotrophic populations at various inoculum dilutions and substrate concentrations, most of whose 16S rRNA sequences have not been detected in mats. A common feature of numerically relevant cyanobacterial, Chloroflexus-like and aerobic chemorganotrophic populations, is that they grow poorly and resist cultivation on solidified medium, suggesting plating bias, and that the medium composition and incubation conditions may not reflect the natural microenvironments these populations inhabit.

Non-NASA Center↗

Lipid Biomarkers for a Hypersaline Microbial Mat Community

The use of lipid biomarkers and their carbon isotopic compositions are valuable tools for establishing links to ancient microbial ecosystems. Various lipids associated with specific microbial groups can serve as biomarkers for establishing organism source and function in contemporary microbial ecosystems (membrane lipids), and by analogy, potential relevance to ancient organic-rich sedimentary rocks (geolipids). As witnessed by the stromatolite record, benthic microbial mats grew in shallow water lagoonal environments. Our recent work has focused on lipid biomarker analysis of a potential analogue for such ancient mats growing in a set of hypersaline evaporation ponds at Guerrero Negro, Baja California Sur, Mexico. The aerobic, surface layer of this mat (0 to 1 mm) contained a variety of ester-bound fatty acids (FA) representing a diverse bacterial population including cyanobacteria, sulphate reducers (SRB) and heterotrophs. Biomarkers for microeukaryotes detected in this layer included sterols, C-20 polyunsaturated FA and a highly branched isoprenoid, diagnostic for diatoms. Cyanobacteria were also indicated by the presence of a diagnostic set of mid-chain methylalkanes. C-28, to C-34 wax esters (WXE) present in relatively small amounts in the upper 3 mm of the mat are considered biomarkers for green non-sulphur bacteria. Ether-bound isoprenoids were also identified although in considerably lower abundance than ester-bound FA (approx. 1:l0). These complex ether lipids included archatol, hydroxyarchaeol and a C-40 tetraether, all in small amounts. After ether cleavage with boron tribromide, the major recovered isoprenyl was a C-30:1. This C(sub 30;1) yelded squalane after hydrogenation, a known geobiomarker for hypersaline environments in ancient oils and sediments. In this mat, it represents the dominant Archaeal population. The carbon isotopic composition of biomarker lipids were generally depleted relative to the bulk organic material (delta C-13 TOC -10%). Most depleted were the cyanobacterial methylalkanes at -27% with FA such as the SRB biomarker, 10- methyl C-I6, somewhat heavier at -16%, and WXE at -17%. The C-30:1 isoprenyl was most enriched with delta C-13 in the -7 to -11% range, much too heavy to represent the methanogen population responsible for mat methane values measured at -60%.

Jahnke, Linda↗

An Unusual Inverted Saline Microbial Mat Community in an Interdune Sabkha in the Rub' Alkhali (the Empty Quarter), UAE: an Analog for Habitats on Present Mars

Salt flats (sabkha) are a recognized habitat for microbial life in desert environments and as analogs for habitats for life on Mars. Here we report on the physical setting and microbiology of interdune sabkhas among the large dunes in the Rub' al Khali (the Empty Quarter) in Liwa Oasis, United Arab Emirates. The salt flats, composed of gypsum and halite, between the dunes are moistened by relatively fresh ground water from below. The result is a salinity gradient that is inverted compared to most salt flat communities with the hypersaline layer at the top and freshwater layers below. We describe and characterize a rich photosynthetically-based microbial ecosystem that is protected from the arid outside environment below the translucent salt crust. Gases collected from sediments under shallow ponds in the sabkha contain methane in concentrations as high as 3400 ppm. The salt layer provides environmental protection to the habitat below and could preserve biomarkers and other evidence for life in the salt after it dries out. Chloride-filled depressions have been identified on Mars and although the surface flow of water is unlikely on Mars today, ground water is possible. Such a near surface system with modern groundwater flowing under ancient salt deposits could be present on Mars and could be accessed by surface rovers.

Hypersaline environments in deserts↗

Responses of Microbes to Modeled Space Radiation

The built environment of spaceships is host to a microbial community that affects crew and craft alike. While the static composition of this community has been characterized and its temporal dynamics examined, the mechanisms controlling its make-up and evolutionary trajectory are not understood. Systematic analyses of microbial diversity show consistent patterns in community composition and function. Understanding these patterns' ecological origins remains a significant challenge, as it requires connecting processes at varying temporal and spatial scales. However, it is clear that the state and trajectories of microbial communities are in-part determined by their physical environment. In this regard, the spaceflight environment includes numerous interacting factors that differentiate it from Earth environments, including an altered atmospheric composition, reduced gravity (and thus altered fluid dynamics), and increased ionizing radiation. These factors impart selective pressures on microbial communities that affect their evolutionary trajectories and thus the risks and benefits these communities represent to crew and craft. The radiation environment of space leads to chronic exposure to low doses and is difficult to mimic on Earth. Thus, little is known about how microbial communities in spacecraft will respond and evolve. Therefore, given the limitations of existing studies, we aim to empirically determine how exposure to low doses of ionizing radiation for thousands of cell divisions affects rates of mutation accumulation in bacteria and the trajectory of their evolution. In this way, we will provide a critical set of data for designing safe and robust space missions. Here we discuss our progress towards this aim, including the construction of exposure facilities, our culturing and analysis approach, and preliminary data.

radiation↗

Microbial mats and modern stromatolites in Shark Bay, Western Australia

Distribution, external morphology, texture, and microbial composition of microbial mats in Hamelin Pool, Shark Bay, Western Australia, have been studied and reviewed along a composite representative profile starting from the permanently submerged zone, across the zones of periodic flooding, toward permanently emerged land and coastal dunes. The following nine types of algal mats have been recognized: colloform, gelatinous, smooth, pincushion, tufted, mamillate, film, reticulate, and blister. Solar ponds represent a particular environment. The mat types represent microbial communities that are characterized by one or more dominant microorganisms. The colonization and stabilization of loose sediment is carried out by a microbial assemblage of generalists that prepare the ground for later replacement and succession by specialized microflora. Lithification of microbial mats takes place periodically, mainly during the austral summer. This process is destructive for the microbial community but increases the preservation potential of the stromatolitic structures.

Golubic, S.↗

Effect of salinity on the preservation of biomarkers in hypersaline microbial mat kerogens

Kerogen is the residue that remains after the minerals and organic compounds have been extracted from a rock using acid and organic solvents, respectively [1]. It is not only the largest organic carbon sink in the geological record but the best-preserved fraction due to its recalcitrance and ability to retain syngenetic source information during deposition [2]. Due to the complex, macromolecular, cross-linked polymeric structure of kerogen, it is difficult to analyse intact, often requiring breakdown into smaller subunits through thermal (pyrolysis) or chemical treatment (chemolysis) [3]. Saline and hypersaline environments host diverse microbial communities, which can trap and bind organic matrices such as cellular components (e.g., extracellular polymeric substances, cell wall, peptidoglycan, sheath material, etc.) with sediments, forming layered, laminated structures called microbial mats. Here we present depth profiles of two hypersaline cores through a microbial mat with differing total salinities. We looked at the composition of organic compounds released from kerogen during high temperature, hydrogen-assisted, pyrolysis to understand the effect of salinity on the composition and timing of their incorporation [4]. Field Site: The Exportadora del Sal S. A. (ESSA) saltern of Guerrero Negro (Baja California Sur, Mexico) lies within a typical sabhka environment adjacent to Laguna Ojo de Liebre. Note the spatial extent of the 13 major ponds of the ESSA system. The GN ponds thrive under well-monitored and stable conditions with steady state accretion and degradation of mat layers (around 0.5–1.4 cm per year). Organic mats typically attain up to 10 cm in thickness, depending on the location, which represents about 60 years of growth since these ponds were established. Sedimentation rate is rapid in these non-lithifying and non-mineralising mats, 1–1.5 cm per year. This environment is ideal for investigating in situ lipid preservation and diagenesis without the influence of constantly changing physical parameters. Methods: P4n5 and P5AB cores were collected as ~9 cm and 6.5 cm fresh cores, respectively, in June 2001 and September 2010, respectively from the ESSA saltern. The P4n5 core (9-9.1% salinity) was taken from Pond 4 near 5 and sub-divided into 10 layers. The P5AB core (11.2% salinity) was taken from Pond 5A near 5B and sub-divided into 8 layers. Lyophilised microbial mat powders were solvent-extracted using a modified Bligh-Dyer method and analysed by gas chromatography-mass spectrometry (GC-MS). Additionally, a subset of layers—layer 4 and layer 7—which represented active biomass and sediment-processed, respectively, were subjected to mild-acid methanolysis. The respective residues from conventional solvent extraction and acid methanolysis were loaded with a molybdenum sulfide catalyst and placed into a stainless-steel reactor. Samples were run on the hydropyrolysis set up (heated to 500 °C with 13-15 MPa of H2 pressure), extracted, and analysed by GC-MS. Results: Molecular profiles from the two ponds were significantly different and reflect the contributions of different photosynthetic and respiratory microbial communities to preserved organic matter. For example, the higher salinity P5AB core showed evidence of greater archaeal inputs (similarly observed in lab culture experiments [5]) as well as potential kerogen-bound carotenoids/carotenoid rearranged products or fragments. A hydrophobic emulsion formed during the acid methanolysis processing of layer 4 from both P4n5 and P5AB and layer 7 of P5AB. Hydropyrolysis of this hydrophobic residues released a greater abundance of polycyclic lipids relative to the pre-extracted control. Implications and Future Work: Catalytic hydropyrolysis of kerogen can rapidly generate abundant saturated pyrolysate products from the bound biomarker pool without altering the structures or stereochemistries of the products [6]. Chemical processing of solvent-extractable residues indicated that cellular matrices such as extracellular polymeric substances (EPS) may play a role in the sequestration of polycyclic lipid biomarkers such as steranes and hopanes. Their higher relative abundance compared to the pre-extracted control in the higher salinity layers and cores preliminarily aligns with this hypothesis, although further experiments are required to confirm this. It has been well-documented that EPS plays an important role in mineralisation, specifically carbonate formation, in microbial ecosystems. EPS can enhance calcium carbonate precipitation by providing diffusion-limited sites that create alkalinity gradients in response to microbial processes [7]. It has been experimentally demonstrated that salinity influences the total amount of EPS (both loosely- and tightly-bound) which increases with increasing salinity [8]. The higher salinity at Guerrero Negro switches the microbial population from filamentous Microcoleus to being dominated by Phormidium, Oscillatoria, and unicellular cyanobacteria. We demonstrated that lipid binding into kerogen via strong covalent linkages occurs at the very earliest stages of sedimentary diagenesis. We are currently investigating the influence of salinity on organic preservation through experimental and modelling approaches. Understanding how key biomarkers transform into preserved organic matter (i.e., from precursor biolipids to bound geolipids) in brine ecosystems will aid the search for organic biosignatures on other planetary bodies, especially Icy Moons and modern Mars.

hypersaline↗

Surface, Water and Air Biocharacterization - A Comprehensive Characterization of Microorganisms and Allergens in Spacecraft Environment

A Comprehensive Characterization of Microorganisms and Allergens in Spacecraft (SWAB) will use advanced molecular techniques to comprehensively evaluate microbes on board the space station, including pathogens (organisms that may cause disease). It also will track changes in the microbial community as spacecraft visit the station and new station modules are added. This study will allow an assessment of the risk of microbes to the crew and the spacecraft. Research Summary: Previous microbial analysis of spacecraft only identify microorganisms that will grow in culture, omitting greater than 90% of all microorganisms including pathogens such as Legionella (the bacterium which causes Legionnaires' disease) and Cryptosporidium (a parasite common in contaminated water) The incidence of potent allergens, such as dust mites, has never been systematically studied in spacecraft environments and microbial toxins have not been previously monitored. This study will use modern molecular techniques to identify microorganisms and allergens. Direct sampling of the ISS allows identification of the microbial communities present, and determination of whether these change or mutate over time. SWAB complements the nominal ISS environmental monitoring by providing a comparison of analyses from current media-based and advanced molecular-based technologies.

Pierson, Duane L.↗

Comparison of the Microbial Diversity and Abundance Between the Freshwater Land-Locked Lakes of Schirmacher Oasis and the Perennially Ice-Covered Lake Untersee in East Antarctica

Extreme conditions such as low temperature, dryness, and constant UV-radiation in terrestrial Antarctica are limiting factors of the survival of microbial populations. The objective of this study was to investigate the microbial diversity and enumeration between the open water lakes of Schirmacher Oasis and the permanently ice-covered Lake Untersee. The lakes in Schirmacher Oasis possessed abundant and diverse group of microorganisms compared to the Lake Untersee. Furthermore, the microbial diversity between two lakes in Schirmacher Oasis (Lake L27C and L47) was compared by culture-based molecular approach. It was determined that L27Chad a richer microbial diversity representing 5 different phyla and 7 different genera. In contrast L47 consisted of 4 different phyla and 6 different genera. The difference in microbial community could be due to the wide range of pH between L27C (pH 9.1) and L47 (pH 5.7). Most of the microbes isolated from these lakes consisted of adaptive biological pigmentation. Characterization of the microbial community found in the freshwater lakes of East Antarctica is important because it gives a further glimpse into the adaptation and survival strategies found in extreme conditions.

Huang, Jonathan↗

Water relations and photosynthesis in the cryptoendolithic microbial habitat of hot and cold deserts

Two cryptoendolithic microbial communities, lichens in the Ross Desert of Antarctica and cyanobacteria in the Negev Desert, inhabit porous sandstone rocks of similar physical structure. Both rock types adsorb water vapor by physical mechanisms unrelated to biological processes. Yet the two microbial communities respond differently to water stress: cryptoendolithic lichens begin to photosynthesize at a matric water potential of -46.4 megaPascals (MPa) [70% relative humidity (RH) at 8 degrees C], resembling thallose desert lichens. Cryptoendolithic cyanobacteria, like other prokaryotes, photosynthesize only at very high matric water potentials [> -6.9 MPa, 90% RH at 20 degrees C].

NASA Discipline Number 52-30↗

Density and composition of microorganisms during long-term (418 day) growth of potato using biologically reclaimed nutrients from inedible plant biomass

A study evaluating alternative methods for long term operation of biomass production systems was recently completed at the Kennedy Space Center (KSC). The 418-day study evaluated repeated batch versus mixed-aged production of potato grown on either standard 1/2-strength Hoagland's nutrient solution or solutions including nutrients recycled from inedible plant material. The long term effects of closure and recycling on microbial dynamics were evaluated by monitoring the microbial communities associated with various habitats within the plant growth system (i.e., plant roots, nutrient solution, biofilms within the hydroponic systems, atmosphere, and atmospheric condensate). Plate count methods were used to enumerate and characterize microorganisms. Microscopic staining methods were used to estunate total cell densities. The primary finding was that the density and composition of microbial communities associated with controlled environmental plant growth systems are stable during long term operation. Continuous production resulted in slightly greater stability. Nutrient recycling, despite the addition of soluble organic material from the waste processing system, did not significantly increase microbial density in any of the habitats.

NASA Center KSC↗

Dormancy and Recovery Testing for Biological Wastewater Processors

Bioreactors, such as aerated membrane type bioreactors have been proposed and studied for a number of years as an alternate approach for treating wastewater streams for space exploration. Several challenges remain before these types of bioreactors can be used in space settings, including transporting the bioreactors with their microbial communities to space, whether that be the International Space Station or beyond, or procedures for safing the systems and placing them into dormant state for later start-up. Little information is available on such operations as it is not common practice for terrestrial systems. This study explored several dormancy processes for established bioreactors to determine optimal storage and recovery conditions. Procedures focused on complete isolation of the microbial communities from an operational standpoint and observing the effects of: 1) storage temperature, and 2) storage with or without the reactor bulk fluid. The first consideration was tested from a microbial integrity and power consumption standpoint; both room temperature (25 C) and cold (4 C) storage conditions were studied. The second consideration was explored; again, for microbial integrity as well as plausible real-world scenarios of how terrestrially established bioreactors would be transported to microgravity and stored for periods of time between operations. Biofilms were stored without the reactor bulk fluid to simulate transport of established biofilms into microgravity, while biofilms stored with the reactor bulk fluid simulated the most simplistic storage condition to implement operations for extended periods of nonuse. Dormancy condition did not have an influence on recovery in initial studies with immature biofilms (48 days old), however, a lengthy recovery time was required (20+ days). Bioreactors with fully established biofilms (13 months) were able to recover from a 7-month dormancy period to steady state operation within 4 days (approximately 1 residence cycle). Results indicate a need for future testing on biofilm age and health and further exploration of dormancy length.

Dormancy↗

VEG-05 Tomato Crop Testing on the International Space Station

Production of fresh, nutritious, and tasty produce for astronauts during spaceflight may provide health-promoting, bioavailable nutrients, enhance the dietary experience, and reduce menu fatigue as we move into longer-duration missions. Growing and caring for plants may also reduce the psychological stresses associated with spaceflight and enhance connection to Earth. A diversity of crops will be required to provide nutrition, variety, and resiliency, however requirements for consistent growth under spaceflight environmental conditions remain unclear. VEG-05 is part of a series of experiments with pick-and-eat salad crops to better define best practices for crop production in space. VEG-05 and predecessor experiments VEG-04A and VEG-04B, grew salad crops in the Veggie vegetable production facilities on the International Space Station using different lighting treatments. In VEG-05 we grew ‘Red Robin’ dwarf cherry tomatoes under two different red: blue lighting spectra. Light can impact the growth habit, yield, nutritional composition, microbial levels, and even flavor attributes within crops, and our goal was to assess these characteristics in ‘Red Robin’ tomatoes during VEG-05. Considerable pre-flight verification testing was performed prior to launch in Nov. 2022. Prior to the flight and ground experiments, lighting in both Veggie units on ISS was measured and lights were standardized between flight and ground hardware. VEG-05 flight operations ran between December 2022 and March 2023, with a ground control initially 48 hours delayed. Due to environmental challenges of very low humidity during the first week of the experiment, seed germination was low for both flight and ground plants. The flight experiment ultimately had 4 - 5 plants per treatment out of the planned 6 plants, but the initial ground control had only three plants in one treatment and none in the other, so this ground control was restarted at the beginning of Feb. 2023 and ran through May, with successful growth of all 12 plants. Both flight and ground control ran 100 days, with harvests of fruit at day 83, day 90, and day 100. Flight plants had uneven growth, and following the early drying events, excess water was frequently observed, which led to a variety of plant stress responses including uneven plant growth, excess adventitious root formation, flower and fruit abortion, and visible microbial growth. In total, from the five surviving red-rich lighted plants, only 5 ripe fruit were produced, and from the four surviving blue-rich lighted plants, 10 fruit were produced with only 6 of these ripe by day 100. Because of the small fruit number and the unsatisfactory growth, crew members were not allowed to consume the tomatoes, and all fruit, as well as large branches with leaves, samples of the adventitious roots, two plant rooting pillows from each treatment, microbial sampling swabs, and some water samples were returned for analysis. Because of the small sample sizes and factors affecting growth on the ISS, objectives of assessing light quality effect (red: blue light treatments) will not be achieved. Revised objectives of this study include to compare stressed flight plants with normal ground plants to determine the impact of plant overwatering stress in space on food safety and the plant microbial community, to determine nutrient content changes in fruit and leaves from stressed plants, and to evaluate stress metabolism changes in returned tissue by transcriptomic analysis. Postflight analysis is underway with the following analyses being conducted: A. culturable microbiology and food safety as well as molecular microbial community analysis of 1. ripe fruit, 2. leaves, stems, and adventitious roots, 3. pillow components (roots, wicks, and substrates), 4. swabs, and 5. water samples from root mats before and after growth. B. transcriptomics of leaf tissue and adventitious roots, and C. elemental analysis of leaf tissue. If sufficient tissue remains elemental analyses will also be conducted on fruit. While not generating the desired information on spaceflight growth responses of healthy crops, our team is hopeful that these analyses will shed light on tomato responses to stress in this environment as plant overwatering stress is a mission-relevant condition that could occur in future space crop growth systems. This research was co-funded by the Human Research Program and Space Biology (MTL#1075) in the ILSRA 2015 NRA call.

Gioia D. Massa↗

Novel long-chain anteiso-alkanes and anteiso-alkanoic acids in Antarctic rocks colonized by living and fossil cryptoendolithic microorganisms

Saponified extracts of rock samples colonized by cryptoendolithic microbial communities from the McMurdo Dry Valleys of Southern Victoria Land, Antarctica, were separated into hydrocarbon and fatty acid fractions by silica gel column chromatography. Hydrocarbons and methyl esters of fatty acids were analyzed by capillary gas chromatography-mass spectrometry. Unusually, a suite of long-chain anteiso-alkanes (a-C20 to a-C30) and anteiso-alkanoic acids (a-C20 to a-C30) were detected in many samples, together with straight-chain, branched and/or cyclic and acyclic isoprenoid compounds. These novel compounds are probably derived from unidentified heterotrophic bacteria or symbiotic processes in a unique microbial community in the Antarctic cold desert and suggest the occurrence of a special biosynthetic pathway. Long-chain anteiso-alkanes are probably formed through microbial decarboxylation of corresponding anteiso-alkanoic acids. They may serve as new biomarkers in environmental and geochemical studies.

NASA Program Exobiology↗