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NASA NTRSDate not supplied

Method Development for In-situ Detection of Latent Herpesvirus DNA from Saliva using Nanopore Sequencing

Research toward latent herpesvirus reactivation has been intensively addressed through Space Shuttle and International Space Station (ISS) investigations. This work has provided the understanding that persistent reactivation of herpesviruses from asymptomatic crew can be detected through viral shedding in saliva, urine, and blood. Occasionally, viral reactivation from the latency stage can pose a threat to crew health (clinical manifestation) before, during, and after flight missions. Furthermore, previous work detailing correlations with immunity indicate that monitoring viral reactivation could be implemented to assess potential immune dysfunction. While in-flight monitoring is desirable, there is no well-established procedure or method for real-time evaluations, and research to date has relied on postflight, ground-based analysis. The development of portable molecular technologies like the miniPCR™ (miniPCR Bio) thermal cycler and the MinION™ sequencer (Oxford Nanopore Technologies) have confirmed that real-time monitoring is possible in extreme and low resource environments. These devices, combined with simple sample preparation methods, have been used to demonstrate bacterial identification onboard the ISS, as well as rapid viral detection in remote locations on Earth. The work here builds upon previous molecular advancements onboard the ISS toward the development and validation of a spaceflight-compatible method for viral detection from crew samples. Several herpesviruses can be detected in saliva, which provides a non-invasive means to collect samples for monitoring. While the basis for this method lies in previous spaceflight investigations, key points for method optimization include DNA extraction from saliva, viral primer selection, and bioinformatic processes for data analysis. To increase viral yield, numerous DNA extraction methods have been evaluated and will be discussed in detail. For initial development and testing, varicella-zoster virus (VZV) is being targeted though open reading frame 51 and 63 (ORF51, ORF63), as the replication origin-binding protein is highly expressed during latency. Optimization of the thermal cycling parameters has resulted in the ability to test the entire process. The full method has been tested with both viral VZV DNA standards and saliva spiked with varying concentrations of VZV. Viral sequence data were mapped to the reference sequence using minimap2. Prior to mapping, DNA sequencing reads were filtered for length and quality, barcodes were removed, and alignment identity calculated. Following further assessments, statistics were compared across multiple sequencing experiments and are being used to determine the success of the protocols. Forward work will include the incorporation of herpes simplex virus 1 (HSV-1) and Epstein-Barr virus (EBV) primers as well as the validation of results to the terrestrial qPCR standard assay. Upon full validation of the developed method, saliva will be collected from 20 healthy subjects and spiked with viral DNA. These samples will be split and assayed with the MinION and standard qPCR assay.

Hang N Nguyen↗

NASA NTRS2022

How to Build a Rover: An Overview of the Mars 2020 Mission’s Vehicle System Testbed

While NASA’s Mars rover Perseverance continues to make groundbreaking achievements on the Red Planet, its twin is hard at work here on Earth. The Operational Perseverance Twin for the Integration of Mechanisms and Instruments Sent to Mars, or OPTIMISM, is the Mars 2020 Vehicle System Testbed (VSTB) rover operated by NASA Jet Propulsion Laboratory (JPL) in Pasadena, California. OPTIMISM’s home is the JPL Mars Yard; an outdoor field with red soil that simulates the terrain encountered by Perseverance. The VSTB is a full-scale engineering model of the flight rover, serving a number of functions to ensure mission operations can continue smoothly and on schedule. The VSTB possesses instrumentation, computers, mechanisms, cameras, and a Mobility subsystem that are nearly identical to its extraterrestrial twin. Its high fidelity allows the rover to be a highly effective tool to fully test system functionality and performance prior to commanding the flight rover. The early stages of building OPTIMISM began a few months prior to Perseverance departing JPL for Cape Canaveral, FL in early 2020. Electrical integration of the flight system avionics, and compatibility checkouts of the electrical ground support equipment ensured that the foundation of the electrical system was operational and in place. Next, the internal harnessing was installed and compatibility checks of the rover instrumentation and mechanisms were performed to confirm the system was prepared for full buildup. Finally, mechanical assembly of the rover chassis with its external components completed the integration of the system before it was moved to the Mars Yard for its initial phase of testing to perform verification & validation (V&V) of the Mobility subsystem requirements. By the time Perseverance landed at Jezero Crater in February 2021, the first phase of VSTB operations was underway. Surface guidance, navigation, and control (SGNC) testing for the Mobility subsystem ensured functionality and performance requirements were met for various capabilities such as visual odometry (VO), mapping, and automatic navigation (AutoNav). Subsequent integration of the robotic arm (RA) onto the VSTB enabled the V&V campaign for surface sampling operations (SSO) to commence. As the mission’s engineering operations (EO) have gotten underway, the VSTB has been utilized for an array of purposes including troubleshooting software anomalies, and performing dry-runs for first time activities (FTAs) prior to sending the commands to Perseverance. OPTIMISM will continue to serve mission critical functions as long as Perseverance is roving the Red Planet.

Rojas, Jose Trujillo↗

NASA NTRS1993

COSMIC monthly progress report

Activities of the Computer Software Management and Information Center (COSMIC) are summarized for the month of August, 1993. Tables showing the current inventory of programs available from COSMIC are presented and program processing and evaluation activities are discussed. Ten articles were prepared for publication in the NASA Tech Brief Journal. These articles (included in this report) describe the following software items: (1) MOM3D - A Method of Moments Code for Electromagnetic Scattering (UNIX Version); (2) EM-Animate - Computer Program for Displaying and Animating the Steady-State Time-Harmonic Electromagnetic Near Field and Surface-Current Solutions; (3) MOM3D - A Method of Moments Code for Electromagnetic Scattering (IBM PC Version); (4) M414 - MIL-STD-414 Variable Sampling Procedures Computer Program; (5) MEDOF - Minimum Euclidean Distance Optimal Filter; (6) CLIPS 6.0 - C Language Integrated Production System, Version 6.0 (Macintosh Version); (7) CLIPS 6.0 - C Language Integrated Production System, Version 6.0 (IBM PC Version); (8) CLIPS 6.0 - C Language Integrated Production System, Version 6.0 (UNIX Version); (9) CLIPS 6.0 - C Language Integrated Production System, Version 6.0 (DEC VAX VMS Version); and (10) TFSSRA - Thick Frequency Selective Surface with Rectangular Apertures. Activities in the areas of marketing, customer service, benefits identification, maintenance and support, and dissemination are also described along with a budget summary.

Source record↗

NASA NTRS1992

Dielectric measurement using an open ended coaxial line with an extended center conductor

Permittivity may be determined by measuring the complex reflection coefficient of an open-ended coaxial line placed in contact with a test sample. This method works best for liquid samples. For the measurement of non-liquid materials problems can arise. A perfect preparation is needed to provide a good contact between the tip and the dielectric medium. At times, the dimension of the sensor cannot be freely chosen, as is the case for the measurement of the dielectric constant of the xylem layer of a living tree. The influence of the installation of the sensor on a tree's water status must be minimized by using a small sensor. In such cases the capacitance cannot be optimized. By extending the center conductor of the open-ended coaxial line, some of these problems can be avoided. This provides an additional tool to tune the capacitance of the sensor by adjusting the length of the extension. Therefore the measurement accuracy can be optimized. The sensor also becomes sensitive to a larger volume. A comparative study of a flush and extended tipped probes shows that the ability to measure the dielectric constant of trees has been notably increased due to the extension of the center conductor.

Wegmueller, Urs↗

NASA NTRS1987

The suprastructure of the saccular macula

The ultrastructure of the rat's macular end organ was examined. Primary fixatives containing 4 or 8 percent tannic acid (with 1 hr fixation time) were found to be optimal for preserving the macular suprastructure from the macular surface to the otoconia. In agreement with observations of other workers, the suprastructure of the macular end organ showed two kinds of filamentous material of different organization: the material which supports the otoconia, the 'otoconial membrane', and the filamentous material, the 'supramacular substance'. However, in contrast to earlier reports, the fluid-filled channels around the stereociliary tufts and a slitlike space immediately above the macula were found to be artefacts of tissue preparation. The results were confirmed in decalcified samples.

Ross, Muriel D.↗

NASA NTRS2013

Transmission Electron Microscopy of Itokawa Regolith Grains

Introduction: In a remarkable engineering achievement, the JAXA space agency successfully recovered the Hayabusa space-craft in June 2010, following a non-optimal encounter and sur-face sampling mission to asteroid 25143 Itokawa. These are the first direct samples ever obtained and returned from the surface of an asteroid. The Hayabusa samples thus present a special op-portunity to directly investigate the evolution of asteroidal sur-faces, from the development of the regolith to the study of the effects of space weathering. Here we report on our preliminary TEM measurements on two Itokawa samples. Methods: We were allocated particles RA-QD02-0125 and RA-QD02-0211. Both particles were embedded in low viscosity epoxy and thin sections were prepared using ultramicrotomy. High resolution images and electron diffraction data were ob-tained using a JEOL 2500SE 200 kV field-emission scanning-transmission electron microscope. Quantitative maps and anal-yses were obtained using a Thermo thin-window energy-dispersive x-ray (EDX) spectrometer. Results: Both particles are olivine-rich (Fo70) with μm-sized inclusions of FeS and have microstructurally complex rims. Par-ticle RA-QD02-0125 is rounded and has numerous sub-μm grains attached to its surface including FeS, albite, olivine, and rare melt droplets. Solar flare tracks have not been observed, but the particle is surrounded by a continuous ~50 nm thick, stuctur-ally disordered rim that is compositionally similar to the core of the grain. One of the surface adhering grains is pyrrhotite show-ing a S-depleted rim (~8-10 nm thick) with nanophase Fe metal grains (<5 nm) decorating the outermost surface. The pyrrhotite displays a complex superstructure in its core that is absent in the S-depleted rim. Particle RA-QD02-0211 contains solar flare particle tracks (~2x109 cm-2) and shows a structurally disordered rim ~100 nm thick. The track density corresponds to a surface exposure of ~103-104 years based on the track production rate of [1]. The dis-ordered rim is nanocrystalline with minor amorphous material between crystalline domains. Quantitative element maps show the outermost ~10 nm of the disordered rim is Si-rich. Discussion and Conclusions: Both particles record the ef-fects of space weathering processes on Itokawa. Noguchi et al. [2] proposed that the disordered rims they observed on Itokawa particles largely result from solar wind radiation damage and we arrive at a similar conclusion for the two particles we analyzed. The microstructure of the S-depleted layer on the pyrrhotite grain in RA-QD02-0125 is similar to that observed in troilite irradiated with ~1018 4 kV He+ [3, 4]. Prolonged irradiation has also been shown to disorder pyrrhotite such that the superstructure reflec-tions are lost [5].

Keller, Lindsay P.↗

NASA NTRS1978

Spatial land-use inventory, modeling, and projection/Denver metropolitan area, with inputs from existing maps, airphotos, and LANDSAT imagery

A landscape model was constructed with 34 land-use, physiographic, socioeconomic, and transportation maps. A simple Markov land-use trend model was constructed from observed rates of change and nonchange from photointerpreted 1963 and 1970 airphotos. Seven multivariate land-use projection models predicting 1970 spatial land-use changes achieved accuracies from 42 to 57 percent. A final modeling strategy was designed, which combines both Markov trend and multivariate spatial projection processes. Landsat-1 image preprocessing included geometric rectification/resampling, spectral-band, and band/insolation ratioing operations. A new, systematic grid-sampled point training-set approach proved to be useful when tested on the four orginal MSS bands, ten image bands and ratios, and all 48 image and map variables (less land use). Ten variable accuracy was raised over 15 percentage points from 38.4 to 53.9 percent, with the use of the 31 ancillary variables. A land-use classification map was produced with an optimal ten-channel subset of four image bands and six ancillary map variables. Point-by-point verification of 331,776 points against a 1972/1973 U.S. Geological Survey (UGSG) land-use map prepared with airphotos and the same classification scheme showed average first-, second-, and third-order accuracies of 76.3, 58.4, and 33.0 percent, respectively.

Tom, C.↗

NASA NTRS1995

Carbon isotopic analysis of atmospheric methane by isotope-ratio-monitoring gas chromatography-mass spectrometry

Less than 15 min are required for the determination of delta C(sub PDB)-13 with a precision of 0.2 ppt(1 sigma, single measurement) in 5-mL samples of air containing CH4 at natural levels (1.7 ppm). An analytical system including a sample-introduction unit incorporating a preparative gas chromatograph (GC) column for separation of CH4 from N2, O2, and Ar is described. The 15-min procedure includes time for operation of that system, high-resolution chromatographic separation of the CH4, on-line combustion and purification of the products, and isotopic calibration. Analyses of standards demonstrate that systematic errors are absent and that there is no dependence of observed values of delta on sample size. For samples containing 100 ppm or more CH4, preconcentration is not required and the analysis time is less than 5 min. The system utilizes a commercially available, high-sensitivity isotope-ratio mass spectrometer. For optimal conditions of smaple handling and combustion, performance of the system is within a factor of 2 of the shot-noise limit. The potential exists therefore for analysis of samples as small as 15 pmol CH4 with a standard deviation of less than 1 ppt.

Merritt, Dawn A.↗

NASA NTRS2004

Characterization of Two Microbial Isolates from Andean Lakes in Bolivia

We are currently investigating the biological population present in the highest and least explored perennial lakes on earth in the Bolivian and Chilean Andes, including several volcanic crater lakes of more than 6000 m elevation, in combination of microbiological and molecular biological methods. Our samples were collected in saline lakes of the Laguna Blanca Laguna Verde area in the Bolivian Altiplano and in the Licancabur volcano crater (27 deg. 47 min S/67 deg. 47 min. W) in the ongoing project studying high altitude lakes. The main goal of the project is to look for analogies with Martian paleolakes. These Bolivian lakes can be described as Andean lakes following the classification of Chong. We have attempted to isolate pure cultures and phylogenetically characterize prokaryotes that grew under laboratory conditions. Sediment samples taken from the Licancabur crater lake (LC), Laguna Verde (LV), and Laguna Blanca (LB) were analyzed and cultured using enriched liquid media under both aerobic and anaerobic conditions. All cultures were incubated at room temperature (15 to 20 C) and under light exposure. For the reported isolates, 36 hours incubation were necessary for reaching optimal optical densities to consider them viable cultures. Ten serial dilutions starting from 1% inoculum were required to obtain a suitable enriched cell culture to transfer into solid media. Cultures on solid medium were necessary to verify the formation of colonies in order to isolate pure cultures. Different solid media were prepared using several combinations of both trace minerals and carbohydrates sources in order to fit their nutrient requirements. The microorganisms formed individual colonies on solid media enriched with tryptone, yeast extract and sodium chloride. Cells morphology was studied by optical and electronic microscopy. Rodshape morphologies were observed in most cases. Total bacterial genomic DNA was isolated from 50 ml late-exponential phase culture by using the CTAB miniprep protocol. The 16S rRNA genes were amplified by PCR using both Bacteria- and Archaeauniversal primer sets: 27f and 1492r, 21f and 1492r respectively. Sequences of 16S rRNA gene were determined and initially compared with reference sequences contained in the EMBL nucleotide sequence database by using the BLAST program and were subsequently aligned with 16S rRNA reference sequences in the ARB package (http://www.mikro.biologie.tu-muenchen.de). Aligned sequences were inserted within a stable phylogenetic tree by using the ARB parsimony tool. In this work we report the morphology and phylogenetic characterization of two isolates belonged to Laguna Blanca sediments.

Demergasso, C.↗

NASA NTRS2019

Differences Between OCO‐2 and GOME‐2 SIF Products From a Model‐Data Fusion Perspective

Space-borne retrievals of solar-induced chlorophyll fluorescence (SIF) over land surfaces have recently become a resource for studying and quantifying the broad scale dynamics of gross carbon uptake (gross primary productivity—GPP) across ecosystems. To prepare for the assimilation of SIF data in terrestrial biosphere models, we examine how differences between SIF products (due to differences in acquisition characteristics and processing chain) may affect the optimization of model parameters and the resultant GPP estimate. We compare recent daily mean SIF products (one from the Orbiting Carbon Observatory-2 [OCO-2] and two from the Global Ozone Monitoring Experiment–2 [GOME-2], GlobFluo [GF] and NASA-v28 [N28], missions), averaged at 0.5° × 0.5° spatial resolution and 16-day temporal resolution, at the biome level. Phase differences between these products are relatively small. A first-order correction of the difference in spectral sampling between the two instruments shows that OCO-2 and N28 are consistent in terms of magnitude and amplitude, while GF is twice as large as the others. Using a bias-blind toy data assimilation framework, we analyze how biases between SIF products, and between model and products, can be partially alleviated by optimizing the slope and intercept parameters of a linear GPP-SIF operator. As observation biases can transfer to biases in other optimized process-based parameters and to modeled carbon fluxes— thereby resulting in unidentified inaccurate parameter values—we argue that potential SIF biases should be treated cautiously in real-world experiments in order to achieve realistic and reliable future simulations.

Gross primary production↗

NASA NTRS2000

Percussive Force Magnitude in Permafrost

An in-depth look at percussive drilling shows that the transmission efficiency is very important; however, data for percussive drilling in hard rock or permafrost is rarely available or the existing data are very old. Transmission efficiency can be used as a measurement of the transmission of the energy in the piston to the drill steel or bit and from the bit to the rock. Having a plane and centralized impact of the piston on the drill steel can optimize the transmission efficiency from the piston to the drill steel. A transmission efficiency of near 100% between piston and drill steel is possible. The transmission efficiency between bit and rock is dependent upon the interaction within the entire system. The main factors influencing this transmission efficiency are the contact area between cutting structure and surrounding rock (energy loss due to friction heat), damping characteristics of the surrounding rock (energy dampening), and cuttings transport. Some of these parameters are not controllable. To solve the existing void regarding available drilling data, an experiment for gathering energy data in permafrost for percussive drilling was designed. Fifteen artificial permafrost samples were prepared. The samples differed in the grain size distribution to observe a possible influence of the grain size distribution on the drilling performance. The samples were then manually penetrated (with a sledge-hammer) with two different spikes.

Eustes, A. W., III↗

NASA NTRS2005

An Improved Thermal Conductivity Polyurethane Composite for a Space Borne 20KV Power Supply

This effort was designed to find a way to reduce the temperature rise of critical components of a 20KV High Voltage Power Supply (HVPS) by improving the overall thermal conductivity of the encapsulated modules. Three strategies were evaluated by developing complete procedures, preparing samples, and performing tests. The three strategies were: 1. Improve the thermal conductivity of the polyurethane encapsulant through the addition of thermally conductive powder while minimizing impact on other characteristics of the encapsulant. 2. Improve the thermal conductivity of the polyurethane encapsulated assembly by the addition of a slab of thermally conductive, electrically insulating material, which is to act as a heat spreader. 3. Employ a more thermally conductive substrate (Al203) with the existing encapsulation scheme. The materials were chosen based on the following criteria: high dielectric breakdown strength; high thermal conductivity, ease of manufacturing, high compliance, and other standard space qualified materials properties (low out-gassing, etc.). An optimized cure was determined by a statistical design of experiments for both filled and unfilled materials. The materials were characterized for the desired properties and a complete process was developed and tested. The thermal performance was substantially improved and the strategies may be used for space flight.

Pb free↗

NASA NTRS2024

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

There are numerous applications for microalgae in spaceflight missions and on Earth, such as oxygen production, carbon dioxide removal, nutrition, wastewater processing, and biofuel production. Space Algae-2 aims to test the genetic stability of Arthrospira platensis, commonly known as spirulina, during six-months of continuous culture on the International Space Station. Long-duration exposure to ionizing radiation and microgravity may impact growth, nutrient composition, and genetic stability. The high protein, vitamin, antioxidant content, and radiation resistance make spirulina a promising candidate for bioregenerative life support systems. A concept of operations was developed to grow and harvest algal biomass in space. Preflight testing experiments optimized conditions for an extended growth period in a gas permeable bioreactor bag. Preflight and post-harvest storage methods were developed in addition to a novel cryopreservation method. After sample return, multi-omics analysis will be conducted to determine the mutation rate, gene expression, and the protein and metabolite profile. The concept of operations for Space Algae-2 was tested during a lunar mission simulation within a semi-controlled environment. During a six-day lunar analog mission at the Hawai’i Space Exploration Analog and Simulation (HI-SEAS) A. platensis was successfully grown using flight-like hardware. The cyanobacteria were harvested and used to supplement bread as an example of spirulina biomass utilization. Overall, the data collected from Space Algae-2 will inform potential bioengineering of spirulina for space and terrestrial applications.

Algae↗

NASA NTRSDate not supplied

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

Algae↗

NASA NTRS2000

Over-Expression, Purification and Crystallization of Human Dihydrolipoamide Dehydrogenase

Dehydrolipoamide dehydrogenase (E3; dihydrolipoan-tide:NAD+ oxidoreductase, EC 1.8.1.4) is a common catalytic component found in pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex, and branched-chain cc-keto acid dehydrogenase complex. E3 is also a component (referred to as L protein) of the glycine cleavage system in bacterial metabolism (2). Active E3 forms a homodimer with four distinctive subdomain structures (FAD binding, NAD+ binding, central and interface domains) with non-covalently but tightly bound FAD in the holoenzyme. Deduced amino acids from cloned full-length human E3 gene showed a total of 509 amino acids with a leader sequence (N-terminal 35 amino acids) that is excised (mature form) during transportation of expressed E3 into mitochondria membrane. So far, three-dimensional structure of human E3 has not been reported. Our effort to achieve the elucidation of the X-ray crystal structure of human E3 will be presented. Recombinant pPROEX-1 expression vector (from GIBCO BRL Life Technologies) having the human E3 gene without leader sequence was constructed by Polymerase Chain Reaction (PCR) and subsequent ligation, and cloned in E.coli XL1-Blue by transformation. Since pPROEX-1 vector has an internal His-tag (six histidine peptide) located at the upstream region of a multicloning site, one-step affinity purification of E3 using nickelnitriloacetic acid (Ni-NTA) agarose resin, which has a strong affinity to His-tag, was feasible. Also a seven-amino-acid spacer peptide and a recombinant tobacco etch virus protease recognition site (seven amino acids peptide) found between His-tag and first amino acid of expressed E3 facilitated the cleavage of His-tag from E3 after the affinity purification. By IPTG induction, ca. 15 mg of human E3 (mature form) was obtained from 1L LB culture with overnight incubation at 25C. Over 98% of purity of E3 from one-step Ni-NTA agarose affinity purification was confirmed by SDS-PAGE analysis. For crystallization, E3 samples were prepared with and without His-tag. To minimize the aggregation of E3, apo- and holo- forms of E3s were tested, as well as a mutated E3. Dynamic light scattering measurements revealed that the E3 preparations without His-tag and substrate are highly monodispersive with regard to homodimers. Consequent crystallization trials of this E3 preparation led to single crystals of E3 grown by the vapor diffusion method. Crystals were obtained within a few days from solution containing poly (ethylene glycol) monomethyl ether 5000 as a precipitant. Autoindexing and integration of the X-ray diffraction data showed that E3 crystals belong to an orthorhombic system with unit cell parameters a-- 123. 1, b= 165.3 and c=214.3A. Further optimization of protein preparation and crystallization experiments for the structural determination will be discussed.

Hong, Y. S.↗

NASA NTRSDate not supplied

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

There are numerous applications for microalgae in spaceflight missions and on Earth, such as, oxygen production, carbon dioxide removal, nutrition, wastewater processing, and biofuel production. Space Algae-2 aims to test the genetic stability of Arthrospira platensis, commonly known as spirulina, during six-months of continuous culture in spaceflight on the International Space Station. Long-duration exposure to ionizing radiation and microgravity may impact growth, nutrient composition, and genetic stability. The high protein, vitamin, antioxidant content, and radiation resistance make spirulina a promising candidate for bioregenerative life support systems during long-duration missions. A concept of operations was developed to grow and harvest algal biomass in space. Preflight testing experiments were conducted to optimize conditions for an extended growth period in a gas permeable bioreactor bag. Preflight and post-harvest storage methods were developed in addition to a novel cryopreservation method. After sample return, multi-omics analysis will be conducted to determine the mutation rate, gene expression, and protein and metabolite profile. The concept of operations for Space Algae-2 was tested at HI-SEAS (Hawai’i Space Exploration Analog and Simulation) during a six-day lunar analog mission (EMMIHS23, EuroMoonMars, International MoonBase Alliance, HI-SEAS, 2023). A. platensis was grown in the semi-controlled environment using flight-like hardware and solar powered LED lights. Then, the biomass was harvested and used to supplement bread as an example of A. platensis utilization. Overall, the data collected from Space Algae-2 will inform potential bioengineering of spirulina for space and terrestrial applications.

Algae↗

NASA NTRS2004

Phosphate binding protein as the biorecognition element in a biosensor for phosphate

This work explores the potential use of a member of the periplasmic family of binding proteins, the phosphate binding protein (PBP), as the biorecognition element in a sensing scheme for the detection of inorganic phosphate (Pi). The selectivity of this protein originates from its natural role which, in Escherichia coli, is to serve as the initial receptor for the highly specific translocation of Pi to the cytoplasm. The single polypeptide chain of PBP is folded into two similar domains connected by three short peptide linkages that serve as a hinge. The Pi binding site is located deep within the cleft between the two domains. In the presence of the ligand, the two globular domains engulf the former in a hinge-like manner. The resultant conformational change constitutes the basis of the sensor development. A mutant of PBP (MPBP), where an alanine was replaced by a cysteine residue, was prepared by site-directed mutagenesis using the polymerase chain reaction (PCR). The mutant was expressed, from plasmid pSD501, in the periplasmic space of E. coli and purified in a single chromatographic step on a perfusion anion-exchange column. Site-specific labeling was achieved by attaching the fluorophore, N-[2-(1-maleimidyl)ethyl]-7-(diethylamino)coumarin-3-carboxamide (MDCC), to the protein through the sulfhydryl group of the cysteine moiety. Steady-state fluorescence studies of the MPBP-MDCC conjugate showed a change in the intensity of the signal upon addition of Pi. Calibration curves for Pi were constructed by relating the intensity of the fluorescence signal with the amount of analyte present in the sample. The sensing system was first developed and optimized on a spectrofluorometer using ml volumes of sample. It was then adapted to be used on a microtiter plate arrangement with microliter sample volumes. The system's versatility was finally proven by developing a fiber optic fluorescence-based sensor for monitoring Pi. In all three cases the detection limits for the analyte were in the sub-microMolar range. It was also demonstrated that the sensing system was selective for phosphate over other structurally-similar anions, paving the way for the design and development of a new family of biosensors utilizing the specific binding properties of periplasmic proteins. c2003 Elsevier B.V. All rights reserved.

NASA Program Fundamental Space Biology↗

NASA NTRSDate not supplied

Metabolic Vessel for Impedance Spectroscopy and Electrochemistry (MVISE): The Ground Mapping Unit for the Lunar Explorer Instrument for Space Biology Applications (LEIA)

The BioSensor payload on the upcoming LEIA platform aboard a CLPS lander will carry yeast to the moon to study response to radiation and lunar gravity. The LEIA BioSensor is designed to monitor metabolic activity using absorbance in conjunction with alamarBlue for measuring colorimetric changes as proxy measurement for redox potential. The science data returned from small spacecraft mission modules like LEIA is limited as it relies solely on optical measurements, necessitating a corresponding ground mapping unit that is equipped with multiple electrochemical sensors for accurate mapping of the optical data and operates fully automatically. This technology development work discusses the extensive design and optimization efforts put into the ground mapping unit, MVISE. MVISE is a custom designed, 3D-printed vessel with an agitation system, with six different electrochemical sensor probes, and ports for sample collection and a pressure release valve. The sensors provide real-time data, with dry absorbance measurements aligning with LEIA flight hardware and wet measurements demonstrating invasive sensor design enabling a comparison between the two setups. The 3D printer resin was tested for mechanical robustness, biocompatibility, and resistance to autoclave sterilization. The inner walls were coated with food-grade epoxy, ensuring a smooth finish to prevent microbial lodging and dye staining. The MVISE has successfully passed a week-long leak test and is now undergoing active biology tests. The MVISE prototype will be prepared for radiation tests, with three identical units being tested for varying radiation levels and culture compositions at the NASA Space Radiation Laboratory in November 2024. The integrated sensor approach proposed in this work will enable the accurate mapping of the BioSentinel/LEIA optical flight data to six sensor parameters on the ground unit for better science data return and will enable the first effort to evaluate classical biochemical sensor measurements by comparing and contrasting their responses.

Chinmayee Govinda Raj↗