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All sourcesNASA NTRSDOE OSTI
NASA NTRSDate not supplied

Spaceflight Biospecimen Sharing in Support of Science Discovery and Exploration

For decades, NASA and international partners have flown non-human biological experiments in space to understand the effects of spaceflight and address potential biological hazards. Sending organisms into space is a costly endeavor which makes space-flown biological specimens a valuable resource. To enable maximum scientific return, samples not required by the Principal Investigators are harvested and collected mostly by NASA’s Space Biology Biospecimen Sharing Program. These specimens are collected according to well-established SOPs that maintain quality and integrity. The specimens are then preserved, archived, and made available to the international scientific community through NASA’s Institutional Scientific Collection (ISC) at Ames Research Center (ARC). The ISC-ARC biospecimens and descriptive metadata are findable and accessible for request through the Life Sciences Data Archive (LSDA). The NASA ISC-ARC currently stores over 32,000 specimens from Shuttle, International Space Station, and ground-based investigations (spaceflight analog experiments involving either hindlimb unloading, centrifugation, or partial weight-bearing study designs). Tissues are predominantly from mice and rats, though samples are also available from bacteria and quail. The specimens include tissues from many physiological systems including musculoskeletal, neurosensory, reproductive, respiratory, circulatory, and digestive. Tissues are stored at -80°C, -20°C, +4°C, or ambient and preserved in various fixatives. Descriptive metadata is available for all samples. Historically, these tissues have been used for a wide range of analyses, including histology, genomics, and transcriptomics. Plans are underway to expand the ISC-ARC beyond the mostly-rodent contents, to include a space-relevant microbial culture collection including bacteria, fungi, and yeast. This expansion of the ISC-ARC will now involve identifying and standardizing best practices for microbial curations. To ensure safe long-term storage of microbial isolates, a microbiology laboratory will be dedicated for identification, cell culture, and lyophilization. Awarding of tissue to public science investigators has resulted in 33 publications since 2011, with 48 requests being submitted since 2016. Of note, NASA GeneLab has been awarded ISC-ARC biospecimens in the past few years. GeneLab processes the biospecimens to generate various levels of ‘omics’ data, which are published on GeneLab’s open access online platform for bioinformatics analysis and visualization. This has helped a systems biology community grow around the processed-biospecimens’ datasets, resulting in many new publications and insights. Websites: https://www.nasa.gov/ames/research/space-biosciences/isc-bsp ; https://lsda.jsc.nasa.gov/Biospecimen

Ryan T. Scott↗
DOE OSTI2022

Photobiological production of high-value pigments via compartmentalized co-cultures using Ca-alginate hydrogels

Abstract Engineered cyanobacterium Synechococcus elongatus can use light and CO 2 to produce sucrose, making it a promising candidate for use in co-cultures with heterotrophic workhorses. However, this process is challenged by the mutual stresses generated from the multispecies microbial culture. Here we demonstrate an ecosystem where S. elongatus is freely grown in a photo-bioreactor (PBR) containing an engineered heterotrophic workhorse (either β-carotene-producing Yarrowia lipolytica or indigoidine-producing Pseudomonas putida ) encapsulated in calcium-alginate hydrogel beads. The encapsulation prevents growth interference, allowing the cyanobacterial culture to produce high sucrose concentrations enabling the production of indigoidine and β-carotene in the heterotroph. Our experimental PBRs yielded an indigoidine titer of 7.5 g/L hydrogel and a β-carotene titer of 1.3 g/L hydrogel, amounts 15–22-fold higher than in a comparable co-culture without encapsulation. Moreover, 13 C-metabolite analysis and protein overexpression tests indicated that the hydrogel beads provided a favorable microenvironment where the cell metabolism inside the hydrogel was comparable to that in a free culture. Finally, the heterotroph-containing hydrogels were easily harvested and dissolved by EDTA for product recovery, while the cyanobacterial culture itself could be reused for the next batch of immobilized heterotrophs. This co-cultivation and hydrogel encapsulation system is a successful demonstration of bioprocess optimization under photobioreactor conditions.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS2013

Development Status of the WetLab-2 Project: New Tools for On-orbit Real-time Quantitative Gene Expression.

The primary objective of NASA Ames Research Centers WetLab-2 Project is to place on the ISS a research platform to facilitate gene expression analysis via quantitative real-time PCR (qRT-PCR) of biological specimens grown or cultured on orbit. The WetLab-2 equipment will be capable of processing multiple sample types ranging from microbial cultures to animal tissues dissected on-orbit. In addition to the logistical benefits of in-situ sample processing and analysis, conducting qRT-PCR on-orbit eliminates the confounding effects on gene expression of reentry stresses and shock acting on live cells and organisms. The system can also validate terrestrial analyses of samples returned from ISS by providing quantitative on-orbit gene expression benchmarking prior to sample return. The ability to get on orbit data will provide investigators with the opportunity to adjust experimental parameters for subsequent trials based on the real-time data analysis without need for sample return and re-flight. Finally, WetLab-2 can be used for analysis of air, surface, water, and clinical samples to monitor environmental contaminants and crew health. The verification flight of the instrument is scheduled to launch on SpaceX-5 in Aug. 2014.Progress to date: The WetLab-2 project completed a thorough study of commercially available qRT-PCR systems and performed a downselect based on both scientific and engineering requirements. The selected instrument, the Cepheid SmartCycler, has advantages including modular design (16 independent PCR modules), low power consumption, and rapid ramp times. The SmartCycler has multiplex capabilities, assaying up to four genes of interest in each of the 16 modules. The WetLab-2 team is currently working with Cepheid to modify the unit for housing within an EXPRESS rack locker on the ISS. This will enable the downlink of data to the ground and provide uplink capabilities for programming, commanding, monitoring, and instrument maintenance. The project is currently designing a module that will lyse the cells and extract RNA of sufficient quality for use in qRT-PCR reactions while using a housekeeping gene to normalize RNA concentration and integrity. Current testing focuses on two promising commercial products and chemistries that allow for RNA extraction with minimal complexity and crew time.

quantitative PCR↗
NASA NTRSDate not supplied

Enabling Space Biology Knowledge Discovery Through Biospecimen Sharing: The NASA Biological Institutional Scientific Collection

NASA and international partners have conducted experiments in space to understand the biological impacts and address hazards to health. The resulting basic and applied science is imperative to enabling humanity to venture back to the Moon and then to Mars and beyond. Sending organisms into space is a costly endeavor. All biospecimens not required by spaceflight-relevant Principal Investigators are harvested, preserved, and archived in the NASA Biological Institutional Scientific Collection (NBISC) to maximize the scientific return. The NASA Biological and Physical Sciences (BPS) Division has an ‘Open Science’ endeavor which includes NASA Genelab, the Space Biology Program’s Biospecimen Sharing Program, Physical Sciences Informatics, the Ames Life Sciences Data Archive, and NBISC. Its purpose is to integrate extensive data and biospecimen resources from spaceflight and/or ground-based analog experiments. NBISC biospecimens are collected and preserved according to well-established standard operating procedures to maintain scientific quality and are available on-request by the international scientific community. NBISC currently stores over 32,000 biospecimens from Shuttle, International Space Station, and ground-based space analog investigations. Tissue sharing has resulted in at least 33 publications since 2011 and 48 requests since 2016. Many requests for NBISC biospecimen come from first-time investigators who subsequently submit grants as the port-of-entry into the field of space biology. Some NBISC biospecimens have been awarded to NASA Genelab, who then generate various ‘Open Science’ -omics data sets on their platform for bioinformatics. Other NBISC biospecimen awards have led to multiple studies such as fecal microbiome analysis, DNA damage analysis using single-cell DNA sequencing, enzymatic-pathway identification involved in spaceflight muscle atrophy, and characterization of ocular morphological changes. Of note, NBISC has expanded to include a new Space Microbial Culture Collection (SMCC) for the collection, identification, documentation, long-term preservation, and distribution of space-related microbial isolates.

Ryan T. Scott↗
NASA NTRSDate not supplied

Enabling Biological Discovery Through Biospecimen Sharing: The Nasa Biological Institutional Scientific Collection

Understanding biological impacts from spaceflight hazards and the subsequent development of countermeasures are a high priority to enable humanity to venture back to the Moon, and then to Mars and beyond. Experiments have been conducted with model organisms flown to space and analogous investigations terrestrially, to identify biological mechanistic impacts from spaceflight hazards and to develop mitigation countermeasures, thus contributing towards basic and applied science goals. However, sending organisms into space is a costly endeavor. To maximize scientific return, all biospecimens not required by spaceflight-relevant Principal Investigators are harvested, preserved, and archived in the NASA Biological Institutional Scientific Collection (NBISC). Biospecimens are collected and preserved according to well-established standard operating procedures to maintain scientific quality and are available on-request by the international scientific community. NBISC currently stores over 32,000 biospecimens from Shuttle, International Space Station, and ground-based space analog investigations. Tissue sharing has resulted in at least 33 publications since 2011 and 51 requests since 2016. Many requests for NBISC biospecimens come from first-time investigators who subsequently submit grants as their point-of-entry into the field of spaceflight biology and health. The NBISC biorepository is part of the NASA ‘Open Science for Life in Space’ collaborative group of projects, which includes NASA Genelab, the Space Biology Program’s Biospecimen Sharing Program, Physical Sciences Informatics, and the Ames Life Sciences Data Archive. NBISC biospecimens have been awarded to NASA Genelab, who then generated various open access science ‘omics datasets through the GeneLab Sample Processing laboratory, with resulting data widely used for biological study. Other NBISC biospecimen awards have led to studies on fecal microbiome analysis, DNA damage analysis using single-cell DNA sequencing, enzymatic-pathway identification involved in spaceflight muscle atrophy, and characterization of ocular morphological changes. Of note, NBISC is expanded to include a new Space Microbial Culture Collection (SMCC) for the collection, identification, documentation, long-term preservation, and distribution of space-related microbial isolates.

Biospecimens↗
DOE OSTI2025

In anaerobic reactors the microbial community structure depends on feed type, with no “keystone” species tied to COD removal

Two-stage anaerobic digestion (AD) systems provide treatment for high strength wastewater with high stability and performance. Encapsulation technology can intensify AD to facilitate the separation of the solids retention time from the hydraulic retention time (HRT), offering lower HRTs, smaller reactors, and high effluent quality. To support successful deployment, however, the encapsulated community must contain all the needed microorganisms for successful treatment and be flexible enough to treat a variety of wastewaters. Here, a two-stage system was investigated in which microbial cultures were enriched on various high-strength wastewaters in suspended flow-through systems to determine how feed type influenced performance and microbial community structure. The hypothesis was that specific genera, or so-called “keystone species” would positively correlate to organic carbon degradation for a given feed, enabling construction of a well-functioning community for encapsulation. Results showed that the number of total bacteria (as 16S rRNA gene copies) did not correlate to soluble chemical oxygen demand (sCOD) removal, indicating that the community structure and/or members were important for good performance. Results also showed that feed type strongly influenced carbon removal and microbial community structure for 1st-stage fermenting communities, but not 2nd-stage methanogenic communities. In this study, the “core” community members were defined as organisms common to all of either the 1st- or 2nd-stage reactors irrespective of the feed they received and were present in at least 50% of the samples throughout the entire experiment. “Unique” community members were specific to a single feed, and hence, only present in either the 1st- or 2nd-stage reactors receiving that feed. In both 1st- and 2nd-stage communities, only one core genera and no unique genera were positively and significantly correlated to sCOD removal. Verification experiments performed with encapsulated communities showed that organisms identified in flow-through system and correlated with carbon degradation, though not significantly, seemed to be important for performance. Our results suggest that one cannot construct a community containing specific populations in lieu of enrichment. Nevertheless, a single diverse encapsulated anaerobic community should provide good (>80%) carbon removal when fed a variety of influents, if time is provided for enrichment after deployment.

54 ENVIRONMENTAL SCIENCES↗
NASA NTRS2003

Preliminary Characterization of a Microbial Community of Rock Varnish from Death Valley, California

Rock varnish (also referred to as desert varnish in the literature because it is particularly noticeable in desert environments) is a dark, thin (typically 50-500 m thick), layered veneer composed of clay minerals cemented together by oxides and hydroxides of manganese and iron. Some scientists suggest that varnish may provide a historical record of environmental processes such as global warming and long-term climate change. However, despite more than 30 years of study using modern microanalytical and microbial culturing techniques, the nucleation and growth mechanisms of rock varnish remain a mystery. Rock varnish is of interest to the Mars science community because a varnish-like sheen has been reported on the rocks at the Viking Lander sites. It therefore important for us to understand the formation mechanisms of terrestrial varnish abiotic, biotic, or a combination of the two -- as this understanding may give us clues concerning the chemical and physical processes occurring on the surface of Mars. It is strongly believed by some in the biogeochemistry community that microbes have a role in forming rock varnish, and iron- and manganese-oxidation by microbes isolated from varnish has been extensively investigated. Only two of these studies have investigated the microbial genetics of varnish. These studies examined the morphological, physiological and molecular characteristics of microbes that had previously been cultured from various rock varnishes and identified the cultivars using 16S rDNA sequencing techniques. However, it is well known that most of organisms existing in nature are refractory to cultivation, so many important organisms would have been missed. The currently described work investigates the genetics of rock varnish microbial community from a site in the Whipple Mtns., south of Death Valley, CA, near Parker, Arizona. We employed both cultural and molecular techniques to characterize the microorganisms found within the varnish and surrounding soil with the objectives of (a) identifying microorganisms potentially involved in varnish formation, and (b) discovering microorganisms that simply use the varnish as an extreme habitat.

Kuhlman, K. R.↗
NASA NTRS2023

Molecular Identification of Microbial Contaminants

Microorganisms can have significant impacts on the success of NASA’s missions, including the integrity of materials, the reliability of scientific results, and maintenance of crew health. Robust cleaning and sterilization protocols are currently in place in NASA facilities, but agency experts agree that microbial contamination is unavoidable and its impact on NASA’s missions and science must be minimized. Therefore, it is critical to understand: 1) what specific microorganisms are present, 2) how they may impact scientific objectives, and 3) how to select appropriate mitigation strategies. The Marshall Space Flight Center (MSFC) Planetary Protection (PP) microbiology lab historically relied solely upon enumeration of culturable microbial contamination associated with spacecraft materials or cleanrooms. However, this process is time consuming, many microbes cannot be cultured, and very few can be identified with any fidelity using NASA standard microbiological methods. The work described in this white paper includes the establishment of molecular identification capabilities at MSFC, including DNA isolation, amplification, purification, and Sanger sequencing. This capability will not only improve planetary protection efforts at MSFC (i.e. by identifying contaminating microorganisms in cleanrooms or on spacecraft) but also offers a service center-wide for the identification of contaminants that arise in other projects, processing locations, or during set up and roll out of spacecraft. This work also lays the foundation for higher throughput efforts to identify large populations of microbes across the lifetime of a project and serves as the starting point for future work into whole genome sequencing, non-culture based methods, or additional characterization studies. Ultimately, accurate identification informs appropriate mitigation strategies, increasing the chances of success for NASA’s missions and objectives.

C. D. Cassilly↗
DOE OSTI2026

Closing the loop on plastics: Biological and hybrid routes for converting plastic waste to polyhydroxyalkanoates

Polyhydroxyalkanoate (PHA) production from plastic-derived substrates offers a promising route to mitigate plastic pollution while reducing dependence on conventional PHA feedstocks. Plastic waste represents an abundant carbon source for microbial fermentation, but efficient conversion remains limited by incomplete deconstruction, inhibitory intermediates, low carbon recovery, and challenges in process integration. Plastic-derived streams contain diverse compounds, including fatty acids, hydrocarbons, fatty alcohols, aldehydes, esters, and aromatic compounds generated during depolymerization. These intermediates can be metabolized by selected microorganisms, particularly Pseudomonas species with versatile fatty-acid and hydrocarbon pathways, as well as Cupriavidus necator and mixed microbial cultures. Unlike reviews that address plastic upcycling or PHA biosynthesis separately, this review focuses on the deconstruction–fermentation interface that governs plastic-to-PHA conversion. It consolidates current progress in plastic deconstruction, substrate conditioning, microbial metabolism, fermentation control, polymer recovery, and techno-economic and life-cycle considerations. Here, by emphasizing substrate composition, biological compatibility, plastic‑carbon recovery, and final polymer quality, the review identifies priorities for scalable and environmentally sustainable PHA production from plastic-derived substrates.

42 ENGINEERING↗
DOE OSTI2022

Meta-omics-aided isolation of an elusive anaerobic arsenic-methylating soil bacterium

Soil microbiomes harbour unparalleled functional and phylogenetic diversity. However, extracting isolates with a targeted function from complex microbiomes is not straightforward, particularly if the associated phenotype does not lend itself to high-throughput screening. Here, we tackle the methylation of arsenic (As) in anoxic soils. As methylation was proposed to be catalysed by sulfate-reducing bacteria. However, to date, there are no available anaerobic isolates capable of As methylation, whether sulfate-reducing or otherwise. The isolation of such a microorganism has been thwarted by the fact that the anaerobic bacteria harbouring a functional arsenite S-adenosylmethionine methyltransferase (ArsM) tested to date did not methylate As in pure culture. Additionally, fortuitous As methylation can result from the release of non-specific methyltransferases upon lysis. Thus, we combined metagenomics, metatranscriptomics, and metaproteomics to identify the microorganisms actively methylating As in anoxic soil-derived microbial cultures. Based on the metagenome-assembled genomes of microorganisms expressing ArsM, we isolated Paraclostridium sp. strain EML, which was confirmed to actively methylate As anaerobically. This work is an example of the application of meta-omics to the isolation of elusive microorganisms.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Efficient Hydrogen Delivery for Microbial Electrosynthesis via 3D-Printed Cathodes

The efficient delivery of electrochemically in situ produced H 2 can be a key advantage of microbial electrosynthesis over traditional gas fermentation. However, the technical details of how to supply large amounts of electric current per volume in a biocompatible manner remain unresolved. Here, we explored for the first time the flexibility of complex 3D-printed custom electrodes to fine tune H 2 delivery during microbial electrosynthesis. Using a model system for H 2 -mediated electromethanogenesis comprised of 3D fabricated carbon aerogel cathodes plated with nickel-molybdenum and Methanococcus maripaludis, we showed that novel 3D-printed cathodes facilitated sustained and efficient electromethanogenesis from electricity and CO 2 at an unprecedented volumetric production rate of 2.2 L CH4 /L catholyte /day and at a coulombic efficiency of 99%. Importantly, our experiments revealed that the efficiency of this process strongly depends on the current density. At identical total current supplied, larger surface area cathodes enabled higher methane production and minimized escape of H 2 . Specifically, low current density (<1 mA/cm 2 ) enabled by high surface area cathodes was found to be critical for fast start-up times of the microbial culture, stable steady state performance, and high coulombic efficiencies. Our data demonstrate that 3D-printing of electrodes presents a promising design tool to mitigate effects of bubble formation and local pH gradients within the boundary layer and, thus, resolve key critical limitations for in situ electron delivery in microbial electrosynthesis.

03 NATURAL GAS↗
DOE OSTI2024

Alkaloids are associated with increased microbial diversity and metabolic function in poison frogs

Shifts in host-associated microbiomes can impact both host and microbes. It is of interest to understand how perturbations, like the introduction of exogenous chemicals, impact microbiomes. In poison frogs (family Dendrobatidae), the skin microbiome is exposed to alkaloids that the frogs sequester for defense. These alkaloids are antimicrobial; however, their effect on the frogs’ skin microbiome is unknown. To test this, we characterized microbial communities from field-collected dendrobatid frogs. Then, we conducted a laboratory experiment to monitor the effect of the alkaloid decahydroquinoline (DHQ) on the microbiome of two frog species with contrasting alkaloid loads in nature. In both datasets, we found that alkaloid-exposed microbiomes were more phylogenetically diverse, with an increase in diversity among rare taxa. Further, to better understand the isolate-specific response to alkaloids, we cultured microbial isolates from poison frog skin and found that many isolates exhibited enhanced growth or were not impacted by the addition of DHQ. To further explore the microbial response to alkaloids, we sequenced the metagenomes from high- and low-alkaloid frogs and observed a greater diversity of genes associated with nitrogen and carbon metabolism in high-alkaloid frogs. From these data, we hypothesized that some strains may metabolize the alkaloids. We used stable isotope tracing coupled to nanoSIMS (nanoscale secondary ion mass spectrometry), which supported the idea that some of these isolates are able to metabolize DHQ. Together, these data suggest that poison frog alkaloids open new niches for skin-associated microbes with specific adaptations, such as alkaloid metabolism, that enable survival in this environment.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRSDate not supplied

Differential Gene Expression in A Cross-Feeding Two-Species Model Microbial Community Under Simulated Microgravity and Deep-Space Radiation

A long-term goal of space biology is to understand interspecies microbial interactions in space. Presently, little is known about the combined effect of microgravity and ionizing radiation on bacterial community response when species are interdependent through exchange of metabolites in fluid medium (cross-feeding). Microgravity is expected to slow interspecies mass transfer and growth in cross-feeding communities in the low-shear, diffusion-limited environment, while ionizing radiation may influence stress response to direct (DNA damage) and indirect damage (ROS). Using a well-understood, two-species (Escherichia coli and Salmonella enterica) microbial community engineered to be a model for studying cross-feeding, we simulated galactic cosmic rays (GCRsim) and microgravity to test the hypothesis: exposure to ionizing radiation causes cell damage or stress, altering transcriptomic community responses in metabolically interdependent cells, which is exacerbated by microgravity. We expect to see differential gene expression between cross-feeding and non-cross-feeding communities. We measured GCRsim effects on growth and gene expression in well-mixed versus simulated-microgravity conditions and in cross-feeding and non-cross-feeding medium. Microbial cultures were inoculated into liquid medium in rotating wall vessels (RWV) with different rotation rates: 5 RPM (simulated microgravity) and 50 RPM (well-mixed). The E. coli-S. enterica consortium, under simulated microgravity, were exposed to 500 mGy of Simplified 5-ion Galactic Cosmic Ray Simulation for 2 hours at Brookhaven National Lab. We harvested samples 40 minutes after irradiation for extraction and sequencing (NASA GeneLab). Here we present the differential gene expression analysis results, which reveal altered transcriptomic community responses, even where growth rate differences are not observed. Gene expression of these actively metabolizing microbial communities in GCRsim may illuminate molecular mechanisms of microbial interactions in space. Understanding how microbial community gene expression, metabolism, and other cellular processes are influenced by spaceflight stressors can inform the use of microbes in human life support for low Earth orbit missions and beyond.

microgravity↗
DOE OSTI2020

The Link Between the Ecology of the Prokaryotic Rare Biosphere and Its Biotechnological Potential

Current research on the prokaryotic low abundance taxa, the prokaryotic rare biosphere, is growing, leading to a greater understanding of the mechanisms underlying organismal rarity and its relevance in ecology. From this emerging knowledge it is possible to envision innovative approaches in biotechnology applicable to several sectors. Bioremediation and bioprospecting are two of the most promising areas where such approaches could find feasible implementation, involving possible new solutions to the decontamination of polluted sites and to the discovery of novel gene variants and pathways based on the attributes of rare microbial communities. Bioremediation can be improved through the realization that diverse rare species can grow abundant and degrade different pollutants or possibly transfer useful genes. Further, most of the prokaryotic diversity found in virtually all environments belongs in the rare biosphere and remains uncultivatable, suggesting great bioprospecting potential within this vast and understudied genetic pool. This Mini Review argues that knowledge of the ecophysiology of rare prokaryotes can aid the development of future, efficient biotechnology-based processes, products and services. However, this promise may only be fulfilled through improvements in (and optimal blending of) advanced microbial culturing and physiology, metagenomics, genome annotation and editing, and synthetic biology, to name a few areas of relevance. In the future, it will be important to understand how activity profiles relate with abundance, as some rare taxa can remain rare and increase activity, whereas other taxa can grow abundant. The metabolic mechanisms behind those patterns can be useful in designing biotechnological processes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Foamed glass ceramics—an upcycled scaffold for microbial biofilm development

Glass, a near infinitely recyclable material, can be upcycled to create new products such as foamed glass ceramics, which are essentially a synthetic pumice-like material. This material has been demonstrated to sustain preserved biofilms which have application in various fields based on the deployability of the product and the preserved microbes. Foamed glass ceramics have increased surface area compared to typical soda-lime glass cullet. This material has been explored for variety of applications including the growth, storage and transport of biofilms and microbial colonies which can be preserved and deployed later. Here, we demonstrate the ability for microbial cultures including BioTiger™, Escherichia coli K-12, Bacillus thuringiensis, and two environmental eukaryotic cells to colonize the upcycled glass products, undergo preservation, and regrow after 84 days of storage. The growth of preserved samples is correlated to the time spent incubating prior to preservation. These results demonstrate the applicability of this novel glass-biofilm combination in which various preserved microorganisms are able to be rapidly grown after storage on an upcycled glass product.

36 MATERIALS SCIENCE↗
NASA NTRS2014

WetLab-2: Tools for Conducting On-Orbit Quantitative Real-Time Gene Expression Analysis on ISS

The objective of NASA Ames Research Centers WetLab-2 Project is to place on the ISS a research platform capable of conducting gene expression analysis via quantitative real-time PCR (qRT-PCR) of biological specimens sampled or cultured on orbit. The project has selected a Commercial-Off-The-Shelf (COTS) qRT-PCR system, the Cepheid SmartCycler and will fly it in its COTS configuration. The SmartCycler has a number of advantages including modular design (16 independent PCR modules), low power consumption, rapid ramp times and the ability to detect up to four separate fluorescent channels at one time enabling multiplex assays that can be used for normalization and to study multiple genes of interest in each module. The team is currently working with Cepheid to enable the downlink of data from the ISS to the ground and provide uplink capabilities for programming, commanding, monitoring, and instrument maintenance. The project has adapted commercial technology to design a module that can lyse cells and extract RNA of sufficient quality and quantity for use in qRT-PCR reactions while using a housekeeping gene to normalize RNA concentration and integrity. The WetLab-2 system is capable of processing multiple sample types ranging from microbial cultures to animal tissues dissected on-orbit. The ability to conduct qRT-PCR on-orbit eliminates the confounding effects on gene expression of reentry stresses and shock acting on live cells and organisms or the concern of RNA degradation of fixed samples. The system can be used to validate terrestrial analyses of samples returned from ISS by providing on-orbit gene expression benchmarking prior to sample return. The ability to get on orbit data will provide investigators with the opportunity to adjust experiment parameters for subsequent trials based on the real-time data analysis without need for sample return and re-flight. Researchers will also be able to sample multigenerational changes in organisms. Finally, the system can be used for analysis of air, surface, water, and clinical samples to monitor environmental contaminants and crew health. The verification flight of the instrument is scheduled to launch on SpaceX-7 in June 2015.

quantitative PCR↗
DOE OSTI2023

Waste-to-Energy Pipeline through Consolidated Fermentation–Microbial Fuel Cell (MFC) System

The rise in population, urbanization, and industrial developments have led to a substantial increase in waste generation and energy demand, posing significant challenges for waste management as well as energy conservation and production. Bioenergy conversions have been merged as advanced, sustainable, and integrated solutions for these issues, encompassing energy generation and waste upcycling of different types of organic waste. Municipal solid waste (MSW) and agricultural residues (AR) are two main resources for bioenergy conversions. Bioenergy production involves feedstock deconstruction and the conversion of platform chemicals to energy products. This review provides a detailed overview of waste sources, biofuel, and bioelectricity production from fermentation and microbial fuel cell (MFC) technology, and their economic and environmental perspectives. Fermentation plays a critical role in liquid biofuel production, while MFCs demonstrate promising potential for simultaneous production of electricity and hydrogen. Fermentation and MFCs hold a significant potential to be integrated into a single pipeline, enabling the conversion of organic matter, including a variety of waste material and effluent, into diverse forms of bioenergy via microbial cultures under mild conditions. Furthermore, MFCs are deemed a promising technology for pollutant remediation, reducing COD levels while producing bioenergy. Importantly, the consolidated fermentation–MFC system is projected to produce approximately 7.17 trillion L of bioethanol and 6.12 × 10 4 MW/m 2 of bioelectricity from MSW and AR annually, contributing over USD 465 billion to the global energy market. Such an integrated system has the potential to initiate a circular economy, foster waste reduction, and improve waste management practices. This advancement could play a crucial role in promoting sustainability across the environmental and energy sectors.

09 BIOMASS FUELS↗
DOE OSTI2021

Precipitation of low-temperature disordered dolomite induced by extracellular polymeric substances of methanogenic Archaea Methanosarcina barkeri : Implications for sedimentary dolomite formation

Abstract A correlation between methanogenesis and dolomite formation has been reported; however, the mechanism underlying this association is not fully understood. In this study, we conducted forced carbonate precipitation experiments at room temperature in calcite-seeded Ca/Mg carbonate solutions containing either purified non-living biomass or bound extracellular polymeric substances (EPS) of the methanogen Methanosarcina barkeri. Purified non-living biomass and bound EPS was used so as to avoid the possible influence of the complex components of the growing microbial culture on carbonate crystallization. Our results demonstrated that non-living biomass of M. Barkeri can enhance the Mg incorporation into calcitic structure and induce the crystallization of disordered dolomite. In the presence of ~113 mg L–1 of non-living biomass, disordered dolomite with ~41 and 45 mol% of MgCO3 was precipitated in solutions with initial Mg:Ca ratios of 5:1 and 8:1, respectively. A systematic increase in the MgCO3 contents of the precipitated Ca-Mg carbonates was also observed with the increased non-living biomass concentration. Bound EPS was shown to be the component of non-living biomass that catalyzed the precipitation of disordered dolomite. At only ~25 mg L–1 of bound EPS, disordered dolomite with ~47 and 48 mol% of MgCO3 was precipitated in solutions with initial Mg:Ca ratios of 5:1 and 8:1, respectively. We propose that adsorption of bound EPS to growing carbonate surfaces through hydrogen bonding is the key to catalyzing disordered dolomite crystallization, and that this mechanism is also applicable to natural EPS-induced dolomite formation. This study provides significant insight into the formation mechanism of microbial-induced dolomite with high δ13C values.

Geochemistry & Geophysics↗