Microbial Analysis of Chile Peppers Grown in NASA's Advanced Plant Habitat on the International Space Station
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The methodologies for studying marine microbiology typically consist of utilizing instrumentation within a laboratory. This typically requires extracting a sample from its place of origin prior to examination, which may be days after collection. Oftentimes, the solution is to bring the lab to the ocean, which may be costly and provide further limitations for a sterile and stable laboratory environment. Here we present a low-cost, submersible, digital holographic microscope (DHM) designed to image marine microorganisms (such as bacteria and plankton) in their natural underwater environment. Our instrument eliminates the need to transport samples and allows for instantaneous data collection of microbes in-situ. The DHM achieves sub-micron spatial resolution and is paired with artificial intelligence for the detection and tracking of specimens to reduce the overall collected data. This instrument also aims to reduce the cost of manufacturing and field expenses relative to marine microbiological research. “Off the shelf” components were selected in the design process of this instrument which allows us to achieve precise results without sacrificing data quality. The DHM itself costs under one thousand dollars and features a low-cost high-resolution camera, the Arducam MT9J001. Included in the design were five main subsystems: optical, mechanical, electrical, power, and machine learning. Our on board computer and artificial intelligence consist of a Raspberry Pi 4 (8 Gb) and Google Coral USB Tensorflow accelerator. Instrument testing has successfully proven our abilities of data acquisition for at least two hours in depths of at least forty meters below sea level. Additionally, our artificial intelligence system is currently capable of tracking up to ten areas of interest in a fraction of a second with over ninety percent confidence via the neural net driven by the tensor cores on the Google Coral. Furthermore, we have demonstrated the versatility of our instrument by mounting it on an ocean-going ROV, the BlueRov2 by BlueRobotics. Our tests on the BlueRov2 exemplified the cost-effective nature of a submersible and reusable instrument that can be implemented in moderate environments and on most vessels.
As the National Aeronautics and Space Association (NASA) begins to pursue long-duration space flights, they will need to be able to provide astronauts with a nutritious and reliable food source. To meet the administration’s goal of traveling to the Moon and Mars, astronauts will need to begin to grow their own food in space. To protect their food source, extensive monitoring will occur to test for the effects of a space flight environment (e.g., radiation) as well as for early pathogen and disease detection. Genomic sequencing allows for both concerns to be tested on a regular basis. However, NASA’s current sequencer is unable to process plant tissues. Therefore, a novel method for plant DNA extraction using microneedle (MN) patches that will be able to feed into NASA’s existing system, but also require minimal human input is proposed. To support this, the design was broken down into four components (1) MN patch fabrication (2) MN patch extraction, (3) automated sampling motion control, and (4) a processing module. The MN patch is fabricated using a custom mold with conically shaped needles. The mold is filled with Polyvinyl alcohol (PVA) solution and placed in a vacuum desiccator. The mold is left in the vacuum overnight until the patch is dry and ready for use. The protocol was tested with varying pressures, drying times, volume amounts, and preparation methods to determine if highquality needles can be produced. A MN is a method of DNA extraction where the patch is applied to a leaf, the needles penetrate the leaf, breaking the rigid plant cell wall to isolate the DNA. A protocol for this method of extraction was tested to ensure the patch could produce the needed yield and purity. The tests varied by the number of patches, number of applications, and plant type. To automate the MN extraction method, motion control will utilize two separate axis tables which move in the x and y directions. The y-axis table will have an end effector that fits a MN patch and will have the ability to apply the patch to the leaf sample. This end effector will also act as a lid for a downstream processing module. The other axis will position the leaf sample and processing container so that the patch can be applied accurately. The Joint Comprehensive Sequencing System (JCSS) module integrates all the components together. The output of this module feeds into the NASA Charged Information-Storage Polymer Preparation System (CHIPPS) for genomic sequencing. The extraction module operates using a series of syringes and tubing to pump the varying reagents needed for the extraction protocol. The results of the study proved that MN patches are a viable method of DNA extraction. While fabrication of high-quality needles was unsuccessful, the protocol was able to be further developed using centrifugation. The integrated design between the motion control and the JCSS enabled the potential for automation with a complete conceptual design and prototype. Future research and development for this study would include (1) further testing for fabrication (2) expanding the range of plant species compatible with the MN patch, and (3) building a working prototype for the integrated system.
The White House Office of Science and Technology Policy (OSTP) has declared 2023 to be the Year of Open Science following an initiative to democratize scientific knowledge. Simultaneously, new sensor technologies have broadened the experimental space available to bioastronautics research. With these open-science goals and technological advances in mind, we have designed and constructed a data acquisition platform for high-precision, real-time monitoring of liquid culture systems. The vessel rig is fitted with six Atlas Scientific probes (micro pH, electrical conductivity, dissolved oxygen, oxidation-reduction potential, liquid temperature, air CO2) and a custom optical density probe similar to the one on BioSentinel’s BioSensor payload. A custom dielectric spectroscopy probe is also planned. The structure of the vessel is resin 3-D printed on a hobbyist-level machine, reducing the production cost and iteration time by over 60% each while increasing extensibility. Data acquisition and storage is controlled with a standalone C state machine-based program running on a Raspberry Pi 3 Model B. When not running headless, an additional program automatically generates and updates plots for live data visualization. Validation of the rig as a data collection system was performed with a yeast liquid culture experiment. While the vessel rig is currently used for standalone experiments, it can also be used as the base perception unit in a self-driving laboratory (SDL). SDLs are high-throughput data collection systems that employ automation and artificial intelligence to conduct and manage routine experiments. Here, we envision an SDL driven by several vessel rigs in which an automated script compares key results, informing the design of future experiments. A vessel rig SDL would streamline many operations, including 1) strain selection for the Lunar Explorer Instrument for space biology Applications (LEIA) investigation and 2) the study of bioregenerative life support systems (BLSS). Ultimately, the datasets that can now be acquired will provide crucial information for accelerating bioastronautics application development in the era of commercial space.
The GERMS (Gravitational Effects and Radiobiology of Microbes in Space) lab conducts fundamental research into the responses of microorganisms to the space environment. This talk introduces concepts in space microbiology and describes some of the work ongoing in the lab.
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Explore the source record for details and available documents.
As humans move to long term stays in space their microbes move with them. This poses problems as the microbes can colonize various life support systems, creating biofilms that can grow and spread, which in turn causes obstructions and ultimately degrades function of these systems. MSFC currently has no protocol in place for the Scanning Electron Microscope (SEM) imaging of microbes.
In the search for life on Mars, keeping its surface pristine and free of microbes due to human presence is of utmost importance. Astronauts will use emptied logistics containers during their Mars stay to store waste. These containers will be left on the planet’s surface and are a potential source of microorganism contamination. It is desired for these containers to be free from leaks for a minimum of 50 years. If leaks do occur, a likely location will be at any ports. At these locations the mating surfaces will be sealed using a flexible gasket material like a thin metal or a polymer. Eventually these seals will fail. To mitigate these effects, a particulate filter can be employed to equilibrate the pressure between the Martin environment and inside the container. To meet planetary protection requirements, the filter can only allow particulates smaller than 0.2 m diameter to pass. This paper discusses why a particulate filter may be needed and how such a filter could be tested to survive and operate in the harsh Martian environment.
Carotenoids are a class of vibrant biological pigments that have a characteristic chemical structure centered around a polyene core (Lu et al. 2018). Carotenoids and their derivatives are candidate biosignatures because they can persist in the terrestrial geologic record for up to 1.73 billion years (Vinnichenko et al. 2020), have specific structures that are likely the result of complex pathways, mediate the survival of many microorganisms in Mars and Ocean Worlds analog environments, and are detectable with multiple techniques, including Raman spectroscopy. In this project, we aim to investigate the detectability of carotenoid pigments with different spectroscopic methods to inform future instrument selection. We compare the spectra of five unaltered carotenoids, two model compounds, and carotenoid-forming archaeon with visible and deep UV Raman spectroscopy and UVVis absorption spectrophotometry. We then use one model pigment, beta-carotene, to evaluate the likelihood that unique spectral properties of carotenoids, or their refractory byproducts, would be preserved and detectable on a remote planetary surface by exposing it to simulated conditions for Mars. Sample Acquisition. Pigments betacarotene, lutein, zeaxanthin, astaxanthin, and lycopene were purchased from Sigma Aldrich. Halobacterium salinarum NRC-1 was acquired from Carlina Biological and grown in Halobacterium media. Mineral salts including sodium sulfate, sodium carbonate, and halite were used to form matrices in which the beta-carotene was embedded before exposure. Pigment-mineral mixes were at a 1:10 ratio in water. Analytical Techniques. Deep UV Raman data were collected on a custom laboratory mapping spectrometer called MOBIUS (Mineral and Organic Based Investigations using Ultraviolet Spectroscopy), which is an analog to the SHERLOC instrument on the Mars 2020 Perseverance rover (Bhartia et al. 2021). It features a 248.56 nm NeCu pulsed laser, liquid nitrogen-cooled detector, and tunable optical setup. Visible Raman data were collected using a Horiba Jobin Yvon LabRam HR spectrometer with a frequencydoubled Nd:YAG laser (532 nm) and a HeNe laser (633 nm). A VWR 6300 PC UV/Visible Spectrophotometer was used to collect absorption data for carotenoid solutions, model compounds, and solvents in UVpermissible capped cuvettes. Data were collected from 190-1100 nm at 1 nm increments. All spectral data were analyzed using Igor Pro 9 (Wavemetrics). Irradiation. We used a vacuum chamber equipped with a cryostat and a flood electron gun to simulate Martian surface temperatures, low pressures, and ionizing radiation (10keV, 10μA for 6h at 200K for our initial tests). The samples were prepared by drying the pigment-mineral mix onto polished metal tabs, then mounted on the cryostat for processing. Samples were then analyzed directly on the tabs after exposure. Results: In comparing the visible and deep UV Raman spectra of unaltered pigments, we found that they differed drastically. Carotenoids are often studied with visible Raman and typically have peaks at 1525 cm-1 and 1157 cm-1, due to the stretching of the C=C and C-C bonds in the polyene structure. However, in deep UV, the strongest feature is at ~1630 cm-1 and is broad, possibly indicating that multiple peaks are forming this feature. This stark difference is likely due to different preresonant enhancement effects. The UV-Vis results show that there is an absorption band in the deep UV <300 nm, which supports the hypothesis that the 248.6 nm excitation is interrogating another aspect of carotenoids than visible Raman. Our preliminary exposure tests indicated that pigments – even without minerals present - were largely unaltered in the applied conditions, with only a slight broadening in the primary polyene peaks apparent in the visible Raman data. Figure 1. A) Visible vs. deep UV Raman spectra of unaltered beta carotene. B) Schematic of exposure. Conclusions: Our results to date indicate that deep UV and visible Raman spectroscopy, both techniques with planetary mission heritage from Mars 2020 (Wiens et al. 2021, Bhartia et al. 2021), may be used in a complementary manner to observe carotenoids. In addition, we find that beta-carotene is largely resistant to our current exposure conditions, though there may be some amount of amorphization of the material which could cause the broadening of the peaks at 1525 and 1157 cm-1. As a next step, we aim to increase the dosage and duration of exposure to observe degradation of the parent pigment, possibly add UV as a factor via an Ar mini-arc UV lamp and use GC-MS to characterize possible degradation products.
In the search for life on Mars, keeping its surface pristine and free of microbes due to human presence is of utmost importance. Astronauts will use emptied logistics containers during their Mars stay to store waste. These containers will be left on the planet’s surface and are a potential source of microorganism contamination. It is desired for these containers to be free from leaks for a minimum of 50 years. If leaks do occur, a likely location will be at any ports. At these locations the mating surfaces will be sealed using a flexible gasket material like a thin metal or a polymer. Eventually these seals will fail. To mitigate these effects, a particulate filter can be employed to equilibrate the pressure between the Martin environment and inside the container. To meet planetary protection requirements, the filter can only allow particulates smaller than 0.2 μm diameter to pass. This paper discusses why a particulate filter may be needed and how such a filter could be tested to survive and operate in the harsh Martian environment.