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Deep UV Native Fluorescence Imaging of Antarctic Cryptoendolithic Communities
An interdisciplinary team at the Jet Propulsion Laboratory Center for Life Detection has embarked on a project to provide in situ chemical and morphological characterization of Antarctic cryptoendolithic microbial communities. We present here in situ deep ultraviolet (UV) native fluorescence and environmental scanning electron microscopy images transiting 8.5 mm into a sandstone sample from the Antarctic Dry Valleys. The deep ultraviolet imaging system employs 224.3, 248.6, and 325 nm lasers to elicit differential fluorescence and resonance Raman responses from biomolecules and minerals. The 224.3 and 248.6 nm lasers elicit a fluorescence response from the aromatic amino and nucleic acids. Excitation at 325 nm may elicit activity from a variety of biomolecules, but is more likely to elicit mineral fluorescence. The resultant fluorescence images provide in situ chemical and morphological maps of microorganisms and the associated organic matrix. Visible broadband reflectance images provide orientation against the mineral background. Environmental scanning electron micrographs provided detailed morphological information. The technique has made possible the construction of detailed fluorescent maps extending from the surface of an Antarctic sandstone sample to a depth of 8.5 mm. The images detect no evidence of microbial life in the superficial 0.2 mm crustal layer. The black lichen component between 0.3 and 0.5 mm deep absorbs all wavelengths of both laser and broadband illumination. Filamentous deep ultraviolet native fluorescent activity dominates in the white layer between 0.6 mm and 5.0 mm from the surface. These filamentous forms are fungi that continue into the red (iron-rich) region of the sample extending from 5.0 to 8.5 mm. Using differential image subtraction techniques it is possible to identify fungal nuclei. The ultraviolet response is markedly attenuated in this region, apparently from the absorption of ultraviolet light by iron-rich particles coating the filaments. Below 8.5 mm the filamentous morphology of the upper layers gives way to punctate 1-2 micron particles evidencing fluorescent activity following excitation at both deep ultraviolet wavelengths.
Determination of Aromatic Ring Number Using Multi-Channel Deep UV Native Fluorescence
The in situ detection of organic material on an extraterrestrial surface requires both effective means of searching a relatively large surface area or volume for possible organic carbon, and a more specific means of identifying and quantifying compounds in indicated samples. Fluorescence spectroscopy fits the first requirement well, as it can be carried out rapidly, with minimal or no physical contact with the sample, and with sensitivity unmatched by any other organic analytical technique. Aromatic organic compounds with know fluorescence signatures have been identified in several extraterrestrial samples, including carbonaceous chondrites, interplanetary dust particles, and Martian meteorites. The compound distributions vary among these sources, however, with clear differences in relative abundances by number of aromatic rings and by degree of alkylation. This relative abundance information, therefore, can be used to infer the source of organic material detected on a planetary surface.
Determining Biosignatures by Complexity Analysis in Antarctic Cryptoendolithic Communities
One of the most difficult problems of life detection is that of identifying biosignatures across a wide range of scales using multiple co-registered probes. The technique should be of equal utility across a wide range of search spaces from remote sensors probing volumes of space or planetary surfaces, visual eye or camera searches across the surface of a rock in Antarctica, low resolution microscopic scanning of a rock or a space craft in situ, or high resolution electron microscope and computerized tomography scanning of geobiological samples. We describe here an approach to this problem which derives in large part from past work done in the area of astrophysics - namely the analysis of complexity in galactic signals by data compression methods. This approach is a radically new one for geobiology and astrobiology, and allows us to assess the complexity (and thus potential biogenicity) of an object being examined. This is done by considering the information within pixels of an image (regardless the sensor used to gather the information) as an energetic system capable of description in terms of classical thermodynamics. The image data space is searched by an algorithm that judges complexity via data compression (e.g., the more compressible it is, the less complex, and vice versa) and maximum entropy as originally outlined by Shannon. At present we are implementing methods to utilize images from multiple sensors gathering different kinds of information (e.g., visible gray-scale data, color analyses, UV fluorescence, chemical information, etc). We present here preliminary data from deep UV fluorescence and ESEM (Environmental Scanning Electron Microscope) images from a layered cryptoendolithic community of an Antarctic rock.
Advancing Launch Readiness of the Europa Light Microscope
The search for life in our solar system relies on our ability to measure key biomarkers in situ and identify them as likely of biotic origin. One class of biomarkers that would be indicative of life is cell-like features such as the colocalization of key biochemicals, spatial organization of native UV fluorescent compounds, and polarization across membrane compartments. The Europa Lander Science Definition Team has emphasized the need for a light microscope to search for these features on Europa, and the Europa Light Microscope (ELM) has been developed to address this need. This work presents the development towards high launch readiness of ELM by demonstrating its optical resolution under brightfield illumination, deep UV fluorescence imaging of unstained living samples, and multicolor imaging of samples stained with exogenous fluorophores targeting proteins, lipids, and nucleic acids. This work also seeks to extend the capabilities of ELM by incorporating fluorescence-based assays for membrane potential as a key biomarker for living cellular material.
The first 300 sols of the SHERLOC investigation on the Mars 2020 rover
The Mars2020 mission is NASA’s latest flagship mission to Mars. The spacecraft launched in July 2020, and landed in Jezero crater on February 18, 2021 at the Octavia Butler Landing site. The rocks and sediments of Jezero crater have been argued to have preserved records of past, potentially habitable environments. The mission is characterizing the field site by analyzing chemistry, looking for organics and searching for potential biosignatures. The Scanning Habitable Environments with Ra-man and Luminescence for Organics and Chemicals (SHERLOC) is a robotic arm mounted instrument. SHERLOC combines imaging with UV resonance Raman and native deep UV fluorescence spectroscopy to identify potential biosignatures and understand the aqueous history of the Jezero Region. WATSON (Wide Angle Topographic Sensor for Operations and eNgineering), a refight of the Mars Hand Lens Imager (MAHLI) on the Curiosity rover is capable of color imaging over a wide range of resolutions (from infinity to 13.1 micron/pixel) and is used for both science and engineering. A second imager, the Autofocusing Con-textual Imager (ACI), produces gray scale images at 10.1 micron/pixel resolution at a 48 mm range.
Lunar and Planetary Science XXXV: Astrobiology
The presentations in this session are: 1. A Prototype Life Detection Chip 2. The Geology of Atlantis Basin, Mars, and Its Astrobiological Interest 3. Collecting Bacteria Together with Aerosols in the Martian Atmosphere by the FOELDIX Experimental Instrument Developed with a Nutrient Detector Pattern: Model Measurements of Effectivity 4. 2D and 3D X-ray Imaging of Microorganisms in Meteorites Using Complexity Analysis to Distinguish Field Images of Stromatoloids from Surrounding Rock Matrix in 3.45 Ga Strelley Pool Chert, Western Australia 4. Characterization of Two Isolates from Andean Lakes in Bolivia Short Time Scale Evolution of Microbiolites in Rapidly Receding Altiplanic Lakes: Learning How to Recognize Changing Signatures of Life 5. The Effect of Salts on Electrospray Ionization of Amino Acids in the Negative Mode 6. Determination of Aromatic Ring Number Using Multi-Channel Deep UV Native Fluorescence 7. Microbial D/H Fractionation in Extraterrestrial Materials: Application to Micrometeorites and Mars 8. Carbon Isotope Characteristics of Spring-fed Iron-precipitating Microbial Mats 9. Amino Acid Survival Under Ambient Martian Surface UV Lighting Extraction of Organic Molecules from Terrestrial Material: Quantitative Yields from Heat and Water Extractions 10. Laboratory Detection and Analysis of Organic Compounds in Rocks Using HPLC and XRD Methods 11. Thermal Decomposition of Siderite-Pyrite Assemblages: Implications for Sulfide Mineralogy in Martian Meteorite ALH84001 Carbonate Globules 12. Determination of the Three-Dimensional Morphology of ALH84001 and Biogenic MV-1 Magnetite: Comparison of Results from Electron Tomography and Classical Transmission Electron Microscopy 13. On the Possibility of a Crypto-Biotic Crust on Mars Based on Northern and Southern Ringed Polar Dune Spots 14. Comparative Planetology of the Terrestrial Inner Planets: Implications for Astrobiology 15. A Possible Europa Exobiology 16. A Possible Biogeochemical Model for Titan
Deep UV Penetration Depth in Astrobiologically Relevant Rocks & Minerals
No abstract available
Mars 2020 Rover SHERLOC Calibration Target
The Scanning Habitable Environments with Raman & Luminescence for Organics & Chemicals (SHERLOC) instrument is a deep ultraviolet (UV) Raman Fluorescence instrument selected as part of the Mars 2020 rover instrument suite. SHERLOC will be mounted on the rover arm and its primary role is to identify carbonaceous species in martian samples. The SHERLOC instrument requires a calibration target which is being designed and fabricated at JSC as part of our continued science participation in Mars robotic missions. The SHERLOC calibration target will address a wide range of NASA goals to include basic science of interest to both the Science Mission Directorate and Human Exploration and Operations Mission Directorate.
Biochemical Detection and Identification False Alarm Rate Dependence on Wavelength Using Laser Induced Fluorescence
Most organic and many inorganic materials absorb strongly in specific wavelength ranges in the deep UV between about 220nm and 300nm. Excitation within these absorption bands results in native fluorescence emission. Each compound or composite material, such as a bacterial spore, has a unique excitation-emission fingerprint that can be used to provide information about the material. The sensitivity and specificity with which these materials can be detected and identified depends on the excitation wavelength and the number and location of observation wavelengths.We will present data on our deep ultraviolet Targeted Ultraviolet Chemical Sensors that demonstrate the sensitivity and specificity of the sensors. In particular, we will demonstrate the ability to quantitatively differentiate a wide range of biochemical agent targets against a wide range of background materials. We will describe the relationship between spectral resolution and specificity in target identification, as well as simple, fast, algorithms to identify materials.Hand-held, battery operated instruments using a deep UV laser and multi-band detection have been developed and deployed on missions to the Antarctic, the Arctic, and the deep ocean with the capability of detecting a single bacterial spore and to differentiate a wide range of organic and biological compounds.
Perseverance’s Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals (SHERLOC) Investigation
The Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals (SHERLOC) is a robotic arm-mounted instrument on NASA’s Perseverance rover. SHERLOC has two primary boresights. The Spectroscopy boresight generates spatially resolved chemical maps using fluorescence and Raman spectroscopy coupled to microscopic images (10.1 μm/pixel). The second boresight is a Wide Angle Topographic Sensor for Operations and eNgineering (WATSON); a copy of the Mars Science Laboratory (MSL) Mars Hand Lens Imager (MAHLI) that obtains color images from microscopic scales (~13 μm/pixel) to infinity. SHERLOC Spectroscopy focuses a 40 μs pulsed deep UV neon-copper laser (248.6 nm), to a ~100 μm spot on a target at a working distance of ~48 mm. Fluorescence emissions from organics, and Raman scattered photons from organics and minerals, are spectrally resolved with a single diffractive grating spectrograph with a spectral range of 250 to ~370 nm. Because the fluorescence and Raman regions are naturally separated with deep UV excitation (<250 nm), the Raman region ~ 800 – 4000 cm -1 (250 to 273 nm) and the fluorescence region (274 to ~370 nm) are acquired simultaneously without time gating or additional mechanisms. SHERLOC science begins by using an Autofocus Context Imager (ACI) to obtain target focus and acquire 10.1 μm/pixel greyscale images. Chemical maps of organic and mineral signatures are acquired by the orchestration of an internal scanning mirror that moves the focused laser spot across discrete points on the target surface where spectra are captured on the spectrometer detector. ACI images and chemical maps (< 100 μm/mapping pixel) will enable the first Mars in situ view of the spatial distribution and interaction between organics, minerals, and chemicals important to the assessment of potential biogenicity (containing CHNOPS). Single robotic arm placement chemical maps can cover areas up to 7x7 mm in area and, with the < 10 min acquisition time per map, larger mosaics are possible with arm movements. This microscopic view of the organic geochemistry of a target at the Perseverance field site, when combined with the other instruments, such as Mastcam-Z, PIXL, and SuperCam, will enable unprecedented analysis of geological materials for both scientific research and determination of which samples to collect and cache for Mars sample return.
Perseverance’s Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals (SHERLOC) Investigation
The Scanning Habitable Environments with Raman and Luminescence for Organics and Chemicals (SHERLOC) is a robotic arm-mounted instrument on NASA’s Perseverance rover. SHERLOC has two primary boresights. The Spectroscopy boresight generates spatially resolved chemical maps using fluorescence and Raman spectroscopy coupled to microscopic images (10.1 μm/pixel). The second boresight is a Wide Angle Topographic Sensor for Operations and eNgineering (WATSON); a copy of the Mars Science Laboratory (MSL) Mars Hand Lens Imager (MAHLI) that obtains color images from microscopic scales (∼13 μm/pixel) to infinity. SHERLOC Spectroscopy focuses a 40 μs pulsed deep UV neon-copper laser (248.6 nm), to a ∼100 μm spot on a target at a working distance of ∼48 mm. Fluorescence emissions from organics, and Raman scattered photons from organics and minerals, are spectrally resolved with a single diffractive grating spectrograph with a spectral range of 250 to ∼370 nm. Because the fluorescence and Raman regions are naturally separated with deep UV excitation (<250 nm), the Raman region ∼ 800 – 4000 cm−1 (250 to 273 nm) and the fluorescence region (274 to ∼370 nm) are acquired simultaneously without time gating or additional mechanisms. SHERLOC science begins by using an Autofocus Context Imager (ACI) to obtain target focus and acquire 10.1 μm/pixel greyscale images. Chemical maps of organic and mineral signatures are acquired by the orchestration of an internal scanning mirror that moves the focused laser spot across discrete points on the target surface where spectra are captured on the spectrometer detector. ACI images and chemical maps (< 100 μm/mapping pixel) will enable the first Mars in situ view of the spatial distribution and interaction between organics, minerals, and chemicals important to the assessment of potential biogenicity (containing CHNOPS). Single robotic arm placement chemical maps can cover areas up to 7x7 mm in area and, with the < 10 min acquisition time per map, larger mosaics are possible with arm movements. This microscopic view of the organic geochemistry of a target at the Perseverance field site, when combined with the other instruments, such as Mastcam-Z, PIXL, and SuperCam, will enable unprecedented analysis of geological materials for both scientific research and determination of which samples to collect and cache for Mars sample return.
Microscopy Methods for Life Detection on Ocean Worlds
On Earth, light microscopy is commonly used in microbiology to identify organisms and observe their interactions with the environment; this makes it an attractive technique for in situ life detection methods on ocean worlds. As a standalone technique, brightfield microscopy, while able to provide important contextual information, has limited usefulness as a life detection technique because it is often challenging to differentiate between abiotic and biotic particles based solely on their size and shape, which may introduce risks of false positive or false negative interpretations. However, these risks can be reduced by combining brightfield microscopy with fluorescence microscopy to provide a method that correlate sample chemistry with sample morphology. In this work, we have used the Luminescence Imager for Exploration (LIfE), a brightfield and epifluorescence microscope with an integrated sample processing system (matured under the Concepts for Ocean worlds Life Detection Technology and Instrument Concepts of Europa Exploration programs) to develop methods that increase the fidelity of in situ microscopy life detection measurements through two main approaches. First, native fluorescence is excited in molecules that contain aromatic moieties such as proteins (using deep UV excitation), and energy carrying molecules and endogenous chromophores (using visible-light excitation), to correlate the location of these species with cell-like structural features (brightfield imaging). Second, fluorescent stains are used to selectively image cells and cell fragments by targeting proteins, lipids, and nucleic acids. We discuss the results of tests, obtained using ocean world analog samples, that have examined trades associated with implementing these methods autonomously in planetary missions, including the intrinsic properties of candidate fluorescence dyes and long-term storage and radiation stability.
Microscopy Methods for Life Detection on Ocean Worlds
On Earth, light microscopy is commonly used in microbiology to identify organisms and observe their interactions with the environment; this makes it an attractive technique for in situ life detection methods on ocean worlds. As a standalone technique, brightfield microscopy, while able to provide important contextual information, has limited usefulness as a life detection technique because it is often challenging to differentiate between abiotic and biotic particles based solely on their size and shape, which may introduce risks of false positive or false negative interpretations. However, these risks can be reduced by combining brightfield microscopy with fluorescence microscopy to provide a method that correlate sample chemistry with sample morphology. In this work, we have used the Luminescence Imager for Exploration (LIfE), a brightfield and epifluorescence microscope with an integrated sample processing system (matured under the Concepts for Ocean worlds Life Detection Technology and Instrument Concepts of Europa Exploration programs) to develop methods that increase the fidelity of in situ microscopy life detection measurements through two main approaches. First, native fluorescence is excited in molecules that contain aromatic moieties such as proteins (using deep UV excitation), and energy carrying molecules and endogenous chromophores (using visible-light excitation), to correlate the location of these species with cell-like structural features (brightfield imaging). Second, fluorescent stains are used to selectively image cells and cell fragments by targeting proteins, lipids, and nucleic acids. We discuss the results of tests, obtained using ocean world analog samples, that have examined trades associated with implementing these methods autonomously in planetary missions, including the intrinsic properties of candidate fluorescence dyes and long-term storage and radiation stability.
Resonance Energy Transfer in a Genetically Engineered Polypeptide Results in Unanticipated Fluorescence Intensity
The fluorescence intensity of a C-terminal acceptor chromophore, N-(7-dimethylamino-4-methyl coumarin (DACM), increased proportionally with 280 nm irradiation of an increasing number of donor tryptophan residues located on a β-sheet forming polypeptide. The fluorescence intensity of the acceptor chromophore increased even as the length of the β-sheet edge approached 256 Å, well beyond the Förster radius for the tryptophan–acceptor chromophore pair. Here the folding of the peptides under investigation was verified by circular dichroism (CD) and deep UV resonance Raman experiments. Control experiments showed that the enhancement of DACM fluorescence occurred concomitantly with peptide folding. In other control experiments, the DACM fluorescence intensity of the solutions of tryptophan and DACM did not show any enhancement of DACM fluorescence with increasing tryptophan concentrations. Formation of fibrillar aggregates of the substrate peptides prepared for the fluorescence studies was undetectable by thioflavin T (ThT) fluorescence.
Relative Detectability of Iron-Bearing Phases for the Mars 2020 Sherloc Deep UV Raman Instrument: 1. Focusing on Carbonates
A deep ultraviolet (DUV) Raman and fluorescence instrument is a surface standoff instrument mounted on the robotic arm of the Mars 2020 (M2020) rover Perseverance, and it is a key element of the Scanning Habitable Environments with Luminescence for Organics and Chemicals (SHERLOC) investigation [1]. Measurement and science objectives include mineralogical and organic images (~100 μm/pixel) that map sub-millimeter spatial distributions and characterization of primary and secondary minerals, potential organics, and their interaction/alteration products. The results of data analysis pertain to understanding igneous and alteration processes on Mars through time, assessing habitability, evidencing in situ biosignatures, and, along with results from other Perseverance instruments, selecting samples to cache for Mars sample return. Acceptance of the SHERLOC investigation for the NASA M2020 mission [2] created a need for Mars-relevant DUV Raman spectra, particularly for inorganic materials [e.g., 3, 4]. As reported previously [e.g., 4, 5], phases with Fe cations as essential elements (e.g., siderite (FeCO3) and ankerite (FeCa(CO3)2) for carbonates) significantly hinder detection by DUV Raman because of intense absorption of incident and scattered DUV laser radiation by Fe cations. We report here the relative detectability of carbonates (Mg,Ca,Fe,Mn)CO3 by DUV Raman using a SHERLOC analog laboratory instrument.
Pulsed laser-induced heating of mineral phases: Implications for laser-induced breakdown spectroscopy combined with Raman spectroscopy
Laser-induced breakdown spectroscopy (LIBS) and Raman spectroscopy are complementary techniques providing respectively chemical and structural information on the sample target. These techniques are increasingly used in Earth and Planetary sciences, and often together. LIBS is locally destructive for the target, and the laser-induced effects due to LIBS laser shots on the structure and on the Raman fingerprint of a set of geological samples relevant to Mars exploration are here investigated by Raman spectroscopy and electron microscopy. Experiments show that the structure of samples with low optical absorption coefficients is preserved as well as the structural information carried by Raman spectra. By contrast, minerals with high optical absorption coefficient can be severely affected by LIBS laser shots with local amorphization, melting and/or phase transformation. Thermal modeling shows that the temperature can reach several thousands of degrees at the surface for such samples during a LIBS laser shot, but decreases rapidly with time and in space. In 2020, NASA Mars 2020 mission will send a rover equipped with a combined LIBS/Raman instrument for remote analysis (SuperCam) as well as proximity science instruments at fine scale for X-ray fluorescence called PIXL for Planetary Instrument for X-ray Lithochemistry, and deep UV Raman spectroscopy called SHERLOC for Scanning Habitable Environments with Raman & Luminescence for Organics & Chemicals. Here, we discuss the implications of our results for the operation of these instruments and show that (i) the SuperCam analytical footprint for Raman spectroscopy is many times larger than the LIBS crater, minimizing any effects and (ii) SHERLOC and PIXL analysis may be affected if they analyze within a LIBS crater created by SuperCam LIBS.