Extraterrestrial Life Signature Detection Microscopy: Search and Analysis of Cells and Organics on Mars and Other Solar System Bodies

Enya, Keigo; Yoshimura, Yoshitaka; Kobayashi, Kensei; Yamagishi, Akihiko

doi:10.1007/s11214-022-00920-4

Cited by 8 publications

(8 citation statements)

References 142 publications

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“…Fluorescence microscopes are thought to be viable research instruments for extraterrestrial life [15][16][17][18][19][20]. Their targeted sensitivity limit is ~10 4 cells/g of soil; therefore, the results described above suggest that the sensitivity of the culture reported in this work was higher than that of a fluorescence microscope.…”

Section: Comparisons With Microscopymentioning

confidence: 70%

Cell culture experiment using a Mars soil simulant for the detection of extraterrestrial life

Enya

Sasaki

Fujishiro

et al. 2023

SPIE Future Sensing Technologies 2023

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We describe a cell culture experiment utilizing Mars soil simulant and yeast as a study for extraterrestrial-microorganism detection in future. The following conditions were combined to create different sample configurations: solid medium (agar plate) or liquid medium, with the Mars soil simulant (100 mg) or without, and various yeast cell densities (30 μL of a suspension of yeast prepared to achieve dilution rates of 10 −2 , 10 −3 , 10 −4 , and 10 −5 from the original suspension). For redundancy, two samples were created for each setting. On each agar plate, colonies of yeast cells were visible. In contrast, only samples created with a dilution rate of 10 −2 and 10 −3 were proven to show the presence of yeast proliferating in liquid media. The agar plates were observed via imaging using a digital camera. Colony counting was conducted using functions of the ImageJ software; the yeast density in the original suspension was estimated to be 2.5 × 10 7 colony forming units (CFU)/mL. Based on the results of cell culture on the agar plates, the detection limit that was experimentally demonstrated in this work amounted to 7.5 CFU/100 mg of soil, which corresponded to 75 CFU/g of soil. Thus, we confirmed that the Mars soil simulant reduced the number of colonies to one-third of that detected without the Mars soil simulant. In the discussion, we pointed out that the method that used the solid media has potential to provide high sensitivity using a simple, compact, and lightweight hardware. Microscope with no optics is also discussed. The lower limit of detection obtained in this study was compared with those obtained using other methods. We compared and discussed extraterrestrial life search based on (1) in situ experiments by unmanned missions, (2) analysis of samples returned to Earth, and (3) dedicated in situ analyses by manned bases located on extraterrestrial sites in the future. The perspectives discussed herein are applicable not only to the samples from Mars but also to the samples from other solar system bodies. Identifying cell culture conditions for these applications is a critical difficulty. Notably, it is a possible scenario that cell culture (although challenging) will become necessary to obtain clear evidence of extraterrestrial life. We also note that in cases where there are multiple species of microorganisms, it can happen that microorganisms that fit the culture conditions used in an experiment are selectively cultured and detected, even if the culture conditions for each species are not fully discovered. It is worthwhile to conduct exploration in advance designed with the intention of determining culture conditions with the plan to culture microorganisms in the future.

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Section: Comparisons With Microscopymentioning

confidence: 70%

Cell culture experiment using a Mars soil simulant for the detection of extraterrestrial life

Enya

Sasaki

Fujishiro

et al. 2023

SPIE Future Sensing Technologies 2023

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“…The subsequent process includes analyses with the goal of distinguishing the candidates from terrestrial organisms. Some analytical flow diagrams in assigning the candidates as being either terrestrial, non-terrestrial, or solar system originated organic compounds are presented in Enya, Yoshimura, Kobayashi and Yamagishi [47], in which Figures 1, 2, and 3, respectively, provide a flowchart in assessing the origin of the organisms on Mars through amino acid analyses, a flowchart for assessing the origin of Mars organisms through an analysis of the enantiomeric excess of amino acids, and a flowchart for assessing Mars organisms which was based on DNA analyses. These analyses can be performed in the returned samples in a ground-based laboratory on Earth without essential difficulty.…”

Section: Analysis-and-conclusion Stepmentioning

confidence: 99%

Comparative study of methods for finding extraterrestrial life based on solar system exploration and the role of microscopy in it

Enya¹,

Ymagishi²,

Kobayashi³

et al. 2023

SPIE Future Sensing Technologies 2023

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We present a comparative study of methods with a table summarizing them in searching extraterrestrial life. This work represents an update and improvement of our previous publication. In these works, microorganisms are supposed to be the object of extraterrestrial life exploration. We selected several characteristics and molecules which can be detected and analyzed in searching for extraterrestrial life. We also evaluated the search technique which targets those characteristics and molecules from several points of view. In this revision, “Generality for terrestrial microorganism” and “Source of false positives” were resolved into sub-issues and described in detail. We also introduced new issues, i.e., “Possible meaning of negative detection” and “Science objectives other than life search.” We classified life in space into three types: Earth life, Earth-independent life (life that emerged from non-life independently of Earth life), and Earth-kin life (extraterrestrial life sharing a common ancestor life with Earth life). All the methods which are effective in detecting Earth life are potentially useful for the detection of Earth-kin life. In contrast, their applicability is not guaranteed for Earth-independent life. The three major conclusions of the previous article were re-evaluated and confirmed to remain unchanged: (i) there is no realistic single detection method which can conclude on the discovery of life independent of Earth, (ii) there is no single best method which is superior to the other methods in all respects, and (iii) there is no single method which can distinguish between Earth life and life independent of Earth. Subsequently, we discussed the combination of the detection methods and dividing extraterrestrial microorganism search into multiple steps: the survey and- detection step and the analysis-and-conclusion step. We also emphasized the importance of imaging; even with not enough resolution for observing the detailed morphology of a microbial cell, it is highly worthwhile to distinguish whether the detected molecules are concentrated at micron size or diffused thinly. Of course, high-resolution imaging provides information with regard to the morphology of microorganisms. Accordingly, imaging is necessary for both the survey-and-detection and the analysis-and-conclusion steps. To realize a space exploration mission or instrument for extraterrestrial life search, not only in scientific but programmatic sense, it is important to design it so that scientific results can be obtained even if no life is found. The studies included in this article primarily supposed Mars as a target; however, they are valid for a wide variety of planets of the solar system, as well as their moons and small objects.

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“…In rock samples, the cells may be within the rock itself (endolithic); bacteria may also be associated with soil particles, biominerals and other inorganics as well as nonbiogenic organic material such as polycyclic aromatic hydrocarbons. [5][6][7] This material can bind dyes and enhance their fluorescence. Nonspecific binding often is brighter than cells and may resemble bacterial cells, and separation of cells from many soil types is extremely difficult.…”

Section: Introductionmentioning

confidence: 99%

“…However, in complex environments such as soil and rock, it can be challenging to separate the signal of labelled cells from mineral autofluorescence and nonspecific dye binding. In rock samples, the cells may be within the rock itself (endolithic); bacteria may also be associated with soil particles, biominerals and other inorganics as well as nonbiogenic organic material such as polycyclic aromatic hydrocarbons 5–7 . This material can bind dyes and enhance their fluorescence.…”

Section: Introductionmentioning

confidence: 99%

The use of fluorescence lifetime imaging (FLIM) for in situ microbial detection in complex mineral substrates

Chmykh,

Nadeau

2024

Journal of Microscopy

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The utility of fluorescence lifetime imaging microscopy (FLIM) for identifying bacteria in complex mineral matrices was investigated. Baseline signals from unlabelled Bacillus subtilis and Euglena gracilis, and Bacillus subtilis labelled with SYTO 9 were obtained using two‐photon excitation at 730, 750 and 800 nm, identifying characteristic lifetimes of photosynthetic pigments, unpigmented cellular autofluorescence, and SYTO 9. Labelled and unlabelled B. subtilis were seeded onto marble and gypsum samples containing endolithic photosynthetic cyanobacteria and the ability to distinguish cells from mineral autofluorescence and nonspecific dye staining was examined in parallel with ordinary multichannel confocal imaging. It was found that FLIM enabled discrimination of SYTO 9 labelled cells from background, but that the lifetime of SYTO 9 was shorter in cells on minerals than in pure culture under our conditions. Photosynthetic microorganisms were easily observed using both FLIM and confocal. Unlabelled, nonpigmented bacteria showed weak signals that were difficult to distinguish from background when minerals were present, though cellular autofluorescence consistent with NAD(P)H could be seen in pure cultures, and phasor analysis permitted detection on rocks. Gypsum and marble samples showed similar autofluorescence profiles, with little autofluorescence in the yellow‐to‐red range. Lifetime or time‐gated imaging may prove a useful tool for environmental microbiology.LAY DESCRIPTION: The standard method of bacterial enumeration is to label the cells with a fluorescent dye and count them under high‐power fluorescence microscopy. However, this can be difficult when the cells are embedded in soil and rock due to fluorescence from the surrounding minerals and dye binding to ambiguous features of the substrate. The use of fluorescence lifetime imaging (FLIM) can disambiguate these signals and allow for improved detection of bacteria in environmental samples.

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Extraterrestrial Life Signature Detection Microscopy: Search and Analysis of Cells and Organics on Mars and Other Solar System Bodies

Cited by 8 publications

References 142 publications

Cell culture experiment using a Mars soil simulant for the detection of extraterrestrial life

Cell culture experiment using a Mars soil simulant for the detection of extraterrestrial life

Comparative study of methods for finding extraterrestrial life based on solar system exploration and the role of microscopy in it

The use of fluorescence lifetime imaging (FLIM) for in situ microbial detection in complex mineral substrates

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