Morphological Biosignatures Research Articles

Bacterial spore morphology remains highly recognizable after exposure to simulated Enceladus and Europa surface conditions

The subsurface oceans of Enceladus and Europa are thought to be some of the best candidate environments for finding life beyond Earth. Realistically, the first missions aimed at searching for life on these worlds will likely be restricted to the shallow subsurface. Here, we investigated whether indicators of life, or biosignatures, deposited near the surface could persist long enough to be detected, given that the extremely harsh conditions there would tend to degrade them. We exposed Bacillus subtilis spores to Ocean World surface conditions and used electron microscopy combined with spectroscopic approaches to assess the preservation potential of structural and morphological biosignatures derived from spores. Our results show that spore structure is highly resilient in the face of extreme conditions long after they have been inactivated, suggesting that methods targeting cell morphology would be valuable components in a suite of life detection strategies used in future missions to Ocean Worlds.

Unveiling Challenging Microbial Fossil Biosignatures from Rio Tinto with Micro-to-Nanoscale Chemical and Ultrastructural Imaging.

Understanding the nature and preservation of microbial traces in extreme environments is crucial for reconstructing Earth's early biosphere and for the search for life on other planets or moons. At Rio Tinto, southwestern Spain, ferric oxide and sulfate deposits similar to those discovered at Meridiani Planum, Mars, entomb a diversity of fossilized organisms, despite chemical conditions commonly thought to be challenging for life and fossil preservation. Investigating this unique fossil microbiota can elucidate ancient extremophile communities and the preservation of biosignatures in acidic environments on Earth and, potentially, Mars. In this study, we use an innovative multiscale approach that combines the state-of-the-art synchrotron X-ray nanoimaging methods of ptychographic X-ray computed laminography and nano-X-ray fluorescence to reveal Rio Tinto's microfossils at subcellular resolution. The unprecedented nanoscale views of several different specimens within their geological and geochemical contexts reveal novel intricacies of preserved microbial communities. Different morphotypes, ecological interactions, and possible taxonomic affinities were inferred based on qualitative and quantitative 3D ultrastructural information, whereas diagenetic processes and metabolic affinities were inferred from complementary chemical information. Our integrated nano-to-microscale analytical approach revealed previously invisible microbial and mineral interactions, which complemented and filled a gap of spatial resolution in conventional methods. Ultimately, this study contributes to the challenge of deciphering the faint chemical and morphological biosignatures that can indicate life's presence on the early Earth and on distant worlds.

Rapid Diagenesis and Microbial Biosignature Degradation in Spring Carbonates from Crystal Geyser, Utah

Carbonate rocks retain a well preserved record of biologically associated structures at the outcrop to millimeter scale, however microscale features such as cellular fossils are rarely represented. The lack of microscale textural information in ancient carbonates is commonly attributed to processes relating to carbonate diagenesis. However, there are relatively few examples of precisely how and when these destructive processes occur, particularly in active precipitating systems. To better understand the taphonomy of carbonate precipitating environments through early diagenesis, we investigated Crystal Geyser, an active cold water carbonate spring ( about 18 deg C) located in Grand County, Utah. Here we show that rapid precipitation is effective at initially capturing cell-like structures and forming associated microscale laminated stromatolites; however, these morphologies degrade immediately after their formation. We attribute destructive diagenetic effects to the recrystallization of metastable aragonite into the more stable polymorph calcite (i.e., inversion) and the associated textural coarsening that homogenizes and erases the original fabric (i.e., aggrading neomorphism). Despite the loss of microscale morphological information, chemical biosignatures in the form of macromolecular organics remain dispersed throughout the disrupted carbonate textures. These observations provide an example of penecontemporaneous diagenesis that obliterates primary microscale textures in carbonate rocks. Similar mechanisms and their rapid timing, as shown here, likely contributes to the observed lack of microscale morphological biosignatures in many ancient carbonates. This work further highlights that within such systems, permineralization by a more stable crystalline phase, such as chert, must occur rapidly after deposition to effectively retain these signatures over geological timescales.

Connecting molecular biomarkers, mineralogical composition, and microbial diversity from Mars analog lava tubes

Lanzarote (Canary Islands, Spain) is one of the best terrestrial analogs to Martian volcanology. Particularly, Lanzarote lava tubes may offer access to recognizably preserved chemical and morphological biosignatures valuable for astrobiology. By combining microbiological, mineralogical, and organic geochemistry tools, an in-depth characterization of speleothems and associated microbial communities in lava tubes of Lanzarote is provided. The aim is to untangle the underlying factors influencing microbial colonization in Earth's subsurface to gain insight into the possibility of similar subsurface microbial habitats on Mars and to identify biosignatures preserved in lava tubes unequivocally.The microbial communities with relevant representativeness comprise chemoorganotrophic, halophiles, and/or halotolerant bacteria that have evolved as a result of the surrounding oceanic environmental conditions. Many of these bacteria have a fundamental role in reshaping cave deposits due to their carbonatogenic ability, leaving behind an organic record that can provide evidence of past or present life. Based on functional profiling, we infer that Crossiella is involved in fluorapatite precipitation via urea hydrolysis and propose its Ca-rich precipitates as compelling biosignatures valuable for astrobiology.In this sense, analytical pyrolysis, stable isotope analysis, and chemometrics were conducted to characterize the complex organic fraction preserved in the speleothems and find relationships among organic families, microbial taxa, and precipitated minerals.We relate organic compounds with subsurface microbial taxa, showing that organic families drive the microbiota of Lanzarote lava tubes. Our data indicate that bacterial communities are important contributors to biomarker records in volcanic-hosted speleothems. Within them, the lipid fraction primarily consists of low molecular weight n-alkanes, α-alkenes, and branched-alkenes, providing further evidence that microorganisms serve as the origin of organic matter in these formations. The ongoing research in Lanzarote's lava tubes will help develop protocols, routines, and predictive models that could provide guidance on choosing locations and methodologies for searching potential biosignatures on Mars.

Controls on Reactive Oxygen Species Cycles in Yellowstone Hot Springs: Implications for Biosignature Preservation on Mars

Early Earth and Mars had analogous environments. While life developed on our planet, the question of whether it did on Mars remains to be answered. Hot spring deposits are compelling targets for exploration because of their high habitability and potential to retain morphological and chemical biosignatures. As a result in this study, we aim to better understand the potential for biosignature preservation in Fe-bearing hydrothermal systems. Understanding oxidation-reduction reactions involving Fe in hot springs is a key step in elucidating the preservation process. Fe reacts readily with reactive oxygen species (ROS), which are produced in hot spring surface waters through photochemical processes. Furthermore, Fe3+ can bind to cell membranes and preserve complex organic molecules (i.e., biomarkers). ROS formation is typically controlled by photoreactions with dissolved organic matter (DOM). However, Fe redox reactions more likely control ROS formation in these Fe-bearing systems. We deconvolved the relationship of ROS with Fe in hot springs and evaluated the role that DOM and dissolved organic sulfur (DOS) may have in ROS production. To better understand these coupled systems, field and laboratory experiments were conducted in hot springs of Yellowstone National Park. In situ H2O2 concentrations observed in these hot springs were comparable to, or higher than, those of other high-temperature systems. Reaction rates determined by measuring concentrations after specified time intervals varied based on water compositions and the presence of particulate or dissolved matter. Fe speciation (photochemical reactivity), concentration, and solubility further determined ROS cycling rates. Specifically, photochemically active Fe enhanced both ROS formation and decay rates depending on incident UV irradiance, and rates increased along with Fe concentration and solubility (i.e., in acidic conditions). Better understanding how ROS and Fe cycle in predominantly abiotic conditions will eventually aid in distinguishing between biosignatures and abiotic substances in the rock record.

Oligotrophic Growth of Nitrate-Dependent Fe2+-Oxidising Microorganisms Under Simulated Early Martian Conditions.

Nitrate-dependent Fe2+ oxidation (NDFO) is a microbially mediated process observed in many anaerobic, low-nutrient (oligotrophic) neutral–alkaline environments on Earth, which describes oxidation of Fe2+ to Fe3+ in tandem with microbial nitrate reduction. Evidence suggests that similar environments existed on Mars during the Noachian epoch (4.1–3.7 Ga) and in periodic, localised environments more recently, indicating that NDFO metabolism could have played a role in a potential early martian biosphere. In this paper, three NDFO microorganisms, Acidovorax sp. strain BoFeN1, Pseudogulbenkiania sp. strain 2002 and Paracoccus sp. strain KS1, were assessed for their ability to grow oligotrophically in simulated martian brines and in a minimal medium with olivine as a solid Fe2+ source. These simulant-derived media were developed from modelled fluids based on the geochemistry of Mars sample locations at Rocknest (contemporary Mars soil), Paso Robles (sulphur-rich soil), Haematite Slope (haematite-rich soil) and a Shergottite meteorite (common basalt). The Shergottite medium was able to support growth of all three organisms, while the contemporary Mars medium supported growth of Acidovorax sp. strain BoFeN1 and Pseudogulbenkiania sp. strain 2002; however, growth was not accompanied by significant Fe2+ oxidation. Each of the strains was also able to grow in oligotrophic minimal media with olivine as the sole Fe2+ source. Biomineralised cells of Pseudogulbenkiania sp. strain 2002 were identified on the surface of the olivine, representing a potential biosignature for NDFO microorganisms in martian samples. The results suggest that NDFO microorganisms could have thrived in early martian groundwaters under oligotrophic conditions, depending on the local lithology. This can guide missions in identifying palaeoenvironments of interest for biosignature detection. Indeed, biomineralised cells identified on the olivine surface provide a previously unexplored mechanism for the preservation of morphological biosignatures in the martian geological record.

Methanogenesis from Mineral Carbonates, a Potential Indicator for Life on Mars

Priorities for the exploration of Mars involve the identification and observation of biosignatures that indicate the existence of life on the planet. The atmosphere and composition of the sediments on Mars suggest suitability for anaerobic chemolithotrophic metabolism. Carbonates are often considered as morphological biosignatures, such as stromatolites, but have not been considered as potential electron acceptors. Within the present study, hydrogenotrophic methanogen enrichments were generated from sediments that had received significant quantities of lime from industrial processes (lime kiln/steel production). These enrichments were then supplemented with calcium carbonate powder or marble chips as a sole source of carbon. These microcosms saw a release of inorganic carbon into the liquid phase, which was subsequently removed, resulting in the generation of methane, with 0.37 ± 0.09 mmoles of methane observed in the steel sediment enrichments supplemented with calcium carbonate powder. The steel sediment microcosms and lime sediments with carbonate powder enrichments were dominated by Methanobacterium sp., whilst the lime/marble enrichments were more diverse, containing varying proportions of Methanomassiliicoccus, Methanoculleus and Methanosarcina sp. In all microcosm experiments, acetic acid was detected in the liquid phase. Our results indicate that chemolithotrophic methanogenesis should be considered when determining biosignatures for life on Mars.

Planetary Mass Spectrometry for Agnostic Life Detection in the Solar System

For the past fifty years of space exploration, mass spectrometry has provided unique chemical and physical insights on the characteristics of other planetary bodies in the Solar System. A variety of mass spectrometer types, including magnetic sector, quadrupole, time-of-flight, and ion trap, have and will continue to deepen our understanding of the formation and evolution of exploration targets like the surfaces and atmospheres of planets and their moons. An important impetus for the continuing exploration of Mars, Europa, Enceladus, Titan, and Venus involves assessing the habitability of solar system bodies and, ultimately, the search for life—a monumental effort that can be advanced by mass spectrometry. Modern flight-capable mass spectrometers, in combination with various sample processing, separation, and ionization techniques enable sensitive detection of chemical biosignatures. While our canonical knowledge of biosignatures is rooted in Terran-based examples, agnostic approaches in astrobiology can cast a wider net, to search for signs of life that may not be based on Terran-like biochemistry. Here, we delve into the search for extraterrestrial chemical and morphological biosignatures and examine several possible approaches to agnostic life detection using mass spectrometry. We discuss how future missions can help ensure that our search strategies are inclusive of unfamiliar life forms.

Identifying optimal working conditions for close-up imagining during the ExoMars rover mission

A Close-Up Imager named CLUPI is one of the instruments that will be onboard the Rosalind Franklin rover, a robot that will explore the surface of Mars in the framework of the ESA/Roscosmos ExoMars mission. CLUPI will be principally used for acquiring close-up images of rock textures and sedimentary structures, identifying materials that may record information about the hypothetical existence of past microbial life. Although the technical specifications of CLUPI are well known, it is not possible to readily translate such specifications in terms of feasibility to recognize “textures of interest” at a given distance under specific light conditions on Mars. Accurate predictions are important for making fast and informed decisions during the daily tactical planning of the rover. Here, we describe the results of some mission-preparation activities, during which a commercial camera that allows for producing images analogue to those of CLUPI has been used to photograph rock samples in an indoor facility (i.e., the Marslabor of the University of Basel) that has been built ad hoc for simulating a Martian landscape. By varying the working distance and light conditions it has been possible to perform a preliminary assessment of the minimal-working-distance required for interpreting rock textures and sedimentary structures that are potentially present on Mars, including textures that allow for differentiating sedimentary rocks from igneous rocks, grains that allow for classifying sedimentary rocks based on their granulometry, and stromatolitic laminations representing morphological biosignatures. In general, the results suggest that rock textures tend to be recognizable even from distances that exceed those one would predict based on the resolution of the instrument and the size of the structure or particles that defines the rock texture or sedimentary structure. We also show that the angle between the illumination axis (i.e., the direction of incident light) and the target surface plays a significant role for the recognition of textural and compositional heterogeneities within the acquired images. The produced data represents a first step in identifying ideal CLUPI working-distances and illumination, and in preparing an image database that will be of help for optimizing rover operations and the scientific return of CLUPI during the ExoMars mission.

Varietal Classification of Lactuca Sativa Seeds Using an Adaptive Neuro-Fuzzy Inference System Based on Morphological Phenes

Seed varieties are often differentiated via the manual and subjective classification of their external textural, spectral, and morphological biosignatures. This traditional method of manually inspecting seeds is inefficient and unreliable for seed phenotyping. The application of computer vision is an ideal solution allied with computational intelligence. This study used Lactuca sativa seed variants, which are commercially known as grand rapid, Chinese loose-leaf, and iceberg (which serves as noise data for extended model evaluation), in determining their corresponding classifications based on the extended morphological phenes using computational intelligence. Red-green-blue (RGB) imaging was employed for individual kernels. Extended morphological phenes, that is, solidity, roundness, compactness, and shape factors, were computed based on seed architectural traits and used as predictors to discriminate among the three cultivars. The suitability of ANFIS, NB, and CT was explored using a limited dataset. A mean accuracy of 100% was manifested in ANFIS; thus, it was proved to be the most reliable model.

Hyperdiverse archaea near life limits at the polyextreme geothermal Dallol area.

Microbial life has adapted to various individual extreme conditions; yet, organisms simultaneously adapted to very low pH, high salt and high temperature are unknown. We combined environmental 16S/18S rRNA-gene metabarcoding, cultural approaches, fluorescence-activated cell sorting, scanning electron microscopy and chemical analyses to study samples along such unique polyextreme gradients in the Dallol-Danakil area (Ethiopia). We identify two physicochemical barriers to life in the presence of surface liquid water defined by: i) high chaotropicity-low water activity in Mg2+/Ca2+-dominated brines and ii) hyperacidity-salt combinations (pH~0/NaCl-dominated salt-saturation). When detected, life was dominated by highly diverse ultrasmall archaea widely distributed across phyla with and without previously known halophilic members. We hypothesize that high cytoplasmic K+-level was an original archaeal adaptation to hyperthermophily, subsequently exapted during multiple transitions to extreme halophily. We detect active silica encrustment/fossilization of cells but also abiotic biomorphs of varied chemistry. Our work helps circumscribing habitability and calls for cautionary interpretations of morphological biosignatures on Earth and beyond.

Macroscopic and Microscopic Morphological Features of Stromatolites Related To Activity of Eukaryote-Dominated Biofilms in an Acid Mine Drainage Environment: Biosignatures and Understanding Preservation of Stromatolites as Trace Fossils

Euglena-, diatom-, and algae-dominated biofilms are the principal producers of iron-rich biolaminates that result in biosedimentary structures, or stromatolites, in an acid mine drainage (AMD) environment in Indiana. These structures are considered trace fossils because they are produced by organism-sediment interactions and record physicochemical conditions of the environment. Our purpose was to link the biofilm types to specific micro- and micromorphological features and the physicochemical conditions under which they were formed. Analyses revealed that Euglena-dominated biofilm produced thin, porous microlaminae by trapping, binding, and relocating AMD precipitates as the biofilm kept pace with chemical sedimentation. More massive microlaminae were produced by high rates of chemical sedimentation brought on by increased discharge and dilution of acidity. Diatom- and algae-dominated biofilms produced thick, mm–cm-scale, porous, spongelike micro- to macrolaminae through oxygenic photosynthesis and/or metal uptake in extracellular polymeric substances, which promoted mineral precipitation on cell walls to create a rigid, porous structure. The variations in biolaminate textures within the stromatolites record seasonal changes in the microbial populations and physicochemical conditions of the AMD environment. These iron-rich stromatolites represent trace fossils that record morphological biosignatures of eukaryote-dominated microbial biofilms and may serve as appropriate proxies in the search for similar evidence of eukaryotic life in other iron-rich paleoenvironments, such as those on early Earth and Mars.

Nitrate-Dependent Iron Oxidation: A Potential Mars Metabolism.

This work considers the hypothetical viability of microbial nitrate-dependent Fe2+ oxidation (NDFO) for supporting simple life in the context of the early Mars environment. This draws on knowledge built up over several decades of remote and in situ observation, as well as recent discoveries that have shaped current understanding of early Mars. Our current understanding is that certain early martian environments fulfill several of the key requirements for microbes with NDFO metabolism. First, abundant Fe2+ has been identified on Mars and provides evidence of an accessible electron donor; evidence of anoxia suggests that abiotic Fe2+ oxidation by molecular oxygen would not have interfered and competed with microbial iron metabolism in these environments. Second, nitrate, which can be used by some iron oxidizing microorganisms as an electron acceptor, has also been confirmed in modern aeolian and ancient sediment deposits on Mars. In addition to redox substrates, reservoirs of both organic and inorganic carbon are available for biosynthesis, and geochemical evidence suggests that lacustrine systems during the hydrologically active Noachian period (4.1–3.7 Ga) match the circumneutral pH requirements of nitrate-dependent iron-oxidizing microorganisms. As well as potentially acting as a primary producer in early martian lakes and fluvial systems, the light-independent nature of NDFO suggests that such microbes could have persisted in sub-surface aquifers long after the desiccation of the surface, provided that adequate carbon and nitrates sources were prevalent. Traces of NDFO microorganisms may be preserved in the rock record by biomineralization and cellular encrustation in zones of high Fe2+ concentrations. These processes could produce morphological biosignatures, preserve distinctive Fe-isotope variation patterns, and enhance preservation of biological organic compounds. Such biosignatures could be detectable by future missions to Mars with appropriate instrumentation.

The Close-Up Imager Onboard the ESA ExoMars Rover: Objectives, Description, Operations, and Science Validation Activities.

The Close-Up Imager (CLUPI) onboard the ESA ExoMars Rover is a powerful high-resolution color camera specifically designed for close-up observations. Its accommodation on the movable drill allows multiple positioning. The science objectives of the instrument are geological characterization of rocks in terms of texture, structure, and color and the search for potential morphological biosignatures. We present the CLUPI science objectives, performance, and technical description, followed by a description of the instrument's planned operations strategy during the mission on Mars. CLUPI will contribute to the rover mission by surveying the geological environment, acquiring close-up images of outcrops, observing the drilling area, inspecting the top portion of the drill borehole (and deposited fines), monitoring drilling operations, and imaging samples collected by the drill. A status of the current development and planned science validation activities is also given. Key Words: Mars-Biosignatures-Planetary Instrumentation. Astrobiology 17, 595-611.

Increased mud deposition reduces stromatolite complexity

Microbial mat growth can produce diagnostic delicate structures in low-energy environments, but mat morphology, and thus preservation of delicate structures as morphological biosignatures, is affected by mud accumulation. In Lake Joyce, Antarctica, varying rates of siliciclastic mud deposition affected the morphology of partially calcified microbial mats. These mats were characterized along a shore-parallel transect from higher- to lower-sedimentation regions of the lake. Mats with high mud accumulation rates were predominantly flat with sparse pinnacles. At lower sedimentation rates, mats developed more pinnacles and more delicate features, including vertical webs of biofilm suspended between pinnacles. Morphological transitions suggest that mud deposition disrupts growth of delicate microbial structures like biofilm webs and inhibits pinnacle growth at higher accumulation rates. The simplification of mat morphology with increasing mud accumulation has important implications for the distribution of stromatolite types in the rock record. Stromatolites with intricate biofilm structures are rare, possibly due to the paucity of appropriate environments rather than the intrinsic properties of microbial communities. Communities capable of forming delicate microbialites likely produced morphologically simpler pinnacles and flat mat in environments with significant siliciclastic mud or micrite influx. Thus, delicate mats have narrow environmental ranges for growth, and temporal or spatial variations in siliciclastic mud or micrite sedimentation rates bias the record of microbial activity preserved in stromatolites.

In Wave Tanks, Microbial Fragments Wrangle Sands into Wrinkle Structures

Wrinkle structures, found in ancient sedimentary environments along the surfaces of sandy beds, can be formed experimentally by adding microbial fragments to sand layers within wave tanks, according to Giulio Mariotti of the Massachusetts Institute of Technology (MIT) in Cambridge and his collaborators there and at Smith College in Northampton, Mass. They say that wrinkle structures in natural sediments are “morphological biosignatures” of microbial fragments—inscribed at the sediment–water interface in wave-dominated environments, and not beneath larger microbial mats as previously thought. Those wrinkle structures likely formed early during the evolution of animals, possibly while they were grazing on microbial fragments. Details appeared 31 August 2014 in Nature Geoscience Letters (doi:10.1038/ngeo2229).

Microbial shaping of sedimentary wrinkle structures

Wrinkle structures in ancient sedimentary environments are enigmatic. Wave-tank experiments suggest that wrinkle structures are shaped by microbial mat fragments that are moved by waves over sandy-bed surfaces, and thus are morphological biosignatures.

MODERN TERRESTRIAL SEDIMENTARY BIOSTRUCTURES AND THEIR FOSSIL ANALOGS IN MESOPROTEROZOIC SUBAERIAL DEPOSITS

Microbial communities developing on modern clastic sedimentary surfaces of arid lands are dominated by phototrophic microorganisms that form a variety of characteristic “microbially induced sedimentary structures” (MISS) through their interactions with detrital sedimentary grains, aided by secretions of extracellular polymeric substances and other organic materials. In this study, we describe modern MISS from unvegetated arid topsoils and compare them with fossil MISS found within decimeter- to meter-thick sedimentary sequences of Mesoproterozoic siliciclastic outcrops of the Dripping Spring Quartzite formation of the Apache Group in central Arizona, USA. These sequences contain numerous bedding plane exposures with desiccation surfaces including polygonal cracks, curls, and chips. Repetition of these structures within stratigraphic sequences indicates recurring episodes of subaerial exposure. Some of these MISS contain cellular microfossils that exhibit morphological adaptations for surviving desiccation. The strong similarities between modern and ancient MISS in this study provide additional criteria for recognizing morphological biosignatures of terrestrial microbial communities in ancient deposits. Our results provide compelling evidence for the presence of land-based microbial communities by the Mesoproterozoic (∼1200 Ma). The association of MISS features further suggests that the primary producers that had colonized Mesoproterozoic land surfaces were likely desiccation-adapted photosynthetic microbes, similar to modern desert soil crust communities.

Morphology of biogenic iron oxides records microbial physiology and environmental conditions: toward interpreting iron microfossils

Despite the abundance of Fe and its significance in Earth history, there are no established robust biosignatures for Fe(II)-oxidizing micro-organisms. This limits our ability to piece together the history of Fe biogeochemical cycling and, in particular, to determine whether Fe(II)-oxidizers played a role in depositing ancient iron formations. A promising candidate for Fe(II)-oxidizer biosignatures is the distinctive morphology and texture of extracellular Fe(III)-oxyhydroxide stalks produced by mat-forming microaerophilic Fe(II)-oxidizing micro-organisms. To establish the stalk morphology as a biosignature, morphologic parameters must be quantified and linked to the microaerophilic Fe(II)-oxidizing metabolism and environmental conditions. Toward this end, we studied an extant model organism, the marine stalk-forming Fe(II)-oxidizing bacterium, Mariprofundus ferrooxydans PV-1. We grew cultures in flat glass microslide chambers, with FeS substrate, creating opposing oxygen/Fe(II) concentration gradients. We used solid-state voltammetric microelectrodes to measure chemical gradients in situ while using light microscopy to image microbial growth, motility, and mineral formation. In low-oxygen (2.7-28μm) zones of redox gradients, the bacteria converge into a narrow (100μm-1mm) growth band. As cells oxidize Fe(II), they deposit Fe(III)-oxyhydroxide stalks in this band; the stalks orient directionally, elongating toward higher oxygen concentrations. M.ferrooxydans stalks display a narrow range of widths and uniquely biogenic branching patterns, which result from cell division. Together with filament composition, these features (width, branching, and directional orientation) form a physical record unique to microaerophilic Fe(II)-oxidizer physiology; therefore, stalk morphology is a biosignature, as well as an indicator of local oxygen concentration at the time of formation. Observations of filamentous Fe(III)-oxyhydroxide microfossils from a ~170Ma marine Fe-Si hydrothermal deposit show that these morphological characteristics can be preserved in the microfossil record. This study demonstrates the potential of morphological biosignatures to reveal microbiology and environmental chemistry associated with geologic iron formation depositional processes.

Quantitative characterization of preneoplastic progression using single‐cell computed tomography and three‐dimensional karyometry

The development of morphological biosignatures to precisely characterize preneoplastic progression necessitates high-resolution three-dimensional (3D) cell imagery and robust image processing algorithms. We report on the quantitative characterization of nuclear structure alterations associated with preneoplastic progression in human esophageal epithelial cells using single-cell optical tomography and fully automated 3D karyometry. We stained cultured cells with hematoxylin and generated 3D images of individual cells by mathematically reconstructing 500 projection images acquired using optical absorption tomographic imaging. For 3D karyometry, we developed novel, fully automated algorithms to robustly segment the cellular, nuclear, and subnuclear components in the acquired cell images, and computed 41 quantitative morphological descriptors from these segmented volumes. In addition, we developed algorithms to quantify the spatial distribution and texture of the nuclear DNA. We applied our methods to normal, metaplastic, and dysplastic human esophageal epithelial cell lines, analyzing 100 cells per line. The 3D karyometric descriptors elucidated quantitative differences in morphology and enabled robust discrimination between cell lines on the basis of extracted morphological features. The morphometric hallmarks of cancer progression such as increased nuclear size, elevated nuclear content, and anomalous chromatin texture and distribution correlated with this preneoplastic progression model, pointing to the clinical use of our method for early cancer detection.