Jan K. Toporski

The simulated silicification of bacteria--new clues to the modes and timing of bacterial preservation and implications for the search for extraterrestrial microfossils.

Evidence of microbial life on Earth has been found in siliceous rock formations throughout the geological and fossil record. To understand the mechanisms of silicification and thus improve our search patterns for evidence of fossil microbial life in rocks, a series of controlled laboratory experiments were designed to simulate the silicification of microorganisms. The bacterial strains Pseudomonas fluorescens and Desulphovibrio indonensis were exposed to silicifying media. The experiments were designed to determine how exposure time to silicifying solutions and to silicifying solutions of different Si concentration affect the fossilization of microbial biofilms. The silicified biofilms were analyzed using transmission electron microscopy (TEM) in combination with energy-dispersive spectroscopy. Both bacterial species showed evidence of silicification after 24 h in 1,000 ppm silica solution, although D. indonensis was less prone to silicification. The degree of silicification of individual cells of the same sample varied, though such variations decreased with increasing exposure time. High Si concentration resulted in better preservation of cellular detail; the Si concentration was more important than the duration in Si solution. Even though no evidence of amorphous silica precipitation was observed, bacterial cells became permineralized. High-resolution TEM analysis revealed nanometer-sized crystallites characterized by lattice fringe-spacings that match the (10-11) d-spacing of quartz formed within bacterial cell walls after 1 week in 5,000 ppm silica solution. The mechanisms of silicification under controlled laboratory conditions and the implication for silicification in natural environments are discussed, along with the relevance of our findings in the search for early life on Earth and extraterrestrial life.

Polymeric substances and biofilms as biomarkers in terrestrial materials: Implications for extraterrestrial samples

Organic polymeric substances are a fundamental component of microbial biofilms. Microorganisms, especially bacteria, secrete extracellular polymeric substances (EPS) to form slime layers in which they reproduce. In the sedimentary environment, biofilms commonly contain the products of degraded bacteria as well as allochthonous and autochthonous mineral components. They are complex structures which serve as protection for the colonies of microorganisms living in them and also act as nutrient traps. Biofilms are almost ubiquitous wherever there is an interface and moisture (liquid/liquid, liquid/solid, liquid/gas, solid/gas). In sedimentary rocks they are commonly recognized as stromatolites. We also discuss the distinction between bacterial biofilms and prebiotic films. The EPS and cell components of the microbial biofilms contain many cation chelation sites which are implicated in the mineralization of the films. EPS, biofilms, and their related components thus have strong preservation potential in the rock record. Fossilized microbial polymeric substances (FPS) and biofilms appear to retain the same morphological characteristics as the unfossilized material and have been recognized in rock formations dating back to the Early Archaean (3.5 b.y.). We describe FPS and biofilms from hot spring, deep-sea, volcanic lake, and shallow marine/littoral environments ranging up to 3.5 b.y. in age. FPS and biofilms are more commonly observed than fossil bacteria themselves, especially in the older part of the terrestrial record. The widespread distribution of microbial biofilms and their great survival potential makes their fossilized remains a useful biomarker as a proxy for life with obvious application to the search for life in extraterrestrial materials.

Morphologic and spectral investigation of exceptionally well-preserved bacterial biofilms from the Oligocene Enspel formation, Germany

Abstract The fossilised soft tissues of a tadpole and an associated coprolitic structure from the organic-rich volcanoclastic lacustrine Upper Oligocene Enspel sediments (Germany) were investigated using high-resolution imaging techniques and nondestructive in situ surface analysis. Total organic carbon analysis of the coprolite and the sediment revealed values of 28.9 and 8.9% respectively. The soft tissues from the tadpole and the coprolite were found to be composed of 0.5 to 1 μm-sized spheres and rod shapes. These features are interpreted as the fossil remains of bacterial biofilms consisting probably of heterotrophic bacteria and fossilised extracellular polymeric substances. They became fossilised while in the process of degrading the organic matter of the organism and the coprolite. Comparison with a modern marine biofilm revealed morphologic details identical to those observed in the fossil bacterial biofilms. Although the fossil biofilms on both macrofossils exhibited identical microtextures, their mode of preservation was inhomogeneous and varied between calcium phosphate and an as yet unidentified mineral phase consisting mainly of Si, Ca, Ti, P, and S, but also showing the presence of Mg, Al, and Fe. The coprolite consists purely of fossilised bacterial cells in a densely packed arrangement and associated fossilised extracellular polymeric substances. In addition to preliminary imaging and energy-dispersive X-ray analysis, both the fossil biofilms and the sediment were investigated by nondestructive in situ analysis using time of flight-secondary ion mass spectroscopy (ToF-SIMS). The mass spectra obtained on the coprolite in mass-resolved chemical mapping mode allowed the tentative identification of a number of organic secondary ion species. Some spectra appear to indicate the presence of bacterial hopanoids, but further work using standard techniques such as gas chromatography mass spectroscopy is needed to conclusively verify the presence of these substances. Nevertheless, ToF-SIMS chemical maps were successfully correlated with electron microscopy images, allowing the correlation of molecular spectra, the spatial distribution of individual organic species, and specific morphologic features to demonstrate the potential of this approach in the analysis of microfossils.

Life on Mars: evaluation of the evidence within Martian meteorites ALH84001, Nakhla, and Shergotty

Abstract Analyses both support and are in opposition to the hypothesis that the Martian meteorite ALH84001 contains evidence for possible biogenic activity on Mars. New observations in two additional Martian meteorites, Nakhla (1.3 Ga old) and Shergotty (300–165 Ma old) indicate possible biogenic features. Features in the three Martian meteorites compare favorably with the accepted criteria for terrestrial microfossils and evidence for early life on the Earth. There is strong evidence for the presence of indigenous reduced carbon, biogenic magnetite, and the low-temperature formation of carbonate globules. The morphological similarities between terrestrial microfossils, biofilms, and the features found in the three Martian meteorites are intriguing but have not been conclusively proven. Every investigation must recognize the possibility of terrestrial contamination of the meteorites, whether or not the meteorites are Martian. The search for evidence of ancient life in Martian meteorites has emphasized the difficulties confronting the scientific community with the respect to the positive identification of evidence of past biogenic activity.

Time of flight secondary ion mass spectrometry (ToFSIMS) of a number of hopanoids

Abstract Time of flight secondary ion mass spectrometry (ToFSIMS) has been applied to a number of bacterial hopanoids in an attempt to characterise these geologically important molecules in situ by a surface sensitive technique. Our results show that these molecules can be detected using this instrumentation to a high degree of mass accuracy. We believe that ToFSIMS can, therefore, be used to identify these molecules in environmental samples where sample size may be an issue and contraindicate the use of more traditional techniques such as GC–MS.

Fossil Biofilms and the Search for Life on Mars

Microbial biofilms and mats are documented as fossils in rocks throughout the 3.5 b.y.-old morphological fossil record of life on Earth (Westall et al., 2000). The polymer-rich biofilms are, per se, highly robust structures capable of great resistance and durability. Moreover, the abundance of active groups in the polymers which can chelate mineral ions in solution, assures their ready preservation in the rock record. These active groups include the carboxylate, hydroxyl, amine and phosphate groups (Geesey and Jang, 1989). Precipitation of minerals within a microbial biofilm can be influenced by microbial metabolic control of the microenvironment. Furthermore, the presence of an organic template with active nucleation sites also contributes towards bio-catalysed precipitation of minerals. Some of the best-known examples of mineralised biofilms in the geological record are calcified and silicified stromatolites (Krumbein, 1983). Mineralisation of organic templates can occur very rapidly (within a day, Toporski et al., 2001a). Experiments to silicify microorganisms also document the potential faithfulness of reproduction of the original organism, and the fineness of detail obtainable, by silica impregnation (Westall et al., 1995; Westall, 1999; Toporski et al., 2001a).

Bacterial Biofilms in Astrobiology: The Importance of Life Detection

Experience gathered by researchers during their hunt for evidence of early life on Earth has shown the difficulties associated with the interpretation of possible microbial fossils (Schopf and Walter, 1983; Schopf, 1999a; Brazier et al., 2001). Indeed the controversy surrounding the earliest life on Earth is akin to the debate (e.g. Thomas-Keprta, 2000; Golden et al., 2000) on the nature of the nano-structures in Martian meteorite ALH84001 described by McKay et al. (1996). Both these examples emphasise the difficulties involved in (a) conclusively identifying structures as fossil bacterial cells and (b) establishing their indigeneity/syngenicity to the host matrix. Better understanding of biological signatures in rocks is, therefore, needed in order to identify traces of microbial life (Knoll, 1999). These traces include not only the characteristic morphologies of potentially fossilised microorganisms, but also include organic biomarker molecules, isotopic fractionations, biological mineralisation and possibly trace element concentrations. It is thus crucial to tackle the problems emerging from the search for evidence of early life on Earth and in exobiological and — palaeontological research with a multi-disciplinary approach (Knoll, 1999). Techniques traditionally applied in the hunt for evidence of early life on Earth included light and electron microscopy for the detection of morphological biomarkers (Schopf, 1999a; Westall, 2000) and gas chromatography — mass spectroscopy (GC-MS) for molecular biomarkers (Peters & Moldowan, 1993). Neither approach however is capable of combining morphological, isotopic and chemical information from individual structures, which is crucial to obtain unambiguous data.

From Microbial Fossils to Astrobiology

Dr. Jan Toporski is a research Fellow in Geoand Astrobiology at the Carnegie Institute of Washington, Geophysical Laboratory, Washington D.C. He obtained his Ph.D. (2001) from the University of Portsmouth, School of Earth and Environmental Sciences, Portsmouth, UK. His current research focuses on the mineralization, preservation and detection of bacterial biomarkers. He obtained the Gerry Soffen Award (2001) by Mary Ann Liebert Publishing Inc for outstanding young investigators in Astrobiology.