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Project Progress

Biological Studies of Hydrothermal Systems (dm)

Task 1. Field Studies and Laboratory Characterization of Hydrothermal Vent Microbes

We have completed a study showing that the hyperthermophilic archaea isolated from the subseafloor are phylogenetlcally and physiologically different from similar organisms isolated from near-by sulfide structures. These results show that the subseafloor in vent systems is a biotope distinct from massive sulfide environments, with unique characteristics and a separate group of indigenous hyperthermophilic species. In a related study, we have described an extremely novel and deeply rooted hyperthermophilic archaea, which we are naming Saganella petroecbolus. This organism has simple nutritional requirements and can grow on organic acids such as acetate and citrate with Fe (III) as the electron acceptor forming magnetite. It can also grow with CO2 and H2. It appears that the normal life-style for Saganella is to attach to mineral surfaces and form biofilms, a characteristic we believe to be canonical for subseafloor microorganisms.

The molecular-phylogenetic analyses of the subsurface bacterial and archaeal communities from the 1998 deep-sea eruption at Axial Volcano, Juan de Fuca Ridge, has been completed for samples collected in 1998, 1999, and 2000. The results from this study show that the subseafloor archaeal community at diffuse-flow vents is a complex mixture of seawater-entrained and indigenous species. Hyperthermophilic and mesophilic methanogens and Thermococcales are clearly indicator-organisms for an anaerobic subseafloor biosphere. The Thermococcus species, while phylogenetically closely related, have markedly different phenotypic characteristics including different protein patterns and enzyme activities. There are also a large number of archaeal sequences that are not found in seawater that appear to be unique to the subseafloor.

A molecular probe has been designed to detect the NIF (nitrogen fixation) gene in archaea and has been used successfully to detect the NIF gene in hyperthermophiles from subseafloor environments. Data from the 2000 cruise to Axial volcano indicate an extremely high diversity of both bacteria and archaea that have the NIF gene. Most of the archaeal NIF genes appear to be associated with methanogens and uncultured marine Crenarchaeota.

A preliminary description of the microbial ecology of active sulfide chimneys has been completed using a combination of molecular and microscopic analyses. Intact microbes have been observed throughout these sulfide structures, including in mineral zones thought to be at temperatures greater than 150°C. This work has also been expanded to include the newly discovered (December 2000) Lost City vent field on the Mid-Atlantic Ridge. This is a unique vent system since the fluids are devoid of sulfide and enriched in carbonate, hydrogen, and methane. The smoker structures consist of carbonates. The preliminary observations indicate unique micro- and macro-faunal communities.

Task 2. Studies of Neutrophilic, Lithotrophic, Fe-oxidizing Bacteria

Iron is potentially one of the Solar System’s most abundant energy supplies, and it is the fourth most abundant element in the Earth’s crust, yet only in the last 10 years has unequivocal evidence been found to support the notion that neutrophilic Fe-oxidizers are truly lithotrophic microorganisms. Our work during the past year has focused on a combination of biodiversity, morphological, and physiological studies aimed at answering fundamental questions about this unique form of metabolism.

â?¢ Biodiversity: We have discovered a rich habitat for Fe-oxidizing microbes in the rhizosphere of wetland plants where microoxic zones around the root mass of wetland plants create an ideal habitat in an otherwise anoxic subsurface for Fe(II) oxidation. As much as 5% of the total population of microbes (approximately 107 cells/cc of root) can be lithotrophic Fe-oxidizers. Interestingly, these root-associated Fe-oxidizers are closely related to other terrestrial and marine Fe-oxidizers.

â?¢ Morphology: A unique feature of Fe-oxidizing bacteria (FeOB) is the signature morphologies of the Fe-oxides formed as a result of their metabolism, normally in the form of a tubular sheath or a helical stalk. We have isolated a strain of FeOB from the Loihi Seamount hydrothermal vent that forms a unique filamentous Fe-oxide. Recent work by several investigators on micro-fossils from different sources (e.g., ancient hydrothermal vent sites or lake beds) and of different ages (up to at least 500 Myr) have shown Fe-silicate structures that look remarkably like the remnants of modern-day Fe-oxidizers.

â?¢ We have been conducting growth studies using a bioreactor that allows precise control of O2, pH, temperature, and Fe(II) concentration to determine the relative portion of Fe-oxidation carried out by the bacteria compared to abiological Fe-oxidation. These measurements are difficult to make, but preliminary estimates are that 30 – 60% of the Fe-oxidation is microbially mediated, the amount depending upon specific conditions of O2 and Fe(II) concentrations.

â?¢ For the past 15 months, we have collected field data on Fe(II), O2, pH, temperature, and conductivity, as well as samples for molecular community analysis, from a high-iron, freshwater aquatic field site that features both low pH (pH 2 – 4) and neutral (pH 5.5 – 7) flow regimens. These data will be useful background information for future field studies aimed at coupling diversity and physiology studies.

Task 3. Possible Origin of Chirality

We initiated studies of the selective adsorption of L- and D-amino acids on calcite â?? work that has implications for the origin of biochemical homochirality. One of life’s most distinctive biochemical signatures is its strong selectivity for chiral molecular species, notably L-amino acids and D-sugars. Prebiotic synthesis reactions, with the possible exception of some interstellar processes, yield essentially equal amounts of L- and D-enantiomers. A significant challenge in origin-of-life research, therefore, is to identify natural mechanisms for the homochiral selection, concentration, and polymerization of molecules from an initially racemic mixture. Symmetry breaking on a chirally-selective mineral surface in an aqueous environment offers a viable scenario for the origin of life. We demonstrated that crystals of the common rock-forming mineral calcite (CaCO3), when immersed in a racemic aspartic acid solution, display significant adsorption and chiral selectivity of D- and L-enantiomers on pairs of mirror-related crystal growth surfaces. This selective adsorption is greater on crystals with terraced surface textures, which suggests that D- and L-aspartic acid concentrate along step-like, linear growth features. Selective adsorption of D- and L-amino acids on calcite is thus a plausible geochemical mechanism for the chiral selection and subsequent homochiral polymerization of amino acids on the prebiotic Earth.

Task 4. Emergence

We have been investigating the role of emergence in the origin of life. Natural systems with many interacting components, such as atoms, molecules, cells or stars, often display complex, emergent behavior not associated with their individual components. In some instances, as in the emergence of turbulent flow in fluids, the solid-state properties of crystals, or the periodic spacing of sand dunes, such complex behavior can be modeled a posteriori when appropriate interaction parameters have been determined. Other phenomena, such as the emergence of consciousness from collections of neurons or the emergence of social behavior from collections of humans, are, at least for the present, less amenable to quantitative analysis. The geochemical origin of life may be modeled as a sequence of “emergent” events, each of which adds to molecular complexity and order. Each of these steps, if properly formulated, should be amenable to experimental study. Each emergent step, furthermore, may result in characteristic isotopic, molecular, and structural “fossils” that might be measured in extraterrestrial environments that have not been subjected to reworking by biological activity.

The observed emergent behavior of highly ordered systems, including galaxies, planets, and life, points to a universal organizing principal. Natural systems tend to develop local-scale order â?? non-equilibrium regions of spontaneously increased free energy and decreased entropy â?? even as global-scale entropy increases. This universal tendency for systems to display increased order at an energy-rich interface, while consistent with the first and second laws of thermodynamics, is not formally addressed in either of those laws; indeed, some researchers have proposed that this behavior should be systematized in a “fourth law of thermodynamics.”

If the chemical evolution of life occurred as a sequence of successively more complex stages of emergence, then the divide between non-life and life may ill-defined. One might establish a hierarchy of emergent properties â?? a progressive sequence that leads, for example, through a number of steps from a pre-biotic ocean enriched in organic molecules, to a cluster of molecules arranged on a mineral surface, to self-replicating molecular systems, to encapsulation and eventually prokaryotic life. The exact nature and sequence of these steps may vary in different environments, but any definition that distinguishes between non-living and living systems of necessity becomes more arbitrary as the number of discrete emergent steps to life increases.

The concept of a sequence of discrete emergent steps is useful and appealing in experimental and theoretical studies of the origin of life for at least two pragmatic reasons. First, a progression of steps reduces an immensely complex historical process to a succession of several, more manageable chemical episodes. Each step becomes a focused process for laboratory experimentation or theoretical modeling. Second, each of these steps may result in distinctive, measurable isotopic, molecular, and structural signatures. As we search for life elsewhere in the universe, we may thus be able to characterize extraterrestrial environments according to their degree of emergence along this multi-step path. In this context, it is useful to review experimental programs that attempt to elucidate a few of the possible geochemical steps in the emergence of life.