Matthew F. Kirk

THE THERMODYNAMIC LADDER IN GEOMICROBIOLOGY

A tenet of geomicrobiology is that anaerobic life in the subsurface arranges itself into zones, according to a thermodynamic ladder. Iron reducers, given access to ferric minerals, use their energetic advantage to preclude sulfate reduction. Sulfate reducers exclude methanogens in the same way, by this tenet, wherever the environment provides sulfate. Examining usable energy—the energy in excess of a cells internal stores—in subsurface environments, we find that in groundwater of near neutral pH the three functional groups see roughly equivalent amounts. Iron reducers hold a clear energetic advantage under acidic conditions, but may be unable to grow in alkaline environments. The calculations fail to identify a fixed thermodynamic hierarchy among the groups. In long-term bioreactor experiments, usable energy did not govern microbial activity. Iron reducers and sulfate reducers, instead of competing for energy, entered into a tightly balanced mutualistic relationship. Results of the study show thermodynamics does not invariably favor iron reducers relative to sulfate reducers, which in turn do not necessarily have an energetic advantage over methanogens. The distribution of microbial life in the subsurface is controlled by ecologic and physiologic factors, and cannot be understood in terms of thermodynamics alone.

Variation in energy available to populations of subsurface anaerobes in response to geological carbon storage.

Microorganisms can strongly influence the chemical and physical properties of the subsurface. Changes in microbial activity caused by geological CO(2) storage, therefore, have the potential to influence the capacity, injectivity, and integrity of CO(2) storage reservoirs and ultimately the environmental impact of CO(2) injection. This analysis uses free energy calculations to examine variation in energy available to Fe(III) and SO(4)(2-) reducers and methanogens because of changes in the bulk composition of brine and shallow groundwater following subsurface CO(2) injection. Calculations were performed using data from two field experiments, the Frio Formation experiment and an experiment at the Zero Emission Research and Technology test site. Energy available for Fe(III) reduction increased significantly during CO(2) injection in both experiments, largely because of a decrease in pH from near-neutral levels to just below 6. Energy available to SO(4)(2-) reducers and methanogens varied little. These changes can lead to a greater rate of microbial Fe(III) reduction following subsurface CO(2) injection in reservoirs where Fe(III) oxides or oxyhydroxides are available and the rate of Fe(III) reduction is limited by energy available prior to injection.

Effect of Permeable Biofilm on Micro- And Macro-Scale Flow and Transport in Bioclogged Pores

Simulations of coupled flow around and inside biofilms in pores were conducted to study the effect of porous biofilm on micro- and macro-scale flow and transport. The simulations solved the Navier-Stokes equations coupled with the Brinkman equation representing flow in the pore space and biofilm, respectively, and the advection-diffusion equation. Biofilm structure and distribution were obtained from confocal microscope images. The bulk permeability (k) of bioclogged porous media depends on biofilm permeability (kbr) following a sigmoidal curve on a log-log scale. The upper and lower limits of the curve are the k of biofilm-free media and of bioclogged media with impermeable biofilms, respectively. On the basis of this, a model is developed that predicts k based solely on kbr and biofilm volume ratio. The simulations show that kbr has a significant impact on the shear stress distribution, and thus potentially affects biofilm erosion and detachment. The sensitivity of flow fields to kbr directly translated to effects on the transport fields by affecting the relative distribution of where advection and diffusion dominated. Both kbr and biofilm volume ratio affect the shape of breakthrough curves.

Solute Concentrations Influence Microbial Methanogenesis in Coal-bearing Strata of the Cherokee Basin, USA.

Microorganisms have contributed significantly to subsurface energy resources by converting organic matter in hydrocarbon reservoirs into methane, the main component of natural gas. In this study, we consider environmental controls on microbial populations in coal-bearing strata of the Cherokee basin, an unconventional natural gas resource in southeast Kansas, USA. Pennsylvanian-age strata in the basin contain numerous thin (0.4–1.1 m) coalbeds with marginal thermal maturities (0.5–0.7% Ro) that are interbedded with shale and sandstone. We collected gas, water, and microbe samples from 16 commercial coalbed methane wells for geochemical and microbiological analysis. The water samples were Na–Cl type with total dissolved solids (TDS) content ranging from 34.9 to 91.3 g L−1. Gas dryness values [C1/(C2 + C3)] averaged 2640 and carbon and hydrogen isotope ratios of methane differed from those of carbon dioxide and water, respectively, by an average of 65 and 183‰. These values are thought to be consistent with gas that formed primarily by hydrogenotrophic methanogenesis. Results from cultivation assays and taxonomic analysis of 16S rRNA genes agree with the geochemical results. Cultivable methanogens were present in every sample tested, methanogen sequences dominate the archaeal community in each sample (avg 91%), and few archaeal sequences (avg 4.2%) were classified within Methanosarcinales, an order of methanogens known to contain methylotrophic methanogens. Although hydrogenotrophs appear dominant, geochemical and microbial analyses both indicate that the proportion of methane generated by acetoclastic methanogens increases with the solute content of formation water, a trend that is contrary to existing conceptual models. Consistent with this trend, beta diversity analyses show that archaeal diversity significantly correlates with formation water solute content. In contrast, bacterial diversity more strongly correlates with location than solute content, possibly as a result of spatial variation in the thermal maturity of the coalbeds.

Mineral influence on microbial survival during carbon sequestration

Geologic carbon sequestration involves the injection of supercritical carbon dioxide into deep saline aquifers. Some of the CO2 dissolves into the brines, perturbing water chemistry and water-rock interactions, and impacting microbial habitat and survival. In this study 3 model organisms were tested for their ability to survive high pressures of CO2 exposure in batch cultures: the gram-negative Shewanella oneidensis (SO) strain MR-1, the gram-positive Geobacillus stearothermophilus (GS), and the methanogenic archaeon Methanothermobacter thermoautitrophicus (MT). Results indicate that GS can survive the highest pressures of CO2 for the longest periods of time while SO is the most sensitive to CO2 toxicity. Survival was then evaluated for SO with various minerals and rocks representative of deep saline aquifers to determine if minerals enhanced survival. Cultures were exposed to 25 bar of CO2 for 2 to 8 h and were plated for viable cell counts. Results show that biofilm formation on the mineral surface is important in protecting SO from the harmful effects of CO2 with quartz sandstones providing the best protection. The release of toxic metals like Al or As from minerals such as clays and feldspars, in contrast, may enhance microbial death under CO2 stress.

Broad-Scale Evidence That pH Influences the Balance Between Microbial Iron and Sulfate Reduction

Understanding basic controls on aquifer microbiology is essential to managing water resources and predicting impacts of future environmental change. Previous theoretical and laboratory studies indicate that pH can influence interactions between microorganisms that reduce ferric iron and sulfate. In this study, we test the environmental relevance of this relationship by examining broad-scale geochemical data from anoxic zones of aquifers. We isolated data from the U.S. Geological Survey National Water Information System for 19 principal aquifer systems. We then removed samples with chemical compositions inconsistent with iron- and sulfate-reducing environments and evaluated the relationships between pH and other geochemical parameters using Spearmans rho rank correlation tests. Overall, iron concentration and the iron-sulfide concentration ratio of groundwater share a statistically significant negative correlation with pH (P < 0.0001). These relationships indicate that the significance of iron reduction relative to sulfate reduction tends to increase with decreasing pH. Moreover, thermodynamic calculations show that, as the pH of groundwater decreases, iron reduction becomes increasingly favorable relative to sulfate reduction. Hence, the relative significance of each microbial reaction may vary in response to thermodynamic controls on microbial activity. Our findings demonstrate that trends in groundwater geochemistry across different regional aquifer systems are consistent with pH as a control on interactions between microbial iron and sulfate reduction. Environmental changes that perturb groundwater pH can affect water quality by altering the balance between these microbial reactions.

Thermodynamic and Kinetic Response of Microbial Reactions to High CO2

Geological carbon sequestration captures CO2 from industrial sources and stores the CO2 in subsurface reservoirs, a viable strategy for mitigating global climate change. In assessing the environmental impact of the strategy, a key question is how microbial reactions respond to the elevated CO2 concentration. This study uses biogeochemical modeling to explore the influence of CO2 on the thermodynamics and kinetics of common microbial reactions in subsurface environments, including syntrophic oxidation, iron reduction, sulfate reduction, and methanogenesis. The results show that increasing CO2 levels decreases groundwater pH and modulates chemical speciation of weak acids in groundwater, which in turn affect microbial reactions in different ways and to different extents. Specifically, a thermodynamic analysis shows that increasing CO2 partial pressure lowers the energy available from syntrophic oxidation and acetoclastic methanogenesis, but raises the available energy of microbial iron reduction, hydrogenotrophic sulfate reduction and methanogenesis. Kinetic modeling suggests that high CO2 has the potential of inhibiting microbial sulfate reduction while promoting iron reduction. These results are consistent with the observations of previous laboratory and field studies, and highlight the complexity in microbiological responses to elevated CO2 abundance, and the potential power of biogeochemical modeling in evaluating and quantifying these responses.

Geochemical Influence on Microbial Communities at CO2-Leakage Analog Sites

Microorganisms influence the chemical and physical properties of subsurface environments and thus represent an important control on the fate and environmental impact of CO2 that leaks into aquifers from deep storage reservoirs. How leakage will influence microbial populations over long time scales is largely unknown. This study uses natural analog sites to investigate the long-term impact of CO2 leakage from underground storage sites on subsurface biogeochemistry. We considered two sites with elevated CO2 levels (sample groups I and II) and one control site with low CO2 content (group III). Samples from sites with elevated CO2 had pH ranging from 6.2 to 4.5 and samples from the low-CO2 control group had pH ranging from 7.3 to 6.2. Solute concentrations were relatively low for samples from the control group and group I but high for samples from group II, reflecting varying degrees of water-rock interaction. Microbial communities were analyzed through clone library and MiSeq sequencing. Each 16S rRNA analysis identified various bacteria, methane-producing archaea, and ammonia-oxidizing archaea. Both bacterial and archaeal diversities were low in groundwater with high CO2 content and community compositions between the groups were also clearly different. In group II samples, sequences classified in groups capable of methanogenesis, metal reduction, and nitrate reduction had higher relative abundance in samples with relative high methane, iron, and manganese concentrations and low nitrate levels. Sequences close to Comamonadaceae were abundant in group I, while the taxa related to methanogens, Nitrospirae, and Anaerolineaceae were predominant in group II. Our findings provide insight into subsurface biogeochemical reactions that influence the carbon budget of the system including carbon fixation, carbon trapping, and CO2 conversion to methane. The results also suggest that monitoring groundwater microbial community can be a potential tool for tracking CO2 leakage from geologic storage sites.

Dissolved fulvic acids from a high arsenic aquifer shuttle electrons to enhance microbial iron reduction

It was demonstrated more than two decades ago that microorganisms use humic substances, including fulvic acid (FA), as electron shuttles during iron (Fe) reduction in anaerobic soils and sediments. The relevance of this mechanism for the acceleration of Fe(III) reduction in arsenic-laden groundwater environments is gaining wider attention. Here we provide new evidence that dissolved FAs isolated from sediment-influenced surface water and groundwater in the Bengal Basin were capable of electron shuttling between Geobacter metallireducens and Fe(III). Moreover, all four Bangladesh sediment-derived dissolved FAs investigated in this study had higher electron accepting capacity (176 to 245μmol/g) compared to aquatic FAs, such as Suwanee River Fulvic Acid (67μmol/g). Our direct evidence that Bangladesh FAs are capable of intermediate electron transfer to Fe(III) supports other studies that implicate electron shuttling by sediment-derived aqueous humics to enhance Fe reduction and, in turn, As mobility. Overall, the finding of greater electron accepting capacity by dissolved FAs from groundwater and other sediment-influenced environments advances our understanding of mechanisms that control Fe reduction under conditions where electron transfer is the rate limiting step.

pH as a Primary Control in Environmental Microbiology: 1. Thermodynamic Perspective

pH influences the occurrence and distribution of microorganisms. Microbes typically live over a range of 3 to 4 pH units and are described as acidophiles, neutrophiles, and alkaliphiles, depending on the optimal pH for growth. Their growth rates vary with pH along bell- or triangle-shaped curve, which reflects pH limits of cell structure integrity and the interference of pH with cell metabolism. We propose that pH can also affect the thermodynamics and kinetics of microbial respiration, which then help shape the composition and function of microbial communities. Here we use geochemical reaction modeling to examine how environmental pH controls the energy yields of common redox reactions in anoxic environments, including syntrophic oxidation, iron reduction, sulfate reduction, and methanogenesis. The results reveal that environmental pH changes the energy yields both directly and indirectly. The direct change applies to the reactions that consume or produce protons whereas the indirect effect, which applies to all redox reactions, comes from the regulation of chemical speciation by pH. The results also show that the energy yields respond strongly to pH variation, which may modulate microbial interactions and help give rise to the pH limits of microbial metabolisms. These results underscore the importance of pH as a control on microbial metabolisms and provide insight into potential impacts of pH variation on the composition and activity of microbial communities. In a companion paper, we continue to explore how the kinetics of microbial metabolisms responds to pH variations, and how these responses control the outcome of microbial interactions, including the activity and membership of microbial consortia.