Fungi play important geoactive roles in the biosphere, particularly element biotransformations and biogeochemical cycling, metal and mineral transformations, decomposition, bioweathering, and soil and sediment formation. Fungi can apply various mechanisms to effect changes in metal speciation, toxicity and mobility, mineral formation and/or mineral dissolution. This research has examined fungal roles in bioweathering and bioleaching of zinc sulfide ore, together with an investigation of the role of fungal phosphatases in the bioprecipitation of uranium and lead when utilising an organic phosphorus-containing substrate as the sole phosphorus source. The results obtained revealed that test fungal species showed bioweathering effects on zinc sulfide ore, and clear evidence of biotransformation and bioleaching of zinc sulfide was obtained after growth of A. niger. The formation of zinc oxalate dihydrate resulted from oxalic acid excretion. The formation of uranium- and lead-containing biominerals after growth of yeasts and filamentous fungi with organic phosphorus sources have also been demonstrated and characterized. Test fungi were capable of precipitating uranium phosphate and pyromorphite, and also produced mycogenic lead oxalate during this process. This work is the first demonstration that filamentous fungi are capable of precipitating a variety of uranium- and lead-containing phosphate biominerals when grown with an organic phosphorus source. The role of fungal processes in the bioweathing and bioleaching of mineral ores, and the significance of phosphatases in the formation of uranium and lead secondary minerals has thrown further light on potential fungal roles in metal and mineral biogeochemistry as well as the possible significance of these mechanisms for element biorecovery or bioremediation.

Microbial communities influence the kinetics and pathways of reactions involved in the dissolution of a number of minerals (Ehrlich 1996). On a smaller scale these interactions can affect substrate permeability, porosity, and create highly localized biogeochemical conditions. However, a mechanistic understanding of the consequences of microbial surface colonization on calcite dissolution rate has yet to be achieved. More specifically, little is known about the impact of sulfur-oxidizing bacteria activity on the rate of carbonate mineral dissolution, or the nature of the microbe-limestone attachment and interaction. Through a series of laboratory and field experiments the effect of mineral surface colonization by microbial communities, obtained from an active sulfuric acid cave (Lower Kane Cave (LKC), Big Horn Basin, WY), on the dissolution rate of Madison Limestone was quantified. Results from laboratory experiments showed that a microbial biofilm, composed primarily of Epsilonproteobacteria and Gammaproteobacteria growing on a limestone surface oxidized thiosulfate and increased carbonate dissolution rates up to 3.3 times faster than abiotic rates. When all thiosulfate substrate was withheld the community oxidized stored intracellular sulfur, continuing to accelerate limestone dissolution and decreasing pH. This process is sensitive to O2 limitations. Characterization of this aggressive sub-biofilm corrosion was more closely examined by SEM imaging. By comparing mineral surface morphology of colonized chips to non-colonized chips of various carbonate substrates, it was shown that even under conditions near equilibrium with calcite, aggressive dissolution of carbonate substratum occurs exclusively beneath the biofilm. These findings support the hypothesis that (1) sulfur-oxidizing microbial communities aggressively dissolve carbonates in order to buffer the production of excess acidity by neutrophilic communities and (2) biofilm presence affects carbonate mineral dissolution by physically separating a bulk stream water from the sub-biomat environment. Furthermore, it was found that mineralogy affects the degree of establishment of microbial communities in this environment. Results from a series of four laboratory and one in situ reactor experiment showed that limestone and dolostone substratum consistently had higher biomass accumulations than silicate minerals or pure Iceland spar calcite in the same reactor. These results provide evidence to support the hypothesis that mineralogy influences microbial accumulation in sulfuric-acid karst systems. Particularly, neutrophilic sulfur-oxidizing communities accumulate in greater quantities on solid substrates that buffer metabolically-generated acidity. These results also demonstrated the dependence of microorganisms on colonization of a particular mineral surface, possibly in order to gain access to micronutrients bound within solid substrates when exposed to nutrient-limited conditions. / text

Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 217-260).

Meteorites were not acknowledged as truly extraterrestrial until in the 19th century. In this report, the ability of organisms to colonize meteorites was examined with classical and molecular biological methods. A table of organisms found on meteorites was constructed from published data and compared with phylogenetic trees based on a molecular biological database containing small subunit ribosomal gene sequences from different meteorite studies. Statistical analyzes between the southern and northern hemisphere were made using Mann-Whitney U test on the data in the table of organisms. Microbiological growth was also examined on the surface of four different meteorites from the desert of Oman. Twenty-one isolates were selected for further characterization of gram-properties, microscopy and fluorescence in situ hybridization (FISH). The table of organisms contains a wide variety of organisms associated with meteorites, e.g. angiosperms, lichens and different types of microorganisms. The phylogenetic trees for all three domains show that most gene sequences obtained from so far published studies belong to several different phyla. The FISH results of own experiments showed that 62 % of the 21 isolates belong to Actinobacteria, 9 % to Firmicutes and 5 % to Betaproteobacteria. The statistical analysis shows that there is no significant difference between the southern and the northern hemisphere. The number of studies is however too low to allow proper conclusions about correlations between meteorites and colonizing organisms, though the results so far suggest that organisms associated with meteorites often represent novel, unknown genera.

Carbonate precipitates in biofilm were investigated from hot springs near Viterbo, Italy; Salt Pond, San Salvador; and Fowl Cay Reef, Abaco, Bahamas. Features shared by hot springs and salt ponds are supersaturation with CaCO3, abundant Spirulina, and clustered acicular aragonite crystals termed “fuzzy dumbbells.” TEM and FESEM microscopy show fuzzy dumbbells contain a core of amorphous organic matter and subhedral CaCO3 microcrystals arranged in linear fabrics. Micron- to millimeter-scale microenvironments are identified by localized dissolution, the occurrence of gothic calcite inter-grown with organic filaments, and the presence of calcite in biofilm where aragonite is chemically favored. Spherical CaCO3 precipitates in reefs were anticipated, but not encountered in TEM sections of reef biofilm. In conclusion, biofilm creates the microenvironment and organic matter provides substrate for fuzzy dumbbell precipitation. TEM is a novel technique for studying the relationship between organic matter and CaCO3 precipitation, and has potential medical, industrial, and academic applications.

Experiments with microbial biofilms in microgravity and simulated microgravity have revealed altered growth kinetics, but geomicrobial biofilms have not yet been studied in low gravity environments. No characterisation of biofilms, geomicrobial or otherwise, have been conducted at hypergravity. This thesis explores factors affecting microbe-mineral interactions under terrestrial conditions, lays the groundwork for a scheduled microgravity experiment, and provides the first data on biofilms grown at hypergravity. As a first step in understanding microbe-mineral interactions in altered gravity environments, experiments were undertaken to identify factors that constrain attachment in a terrestrial environment. The model organism Sphingomonas desiccabilis and basaltic rock from Iceland were selected, and the minerals that make up the basalt were identified and procured in their pure form. The relative significance of physical factors such as hydrophobicity, surface charge, porosity and nutritional value were examined in relationship to the success with which biofilms colonised the mineral surfaces. Growth was measured by the quantity of biofilm biomass after a ifxed time period, using Crystal Violet stain, in order to draw conclusions about the most influential physical conditions on biofilm attachment to a substrate. It was found that mineral attachment is influenced more by porosity and nutritional value than by hydrophobicity or surface charge. To explore how reduced gravity affects biofilm formation and weathering rates, a European Space Agency experiment, BioRock, is underway. Samples of basalt, with monocultures of three different organisms, will be sent to the International Space Station in 2019 for long-term exposure to Martian and micro-gravity. Research testing proof of concepts, material compatibility, and experimental procedure and equipment is described. Confocal laser scanning microscopy (CLSM) was used to image the biofilms, and inductively coupled plasma mass spectroscopy (ICP-MS) experiments were conducted to compare biotic and abiotic elemental release rates from basalt. Both of these methods will be employed for post-flight analysis of BioRock. Preliminary terrestrial ICP-MS experiments indicated that rare Earth elements (REEs) showed the most reliable reflection of leaching patterns overall, as a consequence of their high molecular weight and low volatility during the ashing procedure. To fully understand gravity's effect on microbiological processes it is important to investigate what occurs when its influences are removed, but also to establish what occurs when extra gravitational force is applied. Using simulated hypergravity, achieved through hyper-acceleration on a geotechnical centrifuge, the effects of 10 x g on biofilm development and the leaching of basalt were investigated. As this was the first time that biofilms had been studied under hypergravity, additional substrates were included with the basalt, to enable characterisation of the more general response of biofilms to hypergravity. In contrast to previous experiments conducted on planktonic bacteria, which found decreased population sizes, the biofilms grown at 10 x g showed greater biomass than the 1 x g samples. ICP-MS showed no difference in the average weathering rates, but greater variability in the higher gravity samples. The data collected here advances our understanding of microbial interactions with geologically important substrates, with implications for an ISS microgravity experiment and future human space exploration. It also presents new intelligence on the previously unstudied effects of hypergravity on biofilms and rock weathering.

Viruses are ubiquitous biological entities whose importance and role in aquatic habits is beginning to take form. However, several habitats have undergone limited to no examination with viral-geochemical parameters minimally examined and viral-mineral relationships in the natural environment and the role of mineralization on viral-host dynamic completely lacking. To further develop knowledge on the presence and abundances of viruses, how viruses impact aquatic systems, and how viral-host interactions can be impacted under mineralizing conditions, viruses were examined under a variety of habitats and experimental conditions. Water samples were collected from the deep subsurface (up to 450 m underground) and acid mine drainage (AMD) systems in order to determine the presence, abundance, and viral-geochemical relationships within the systems. Samples were also collected from a variety of freshwater habitats, which have undergone limited examination, to determine viral-geochemical and viral-mineral relationships. Lastly, bacteriophage-host dynamics were examined under authigenic mineral precipitation to determine how mineralization impacts this relationship. Results reveal that not only are viruses present in the deep subsurface and AMD systems, but they are abundant (up to 107 virus-like particles/mL) and morphogically diverse. Viruses are also the strongest predictor of prokaryotic abundance in southern Ontario freshwater systems where potential nutrients are rich. Geochemical variables, such as pH and Eh, were shown to have negative impacts of viral abundance indicting that AMD environments are detrimental for free viruses (i.e. not particle associated). Direct evidence of viral-mineral interactions was found using transmission electron microscopy as viral particles were shown attached to iron-bearing mineral phases (determined through elemental analysis). In addition, evidence of viral participation in mineralization events was found in both AMD and freshwater environments where inverse correlations were noted between viral abundance and jarosite saturation indices (r = -0.71 and r = -0.33, respectively), and goethite saturation indices were also noted to be the strongest predictor of VLP abundance in freshwater habitats explaining 78% of the variability in the data. Lastly, iron precipitation and/or metal ion binding to bacterial surfaces greatly reduced phage replication (~98%) revealing bacterial mineralization has a protective benefit strongly hindering viral replication.

Viruses are ubiquitous biological entities whose importance and role in aquatic habits is beginning to take form. However, several habitats have undergone limited to no examination with viral-geochemical parameters minimally examined and viral-mineral relationships in the natural environment and the role of mineralization on viral-host dynamic completely lacking. To further develop knowledge on the presence and abundances of viruses, how viruses impact aquatic systems, and how viral-host interactions can be impacted under mineralizing conditions, viruses were examined under a variety of habitats and experimental conditions. Water samples were collected from the deep subsurface (up to 450 m underground) and acid mine drainage (AMD) systems in order to determine the presence, abundance, and viral-geochemical relationships within the systems. Samples were also collected from a variety of freshwater habitats, which have undergone limited examination, to determine viral-geochemical and viral-mineral relationships. Lastly, bacteriophage-host dynamics were examined under authigenic mineral precipitation to determine how mineralization impacts this relationship. Results reveal that not only are viruses present in the deep subsurface and AMD systems, but they are abundant (up to 107 virus-like particles/mL) and morphogically diverse. Viruses are also the strongest predictor of prokaryotic abundance in southern Ontario freshwater systems where potential nutrients are rich. Geochemical variables, such as pH and Eh, were shown to have negative impacts of viral abundance indicting that AMD environments are detrimental for free viruses (i.e. not particle associated). Direct evidence of viral-mineral interactions was found using transmission electron microscopy as viral particles were shown attached to iron-bearing mineral phases (determined through elemental analysis). In addition, evidence of viral participation in mineralization events was found in both AMD and freshwater environments where inverse correlations were noted between viral abundance and jarosite saturation indices (r = -0.71 and r = -0.33, respectively), and goethite saturation indices were also noted to be the strongest predictor of VLP abundance in freshwater habitats explaining 78% of the variability in the data. Lastly, iron precipitation and/or metal ion binding to bacterial surfaces greatly reduced phage replication (~98%) revealing bacterial mineralization has a protective benefit strongly hindering viral replication.

Iron oxyhrdroxide exist in a range of crystallinities and subsequent bioavailabilities with the poorly crystalline Fe oxyhrdroxide, ferrihydrite, considered the most bioavailable. Yet, as a result of the instability ferrihydrite it quickly ripens and/or transforms to more thermodynamically stable end-members bringing into question its importance in supporting long-term Fe(III)-reducing microbial communities. Furthermore, while a wide phylogenetic diversity of microorganisms capable of reducing ferrihydrite have been isolated, these organisms show diminished abilities to reduce more stable and dominant crystalline Fe phases. Therefore to address the questions of which microorganisms and what microbial processes are responsible for controlling the reduction of diverse Fe(III) minerals phases, cultivation based approaches using both batch and column-type reactors were employed. Using geochemical and phylogenetic analysis it was revealed that the Fe oxide substrate was important in dictating the mechanisms of Fe(III) reduction, and the structure of the microbial communities. While model dissimilartory Fe reducing microorganisms were capable of reducing ferrihydrite when acetate was provided as a carbon source these organisms did not enrich and were incapable of reducing crystalline Fe(III) oxides. Instead, in enrichments where crystalline Fe(III) oxides were reduced, organisms associated with fermentation and sulfate respiration dominated, this despite using freshwater media low in sulfate (less than 200 µM). In addition, these non-model Fe reducers dominated in ferrihydrite enrichments when carbon compounds other than acetate were given. Interestingly, a strong negative correlation between Fe(III) and sulfate respiration was observed with the canonical thermodynamic view that ferrihydrite should precede sulfate as a terminal electron acceptor being challenged. Further experiments with pure cultures of Desulfovibrio putealis indicated that a catalytic sulfur cycle may be responsible for greater than expected Fe(II) values under low sulfur conditions. These findings, have broad implications in predicting microbially mediated electron flow to oxidized substrates which will dictate the pathways and degree of carbon mineralization and subsequent carbon sequestration within sediments and soils. Further, given the importance of Fe(III)-reducing communities and Fe(II) in the sequestration of both inorganic and organic contaminants, these findings will have direct bearing on contaminant mitigation and remediation. / Engineering and Applied Sciences

Master of Science / Department of Geology / Matthew Kirk / Groundwater from the High Plains aquifer is vital for food production and a growing human population in the Great Plains region of the United States. Understanding how groundwater quality is changing in response to anthropogenic and natural processes is critical to effectively managing this resource. Our study considers variation in groundwater geochemistry in the Great Bend Prairie aquifer, a portion of the High Plains aquifer in southcentral Kansas. We collected samples during summer 2016 from 24 monitoring wells and compared our results to data collected previously from the same wells from 1979 to 1987. We sampled 13 wells screened in the upper portion of the aquifer (avg. depth 72 ft), 10 wells screened near the aquifer base (avg. depth 141 ft), and one well screened in underlying bedrock. Compared to initial samples, samples we collected tended to have higher total dissolved solids (TDS) and nitrate content, particularly those we collected from the upper aquifer. Compared to initial samples, TDS was 78 mg/L higher in samples we collected from the upper aquifer and 373 mg/L lower in samples we collected from the aquifer base on average. Nitrate exceeded the U.S. standard for public supplies of drinking water (10 mg/L as N) in seven of the samples we collected, compared to only two samples collected previously. Compared to previous samples, nitrate concentrations were 9.5 and 3.9 mg/L as N higher on average in samples collected from the upper aquifer and aquifer base, respectively. Based on a mixing analysis, variation in the salinity of our samples primarily reflects the dilution of natural Permian brines by freshwater recharge throughout the area. However, salinity decreases observed in four samples reflects flushing of initial oil brine contamination over time, salinity increases in two samples may be due to evapotranspiration, and salinity increases in two samples may reflect migration of oil-brine contamination towards the site. Stable nitrogen (15N/14N) and oxygen (18O/16O) isotope ratios in our samples primarily fall within the range typical of nitrification of ammonium-based fertilizers with potential contributions from manure or sewage. In our analysis of the microbial community, we observed groups capable of denitrification, including genera within Nitrospirae, Firmicutes, and Proteobacteria. Despite their presence, our results demonstrate that water quality in the aquifer has degraded over the past 30 to 40 years due to nitrate accumulation.