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.

The weathering of shales is a globally important process affecting both natural and built environments. Shales form roughly 70 % of worldwide sedimentary rock deposits and therefore the weathering of these rocks has substantial effects on the geochemical cycling of elements such as carbon, iron and sulfur. Microbes have been shown to play a key role in weathering shales, primarily through the oxidation of the iron and sulfur of embedded pyrite and the resultant production of sulfuric acid. Despite significant interest in the microbial weathering of shales within industrial sectors such as biohydrometallurgy and civil engineering, comparatively few studies have investigated microbial shale weathering in natural environments. Furthermore, the role of microbes in natural shale weathering processes beyond iron oxidation has largely remained unexplored. In this thesis, the weathering capabilities of microbial communities from natural weathered shale was investigated. The North Yorkshire coastline was used as a study location, due to the abundance and diversity of natural cliffs and historic, disused industrial sites. Cliff erosion and recession on the North Yorkshire coastline is a major concern for local authorities and is the focus of current research. The aim of this work has been to evaluate microbial shale weathering processes within these environments, and hypothesise the possible contribution they may have to erosive processes. Phenotypic plate assays inoculated with weathered shale material were used to obtain rock weathering bacterial isolates that tested positive for a specific weathering phenotype, such as iron oxidation or siderophore production. Subsequent 16S rRNA sequencing enabled genera level identification, revealing 15 genera with rock weathering capabilities with several being associated with multiple weathering phenotypes including Aeromonas sp., Pseudomonas sp. and Streptomyces sp. Shale enrichment liquid cultures were incubated with shale rock chips to simulate natural biological weathering conditions, and the concentration of rock-leached elements in the fluid measured. No evidence of microbially-enhanced leaching was found consistently for any element, however the significant reduction in leachate iron concentration under biological conditions indicates that iron precipitation occurred via microbial iron oxidation. Enrichment cultures inoculated with weathered shale and containing organic matter (OM) rich rocks in water or M9 medium, both liquids lacking an organic carbon source, were grown over several months. The cultures yielded microbial isolates that could utilise rock bound OM sources and one bacterial isolate, Variovorax paradoxus, was taken forward for ecophysiological study. The shale rock that the organism was isolated from, along with other OM rich rocks (mudstones and coals), elicited complex responses from V. paradoxus including enhanced growth and motility. Finally, mineral microcosms in vitro and mesocosms in situ investigated microbial colonization and weathering of shale-comprising minerals (albite, calcite, muscovite, pyrite and quartz). Microcosms were established using iron oxidizing enrichment cultures, as based on the results of the simulated rock weathering experiments, while the in situ mesocosms were buried within weathered shale scree within a disused mine level. Levels of colonization significantly varied between minerals within the microcosms (pyrite > albite, muscovite > quartz > calcite). Although differences in mineral colonization were seen in the mesocosms, they did not match those in the microcosms and were not statistically significant. Pyrite incubated in the microcosms became significantly weathered, with extensive pit formation across the mineral surface that is consistent with microbial iron oxidation. In the mesocosms, pit formation was not identified on pyrite surfaces but dark etchings into the pyrite surface were found underneath fungi hyphal growth. The results of this thesis highlights that a range of microbial rock weathering mechanisms are abundant across weathered shale environments. Microbial iron oxidizing activity was a dominant biogeochemical process that altered rock-fluid geochemistry and weathered pyrite surfaces. However, the impact on rock or mineral weathering of other microbial mechanisms was not elucidated by this work. Given the known capabilities of these mechanisms, the conditions under which they are active may not have been met within the experimental setup used. Microbial iron oxidation in shale and shale-derived materials has previously been demonstrated to weaken rock structure through acid production and secondary mineral formation. From the results of this thesis, it is clear that microbial iron oxidation is an active process within some of the weathered shale environments studied, including cliff surfaces. Therefore, it can be hypothesised that microbial activity could play a role in structurally weakening shale rock within cliffs and accelerate their erosion. Future work should attempt to quantify the rate and extent of microbial iron oxidizing activity within shale cliff environments and investigate its contribution to erosive processes.

Les principaux objectifs étaient d'évaluer la stabilité de l'environnement des scories métallurgiques de Cu résultant de différentes périodes d'activités industrielles et de différentes technologies de fusion. Parmi les scories étudiées, on retrouve: les scories historiques cristallines (SH) ainsi que modernes: les scories de four vertical (SFS), les scories granulées (GS) et les scories de plomb (LS). Les différentes approches adoptées dans ce travail de thèse ont tenu compte de: i) la composition chimique et la phase minérale des scories, ii) la sensibilité à la lixiviation des scories sous l’exposition à différentes conditions de pH en mode statique, iii) l’altération des scories sous exposition aux acides organiques couramment trouvés dans l'environnement du sol, iv ) la bio-altération des scories par les bactéries (Pseudomonas aeruginosa) et v) l’application future de la récupération des métaux provenant des scories étudiées en mettant en œuvre la méthode de lixiviation biologique. Résultats cruciaux: Les résultats des tests de lixiviation dépendant du pH ont montré une libération de métal plus élevée dans des conditions fortement acides (pH 2 et 4), alors que la lixiviation dans des conditions alcalines (pH 10.5) était moins importante pour toutes les scories analysées. L'effet de l’altération par le sol a été démontré, la dissolution des scories est notamment sensible à la présence d'exsudats racinaires artificiels (ARE), d’acides humiques (HA) et d’acides fulviques (FA), la contribution des ARE étant la plus forte. Selon les données recueillies, la dissolution relative des scories est strictement liée à leurs caractéristiques (composition chimique et minéralogique) en fonction des différentes conditions étudiées. L'étude concernant l’effet de l’altération biologique a révélé que Pseudomonas aeruginosa améliore considérablement la libération des éléments majeurs (Si et Fe) et métalliques (Cu, Zn, Pb) par rapport aux effets des facteurs abiotiques, indépendamment de la chimie et de la structure des scories. En outre, une récupération élevée (jusqu'à 90%) des métaux (Cu, Zn, Fe) pourrait être obtenue grâce à la lixiviation avec Acidithiobacillus thiooxidans dans des conditions de laboratoire. Conclusions générales : La stabilité des scories dans l'environnement dépend à la fois des caractéristiques chimiques et de la minéralogie. Cependant, les phases minérales hébergeant les métaux sont les facteurs les plus déterminants concernant l'intensité de la lixiviation des métaux. Pour cette raison, l'examen individuel du comportement des scories est important pour empêcher la contamination de l'environnement et devrait être considéré comme une priorité pour la gestion durable des scories. L’optimisation des paramètres de fonctionnement pour le biolessivage et le développement de la technologie à l'échelle industrielle pourrait permettre une bien meilleure gestion (voir l’exploitation) des scories métallurgiques de Cu / Problem statement: Copper pyrometallurgical slags are inevitable waste by-products of Cu smelting operations. These waste are considered to be important due to their production volume and high residual metal content that are inefficiently recovered during industrial process. Due to the lack of sustainable practices in the past, tremendous volumes of Cu-slags have been disposed in many industrial districts, regardless of the weathering and associated environmental risk. Consequently, there are many areas where slags have been proven to be a source of metallic pollution for the surrounding environment. At the present time, the outstanding contradiction between the sustainable development and environmental pollution encourages to undertake the action regarding this aspect. For this reason, slags are currently being used as supplementary materials for civil engineering purposes (e.g. cement and concrete additives, road bed filling materials, hydraulic construction materials) rather than disposed. Additionally, modern-day management strategies require slags to be thoroughly evaluated with respect to their environmental stability prior undertaking any reuse action. Main objectives were to evaluate environmental stability of Cu-metallurgical slags resulting from different periods of industrial activities and different smelting technologies. Those included: historical crystalline slag (HS) as well as modern: shaft furnace slag (SFS), granulated slag (GS) and lead slag (LS). Different approaches undertaken in this PhD work considered: i) chemical and mineral phase compositions of slags, ii) leaching susceptibility of slags under exposure to different pH-stat conditions, iii) slags weathering under exposure to organic acids commonly found in soil environment, iv) bacterially (Pseudomonas aeruginosa) mediated weathering of slags and v) future application of studied slags for metal recovery by implementing the bioleaching method. Crucial results: The results of the pH-dependent leaching tests showed a higher metal release in strong acidic conditions (pH 2 and 4), whereas leachability at alkaline conditions (pH 10.5) revealed a lower importance for all the slags analyzed. The study considering soil weathering scenario demonstrated that Cu-slags are susceptible to dissolution in the presence of artificial root exudates (ARE), humic (HA) and fulvic acids (FA), whereby ARE were found to have stronger contribution than HA and FA. According to data collected, the different behavior of individual slags is strictly related to their characteristics (chemical and phase composition) reflecting various susceptibilities to dissolution under the investigated conditions. The study considering bio-weathering scenario revealed that Pseudomonas aeruginosa considerably enhances the release of major (Si and Fe) and metallic (Cu, Zn, Pb) elements compared to the effects of abiotic factors, regardless of the slags chemistry and structure. Furthermore, a high gain (up to 90%) of metals (Cu, Zn, Fe) could be credited to bioleaching with Acidithiobacillus thiooxidans under laboratory conditions. General conclusions: The environmental stability of slags depends on both, their bulk chemistry and mineralogy. However, mineral phases harbouring the metals are the key players in metal leachability intensity. For, this reason consideration of individual slags behaviour is important for preventing environmental contamination and should be regarded as priority branch of sustainable slag management. Optimization of operating parameters for bioleaching following development of industrial scale technology is an incentive scheme for future management of Cu-metallurgical slags