Function and Adaptation of Acidophiles in Natural and Applied Communities

Abstract: Acidophiles are organisms that have evolved to grow optimally at high concentrations of protons. Members of this group are found in all three domains of life, although most of them belong to the Archaea and Bacteria. As their energy demand is often met chemolithotrophically by the oxidation of basic ions and molecules such as Fe2+, H2, and sulfur compounds, they are often found in environments marked by the natural or anthropogenic exposure of sulfide minerals. Nonetheless, organoheterotrophic growth is also common, especially at higher temperatures. Beside their remarkable resistance to proton attack, acidophiles are resistant to a multitude of other environmental factors, including toxic heavy metals, high temperatures, and oxidative stress. This allows them to thrive in environments with high metal concentrations and makes them ideal for application in so-called biomining technologies.The first study of this thesis investigated the iron-oxidizer Acidithiobacillus ferrivorans that is highly relevant for boreal biomining. Several unresolved nodes of its sulfur metabolism were elucidated with the help of RNA transcript sequencing analysis. A model was proposed for the oxidation of the inorganic sulfur compound tetrathionate. In a second paper, this species’ transcriptional response to growth at low temperature was explored and revealed that At. ferrivorans increases expression of only very few known cold-stress genes, underlining its strong adaptation to cold environments.Another set of studies focused on the environmentally friendly metal winning technology of bioleaching. One of the most important iron-oxidizers in many biomining operations is Leptospirillum ferriphilum. Despite its significance, only a draft genome sequence was available for its type strain.Therefore, in the third paper of this thesis we published a high quality, closed genome sequence of this strain for future use as a reference, revealing a previously unidentified nitrogen fixation system and improving annotation of genes relevant in biomining environments. In addition, RNA transcript and protein patterns during L. ferriphilum’s growth on ferrous iron and in bioleaching culture were used to identify key traits that aid its survival in extremely acidic, metal-rich environments. The biomining of copper from chalcopyrite is plagued by a slow dissolution rate, which can reportedly be circumvented by low redox potentials. As conventional redox control is impossible in heap leaching, paper four explored the possibility of using differentially efficient iron oxidizers to influence this parameter. The facultative heterotrophic Sulfobacillus thermosulfidooxidans was identified as maintaining a redox potential of ~550 mV vs Ag/AgCl, favorable for chalcopyrite dissolution,while L. ferriphilum caused the potential to raise far above this critical value. RNA transcript analysis was used to identify genomic features that may contribute to this behavior.Lastly, six fields in Northern Sweden were examined for the presence of acid sulfate soils in the fifth paper. The study revealed three acid sulfate soils. The presence of acidophiles that likely catalyze the production of acid in the soil was confirmed by community 16S gene amplicon analysis. One site that was flooded in a remediation attempt and is therefore anoxic still exhibited similar bacteria, however, these now likely grow via ferric iron reduction. This process consumes protons and could explain the observed rise in pH at this site.This thesis examines acidophiles in pure culture, as well as natural and designed communities. Key metabolic traits involved in the adaptation to their habitats were elucidated, and their application in mining operations was discussed. Special attention was paid to acidophiles in chalcopyrite bioleaching and in cold environments, including environmental acid sulfate soils in Northern Sweden.