Characterization of NaCl tolerance mechanism and its relation with the antioxidant mechanisms in the acidophilic bacterium Leptospirillum ferriphilum DSM 14647
Chloride bioleaching is considered a promising alternative method to recover copper from chalcopyrite and other primary copper sulfides, because it favors the leaching kinetics and avoids passivation of minerals. Nevertheless, chloride ions are highly toxic for iron-oxidizing microorganisms that par...
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2019 |
| País: | Chile |
| OAI Identifier: | oai:repositorio.anid.cl:10533/246430 |
| Acceso en línea: | https://hdl.handle.net/10533/246430 |
| Access Level: | acceso abierto |
| Palabra clave: | Ciencias Naturales Biotecnología Industrial |
| Sumario: | Chloride bioleaching is considered a promising alternative method to recover copper from chalcopyrite and other primary copper sulfides, because it favors the leaching kinetics and avoids passivation of minerals. Nevertheless, chloride ions are highly toxic for iron-oxidizing microorganisms that participate in the bioleaching process. In addition to the osmotic imbalance, chloride can also induce acidification of the cytoplasm in these microorganisms. We predicted that intracellular acidification produces an increase in respiratory rate and reactive oxygen species generation, and therefore oxidative stress can also be induced. The general goal of this study was to characterize the NaCl tolerance molecular mechanism and establish its relation with the antioxidant mechanism in Leptospirillum ferriphilum DSM 14647. First, the participation of canonical systems of tolerance to osmotic stress and the antioxidant systems, as an early response mechanism, were studied. By bioinformatic analysis, it was determined that genes for a complete or partial repertoire of K+ transporters, the biosynthesis pathways and transporter for compatible solutes (hidroxi)ectoine and trehalose were found in most of the acidophilic iron-oxidizing microorganisms. Additionally, the exposition of L. ferriphilum to 100 mM NaCl immediately up-regulated kdpC and kdpD genes coding for potassium transporters. A prolonged exposure to NaCl also increased the expression of genes encoding for biosynthesis of compatible solutes (hydroxy)ectoine (ectC and ectD) and trehalose (otsB). As a consequence, the intracellular levels of both hydroxyectoine and trehalose increased significantly, suggesting a strong response to keep osmotic homeostasis. On the other hand, the intracellular pH significantly decreased from 6.7 to pH 5.5 and oxygen consumption increased significantly when the cells were exposed to NaCl stress. Furthermore, this stress condition led to a significant increase of the intracellular content of reactive oxygen species, and to a rise of the antioxidative cytochrome c peroxidase (CcP) and thioredoxin (Trx) activities. In agreement with these results, ccp and trx genes were up-regulated under this condition, suggesting that this bacterium displays a transcriptionally regulated response against oxidative stress induced by chloride. In parallel, L. ferriphilum was adapted to 180 mM NaCl to identify the late response strategy. The analysis by transcriptomic profile revealed that the principal mechanisms involved in the adaptation were related with genes associated to the cell membrane integrity, respiration and antioxidant proteins, probably to conserve the pH and redox homeostasis. Inspection of these parameters in the adapted culture proved an increase in the respiratory rate and the maintain of the intracellular ROS levels. On contrary, genes associated with biosynthesis of hydroxyectoine (ectB, ectC, ectD) were repressed, coincident with the lack of detection of this compound in cell extract. Thus, these data suggest that the cells were not under osmotic stress. Finally, according with these results, we were able to conclude that chloride has a dramatic multifaceted effect on acidophile physiology that involves osmotic, acidic and oxidative stresses. The early response mechanism was composed by the osmotic response, pH homeostasis by cell respiration and antioxidant response. Instead, the late response mechanism involved pH homeostasis and antioxidant response. |
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