Analysis of the molecular basis of chloride ion tolerance in acidophilic bacteria used in bioleaching

Author(s)

Thiago Miranda da Silva

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2016-05-07

Examining board members

Laura Maria Mariscal Ottoboni; Edi Lúcia Sartorato; Sueli Matilde da Silva Costa; Edmilson Ricardo Gonçalves; Salete Aparecida Gaziola

• Advisor: Laura Maria Mariscal Ottoboni; Tatiana Teixeira Torres

Abstract

Acidithiobacillus ferrooxidans is a Gram-negative, chemolitotrophic and acidophilic bacterium, important in metal bioleaching processes. However, its utilization is limited in bioleaching operations where only saline water is available for the process due to the bacterial sensitivity to NaCl. This way, the comprehension of the molecular mechanisms involved in salt stress in the bacterium is of great interest for the biomining industry. Taking this into consideration, the aim of this work was to analyze the salt (NaCl) stress in A. ferrooxidans cells for short (salt-shock cells) and long (salt-30 cells) periods of exposition. The RAMAN spectroscopy analysis was performed with A. ferrooxidans salt-shock and salt-30 cells and the results showed several changes in membrane components corresponding to proteins, lipids and carbohydrates. These alterations were observed in both salt-shock and salt-30 cells, showing that cells are affected either by short or long periods of exposure to salt. The proteome analysis showed that the A. ferrooxidans salt-30 cells increased the expression of several membrane related proteins ¿ structural and transport proteins ¿ possibly as a defense mechanism against salt stress. Also, in these cells, the expression of stress-response related proteins, proteases and transcription regulation proteins increased suggesting that the long-term salt stress affects several cellular processes. The expression of proteins related to carbon metabolism, ATP production and transcription processes, among others, decreased in the salt-30 cells. The RNA sequencing was performed with A. ferrooxidans salt-shock cells aiming to evaluate salt stress effects in A. ferrooxidans for short periods. The results showed an alteration in the expression of several genes that encode proteins related to cellular membrane and its permeability, highlighting the importance of the bacterial membrane as a defense mechanism against salt stress. The down-regulation of several genes that encode proteins related to energy production, cellular division and protein synthesis, contrasting with the up-regulation of genes that encode for transposases and transcription processes, suggest that the presence of NaCl affects the salt-shock cells metabolism and growth. Also, in this work, the genome of a salt-tolerant bacterium, Thiobacillus prosperus DSM 14174, was sequenced. The sequencing revealed that the bacterium possess several genes responsible for salt tolerance that are no present in A. ferrooxidans. Among these genes, are the ones that encode for membrane transporters of chloride ions and other ions, and transport and biosynthesis of osmoprotectants. A phylogenetic and phylogenomic analyses indicated that T. prosperus DSM 14174 is a member of the Ectothiorhodospiraceae family, composed of halophilic bacteria. This bacterium belongs to the same genus of Acidihalobacter prosperus DSM 5130. However, they do not belong to the same species (AU)

• FAPESP's process: 10/12476-1 - Molecular basis analysis of chloride ion tolerance in acidophile bacteria used in bioleaching
• Grantee: Thiago Miranda da Silva
• Support Opportunities: Scholarships in Brazil - Doctorate