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In extremely acidic environments, ferric iron can be a thermodynamically favorable electron acceptor during elemental sulfur oxidation by some Acidithiobacillus spp. under anoxic conditions. Quantitative 2D-PAGE proteomic analysis of a resting cell suspension of a sulfur-grown Acidithiobacillus ferrooxidans CCM 4253 subculture that had lost its iron-reducing activity revealed 147 protein spots that were downregulated relative to an iron-reducing resting cell suspension of the antecedent sulfur-oxidizing culture and 111 that were upregulated. Tandem mass spectrometric analysis of strongly downregulated spots identified several physiologically important proteins that apparently play roles in ferrous iron oxidation, including the outer membrane cytochrome Cyc2 and rusticyanin. Other strongly repressed proteins were associated with sulfur metabolism, including heterodisulfide reductase, thiosulfate:quinone oxidoreductase and sulfide:quinone reductase. Transcript-level analyses revealed additional downregulation of other respiratory genes. Components of the iron-oxidizing system thus apparently play central roles in anaerobic sulfur oxidation coupled with ferric iron reduction in the studied microbial strain., (Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

In contrast to iron-oxidizing Acidithiobacillus ferrooxidans, A. ferrooxidans from a stationary phase elemental sulfur-oxidizing culture exhibited a lag phase in pyrite oxidation, which is similar to its behaviour during ferrous iron oxidation. The ability of elemental sulfur-oxidizing A. ferrooxidans to immediately oxidize ferrous iron or pyrite without a lag phase was only observed in bacteria obtained from growing cultures with elemental sulfur. However, these cultures that shifted to ferrous iron oxidation showed a low rate of ferrous iron oxidation while no growth was observed. Two-dimensional gel electrophoresis was used for a quantitative proteomic analysis of the adaptation process when bacteria were switched from elemental sulfur to ferrous iron. A comparison of total cell lysates revealed 39 proteins whose increase or decrease in abundance was related to this phenotypic switching. However, only a few proteins were closely related to iron and sulfur metabolism. Reverse-transcription quantitative PCR was used to further characterize the bacterial adaptation process. The expression profiles of selected genes primarily involved in the ferrous iron oxidation indicated that phenotypic switching is a complex process that includes the activation of genes encoding a membrane protein, maturation proteins, electron transport proteins and their regulators.

Elemental sulfur oxidation by ferric iron in Acidithiobacillus ferrooxidans was investigated. The apparent Michaelis constant for ferric iron was 18.6 mM. An absence of anaerobic ferric iron reduction ability was observed in bacteria maintained on elemental sulfur for an extended period of time. Upon transition from ferrous iron to elemental sulfur medium, the cells exhibited similar kinetic characteristics of ferric iron reduction under anaerobic conditions to those of cells that were originally maintained on ferrous iron. Nevertheless, a total loss of anaerobic ferric iron reduction ability after the sixth passage in elemental sulfur medium was demonstrated. The first proteomic screening of total cell lysates of anaerobically incubated bacteria resulted in the detection of 1599 protein spots in the master two-dimensional electrophoresis gel. A set of 59 more abundant and 49 less abundant protein spots that changed their protein abundances in an anaerobiosis-dependent manner was identified and compared to iron- and sulfur-grown cells, respectively. Proteomic analysis detected a significant increase in abundance under anoxic conditions of electron transporters, such as rusticyanin and cytochrome c(552), involved in the ferrous iron oxidation pathway. Therefore we suggest the incorporation of rus-operon encoded proteins in the anaerobic respiration pathway. Two sulfur metabolism proteins were identified, pyridine nucleotide-disulfide oxidoreductase and sulfide-quinone reductase. The important transcription regulator, ferric uptake regulation protein, was anaerobically more abundant. The anaerobic expression of several proteins involved in cell envelope formation indicated a gradual adaptation to elemental sulfur oxidation.