Extracellular Iron Biomineralization by Photoautotrophic Iron-Oxidizing Bacteria

Iron oxidation at neutral pH by the phototrophic anaerobic iron-oxidizing bacterium Rhodobacter sp. strain SW2 leads to the formation of iron-rich minerals. These minerals consist mainly of nano-goethite (α-FeOOH), which precipitates exclusively outside cells, mostly on polymer fibers emerging from the cells. Scanning transmission X-ray microscopy analyses performed at the C K-edge suggest that these fibers are composed of a mixture of lipids and polysaccharides or of lipopolysaccharides. The iron and the organic carbon contents of these fibers are linearly correlated at the 25-nm scale, which in addition to their texture suggests that these fibers act as a template for mineral precipitation, followed by limited crystal growth. Moreover, we evidence a gradient of the iron oxidation state along the mineralized fibers at the submicrometer scale. Fe minerals on these fibers contain a higher proportion of Fe(III) at cell contact, and the proportion of Fe(II) increases at a distance from the cells. All together, these results demonstrate the primordial role of organic polymers in iron biomineralization and provide first evidence for the existence of a redox gradient around these nonencrusting, Fe-oxidizing bacteria.Fe(II) can serve as a source of electrons for phylogenetically diverse microorganisms that precipitate iron minerals as products of their metabolism (see, e.g., references 3, 5, 25, and 30). For example, mixotrophic or autotrophic bacteria can couple the oxidation of Fe(II) to the reduction of nitrate in anoxic and neutral-pH environments. With Fe(III) being highly insoluble at neutral pH, this metabolism leads to the formation of poorly to well-crystallized iron minerals (3, 18, 26, 27) that precipitate partly within the cell periplasm for some strains (22). Similar Fe minerals are also synthesized by autotrophic bacteria that perform anoxygenic photosynthesis, using Fe(II) as an electron donor and light as a source of energy for CO₂ fixation (8, 12, 30), according to the equation HCO₃⁻ + 4 Fe²⁺ + 10 H₂O ⇆ <CH₂O> + 4 Fe(OH)₃ + 7 H⁺.However, the biological mechanisms of iron oxidation in these bacteria and in particular the way they cope with the formation of minerals within their ultrastructures are still not fully understood. Indeed, iron minerals are potentially lethal since their precipitation may alter cellular ultrastructures but also catalyze the production of free radicals (2). Recent genetic studies of the phototrophic, iron-oxidizing bacteria Rhodobacter sp. strain SW2 (6) and Rhodopseudomonas palustris strain TIE-1 (16) have identified genes (fox and pio operons, respectively) encoding proteins specific for iron oxidation. Interestingly, Jiao and Newman (16) suggested that one of these proteins could have a periplasmic localization. However, in contrast to what has been observed in some other phototrophic iron oxidizers (25) and in some nitrate-reducing, iron-oxidizing bacteria (22), no iron-mineral precipitation occurs within the periplasm of the purple nonsulfur iron-oxidizing bacterium Rhodobacter sp. strain SW2 (3). Similarly to some other anaerobic neutrophilic (22, 25) and microaerobic iron-oxidizing bacteria (5, 10), this strain seems indeed to have the ability to localize iron biomineralization at a distance from the cells, leaving large areas of the cells free of precipitates (17, 25). While it has been shown that the Gallionella and Leptothrix genera, for example, produce extracellular polymers that facilitate the nucleation of iron minerals outside cells (see, e.g., references 5 and 9), only a little is known about the existence and function of such polymers in anaerobic, neutrophilic iron-oxidizing bacteria and particularly in the phototrophic strain SW2. In the present study, we investigate iron biomineralization by the photoautotrophic iron-oxidizing bacterium Rhodobacter sp. strain SW2. We use scanning transmission X-ray microscopy (STXM) to map and identify organic polymers produced by the cells as well as the redox state of iron at the 25-nanometer scale regularly during a 2 week-period. These results demonstrate the primordial role of organic polymers in iron biomineralization and provide the first evidence for the existence of a redox gradient around SW2 cells.