Biomineralisation of calcite by bacterial isolates

Keiner, Robert; Frosch, Torsten; Hanf, Stefan; Rusznyak, Anna; Akob, Denise M; Küsel, Kirsten; Popp, Jürgen

Raman Spectroscopy—An Innovative and Versatile Tool To Follow the Respirational Activity and Carbonate Biomineralization of Important Cave Bacteria Journal Article

Analytical Chemistry, 85 (18), pp. 8708–8714, 2013.

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Rusznyák, Anna; Akob, Denise M; Nietzsche, Sándor; Eusterhues, Karin; Totsche, Kai Uwe; Neu, Thomas R; Frosch, Torsten; Popp, Jürgen; Keiner, Robert; Geletneky, Jörn; Katzschmann, Lutz; Schulze, Ernst-Detlef; Küsel, Kirsten

Calcite Biomineralization by Bacterial Isolates from the Recently Discovered Pristine Karstic Herrenberg Cave Journal Article

Applied and Environmental Microbiology, 78 (4), pp. 1157–1167, 2012.

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Using stalactite material and fluvial sediments from the Blessberg cave, the diversity and activity of the occuring bacteria and their formation of carbonate minerals were investigated. For this purpose, various techniques such as phylogenetic analyses, bacterial cultivation, electron microscopy, X-ray spectroscopy, confocal laser scanning microscopy, and Raman spectroscopy were used.

Bacteria were found on the surface and inside the stalactites using confocal laser scanning microscopy. Phylogenetic analyses showed that Proteobacteria were most abundant in the bacterial communities on and inside the stalactites and also in the fluvial sediments, but also other groups such as Actinobacteria or FirmicutesFirmicutes Eine artenreiche Gruppe innerhalb der Bakterien. Unterscheiden sich von den Actinobakterien unter anderem durch ihren niedrigeren Gehalt an Nukleinbasen in der Bakterien-DNS.. Many of the detected bacteria have not yet been cultivated.

A total of nine new bacterial cultures were isolated from the cave sediments, growing on alkaline carbonate-precipitating medium and belonging to the bacterial genera Arthrobacter, Flavobacterium, Pseudomonas, Rhodococcus, Serratia and Stenotrophomonas. Of these, the two with the most intense precipitate formation were selected for further research: Arthrobacter sulfonivorans SCM3 and Rhodococcus globerulus SCM4. Both produced extracellular polymeric substances (EPS) and grew as cell aggregates.

Scanning electron microscopy of cell aggregates and extracellular polymeric substance (EPS) of the isolates Arthrobacter sulfonivorans SCM3 and Rhodococcus globerulus SCM4
Scanning electron microscopy of cell aggregates and extracellular polymeric substance (EPS) of the isolates Arthrobacter sulfonivorans SCM3 (left) and Rhodococcus globerulus SCM4 (right) (Image: Elektronenmikroskopisches Zentrum Jena, Sándor Nietzsche and Anna Rusznyak).

The carbonate minerals formed were mixtures of calcite, vaterite, and monohydrocalcite. Arthrobacter sulfonivorans SCM3 formed xenomorphic spherical crystals, and Rhodococcus globerulus SCM4 idiomorphic crystals with rhombohedral morphology.

The biomineralisation process of Arthrobacter sulfonivorans SCM3 was further investigated using a combination of Raman macro- and microspectroscopy to obtain a spatially resolved chemical representation of the different types of calcium carbonate minerals. The cell surface of Arthrobacter sulfonivorans SCM3 served as a nucleus for the biomineralisation of vaterite precipitates. These were initially spherical and then continued to grow as chemically stable, rock-forming calcite crystals with rough edges.

Scanning electron microscopy of precipitates of the isolates Arthrobacter sulfonivorans SCM3 and Rhodococcus globerulus SCM4
Scanning electron microscopy of precipitates of the isolates Arthrobacter sulfonivorans SCM3 (left) and Rhodococcus globerulus SCM4 (right) (Image: Elektronenmikroskopisches Zentrum Jena, Sándor Nietzsche and Anna Rusznyak).