@article{Keiner2013,
title = {Raman Spectroscopy—An Innovative and Versatile Tool To Follow the Respirational Activity and Carbonate Biomineralization of Important Cave Bacteria},
author = {Robert Keiner and Torsten Frosch and Stefan Hanf and Anna Rusznyak and Denise M Akob and Kirsten Küsel and Jürgen Popp},
doi = {10.1021/ac401699d},
year = {2013},
date = {2013-09-01},
journal = {Analytical Chemistry},
volume = {85},
number = {18},
pages = {8708–8714},
abstract = {Raman gas spectrometry is introduced as a unique tool for the investigation of the respiratory activity that is indicative for growth of bacteria involved in biomineralization. Growth of these bacteria cannot be monitored using conventional turbidity-based optical density measurements due to concomitant mineral formation in the medium. The respiratory activity of carbonate-precipitating Arthrobacter sulfonivorans, isolated from the recently discovered Herrenberg Cave, was investigated during its lifecycle by means of innovative cavity-enhanced Raman gas analysis. This method allowed rapid and nonconsumptive online quantification of CO2 and O2 in situ in the headspace of the bacterial culture. Carbon dioxide production rates of A. sulfonivorans showed two maxima due to its pleomorphic growth lifecycle. In contrast, only one maximum was observed in control organism Pseudomonas fluorescens with a one-stage lifecycle. Further insight into the biomineralization process over time was provided by a combination of Raman macro- and microspectroscopy. With the help of this spatially resolved chemical imaging of the different types of calcium carbonate minerals, it was elucidated that the surface of the A. sulfonivorans bacterial cells served as nuclei for biomineralization of initially spherical vaterite precipitates. These vaterite biominerals continued growing as chemically stable rock-forming calcite crystals with rough edges. Thus, the utilization of innovative Raman multigas spectroscopy, combined with Raman mineral analysis, provided novel insights into microbial-mediated biomineralization and, therefore, provides a powerful methodology in the field of environmental sciences.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
Raman gas spectrometry is introduced as a unique tool for the investigation of the respiratory activity that is indicative for growth of bacteria involved in biomineralization. Growth of these bacteria cannot be monitored using conventional turbidity-based optical density measurements due to concomitant mineral formation in the medium. The respiratory activity of carbonate-precipitating Arthrobacter sulfonivorans, isolated from the recently discovered Herrenberg Cave, was investigated during its lifecycle by means of innovative cavity-enhanced Raman gas analysis. This method allowed rapid and nonconsumptive online quantification of CO2 and O2 in situ in the headspace of the bacterial culture. Carbon dioxide production rates of A. sulfonivorans showed two maxima due to its pleomorphic growth lifecycle. In contrast, only one maximum was observed in control organism Pseudomonas fluorescens with a one-stage lifecycle. Further insight into the biomineralization process over time was provided by a combination of Raman macro- and microspectroscopy. With the help of this spatially resolved chemical imaging of the different types of calcium carbonate minerals, it was elucidated that the surface of the A. sulfonivorans bacterial cells served as nuclei for biomineralization of initially spherical vaterite precipitates. These vaterite biominerals continued growing as chemically stable rock-forming calcite crystals with rough edges. Thus, the utilization of innovative Raman multigas spectroscopy, combined with Raman mineral analysis, provided novel insights into microbial-mediated biomineralization and, therefore, provides a powerful methodology in the field of environmental sciences.
@article{Rusznyak2012,
title = {Calcite Biomineralization by Bacterial Isolates from the Recently Discovered Pristine Karstic Herrenberg Cave},
author = {Anna Rusznyák and Denise M Akob and Sándor Nietzsche and Karin Eusterhues and Kai Uwe Totsche and Thomas R Neu and Torsten Frosch and Jürgen Popp and Robert Keiner and Jörn Geletneky and Lutz Katzschmann and Ernst-Detlef Schulze and Kirsten Küsel},
doi = {10.1128/AEM.06568-11},
year = {2012},
date = {2012-02-01},
journal = {Applied and Environmental Microbiology},
volume = {78},
number = {4},
pages = {1157–1167},
abstract = {Karstic caves represent one of the most important subterranean carbon storages on Earth and provide windows into the subsurface. The recent discovery of the Herrenberg Cave, Germany, gave us the opportunity to investigate the diversity and potential role of bacteria in carbonate mineral formation. Calcite was the only mineral observed by Raman spectroscopy to precipitate as stalactites from seepage water. Bacterial cells were found on the surface and interior of stalactites by confocal laser scanning microscopy. Proteobacteria dominated the microbial communities inhabiting stalactites, representing more than 70% of total 16S rRNA gene clones. Proteobacteria formed 22 to 34% of the detected communities in fluvial sediments, and a large fraction of these bacteria were also metabolically active. A total of 9 isolates, belonging to the genera Arthrobacter , Flavobacterium , Pseudomonas , Rhodococcus , Serratia , and Stenotrophomonas , grew on alkaline carbonate-precipitating medium. Two cultures with the most intense precipitate formation, Arthrobacter sulfonivorans and Rhodococcus globerulus , grew as aggregates, produced extracellular polymeric substances (EPS), and formed mixtures of calcite, vaterite, and monohydrocalcite. R. globerulus formed idiomorphous crystals with rhombohedral morphology, whereas A. sulfonivorans formed xenomorphous globular crystals, evidence for taxon-specific crystal morphologies. The results of this study highlighted the importance of combining various techniques in order to understand the geomicrobiology of karstic caves, but further studies are needed to determine whether the mineralogical biosignatures found in nutrient-rich media can also be found in oligotrophic caves.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
Karstic caves represent one of the most important subterranean carbon storages on Earth and provide windows into the subsurface. The recent discovery of the Herrenberg Cave, Germany, gave us the opportunity to investigate the diversity and potential role of bacteria in carbonate mineral formation. Calcite was the only mineral observed by Raman spectroscopy to precipitate as stalactites from seepage water. Bacterial cells were found on the surface and interior of stalactites by confocal laser scanning microscopy. Proteobacteria dominated the microbial communities inhabiting stalactites, representing more than 70% of total 16S rRNA gene clones. Proteobacteria formed 22 to 34% of the detected communities in fluvialfluvialDurch fließendes Wasser verursacht; zum Beispiel: "Fluviale Sedimente" = durch fließendes Wasser abgelagertes Material. sediments, and a large fraction of these bacteria were also metabolically active. A total of 9 isolates, belonging to the genera Arthrobacter , Flavobacterium , Pseudomonas , Rhodococcus , Serratia , and Stenotrophomonas , grew on alkaline carbonate-precipitating medium. Two cultures with the most intense precipitate formation, Arthrobacter sulfonivorans and Rhodococcus globerulus , grew as aggregates, produced extracellular polymeric substances (EPS), and formed mixtures of calcite, vaterite, and monohydrocalcite. R. globerulus formed idiomorphous crystals with rhombohedral morphology, whereas A. sulfonivorans formed xenomorphous globular crystals, evidence for taxon-specific crystal morphologies. The results of this study highlighted the importance of combining various techniques in order to understand the geomicrobiology of karstic caves, but further studies are needed to determine whether the mineralogical biosignatures found in nutrient-rich media can also be found in oligotrophic caves.
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 FirmicutesFirmicutesEine 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 (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 (left) and Rhodococcus globerulus SCM4 (right) (Image: Elektronenmikroskopisches Zentrum Jena, Sándor Nietzsche and Anna Rusznyak).
