Abstract
The inner wall of the Tsukioka hot spring well was covered with green microbial mats which contained black crude oil. The hot spring is located in Tsukioka, Niigata, Japan. The hot spring water (pH 7.3, Eh -184 mV and 49.2℃) that was produced from a 280m in depth originated from fossil seawater. The aim of this study was to investigate of covellite biomineralization in the green microbial mats with crude oil from the Tsukioka hot spring water. Observations by optical and electron microscopy, phospholipid analysis and bacterial cultivation showed that the biomineralization of covellite (CuS) was found on the surface of crude oil droplets. ED-XRF analysis showed that the crude oil contained SO3 (67.6wt%), CaO (24.8wt%) and Cu2O (2.6wt%). Observations by optical and epifluorescence microscopy as well as scanning electron microscopy (SEM) showed that two morphological types of bacteria (i.e., filamentous and coccus typed bacteria) were found on crude oil droplets. Transmission electron microscopic (TEM) observations further revealed that the cell wall of the filamentous bacteria had a double membrane and was surrounded by Extracellular Polymeric Substance (EPS), which might provide the favorable nucleation sites for CuS. X-ray diffraction and electron diffraction analyses of the microbial mats showed that the materials precipitated on the surface of the oil droplets were highly crystallized covellite (CuS). FT-IR spectra of the microbial mats that contained covellite showed the appearance of C-N-H, P=O, P-O-C and N-H bands, indicating the presence of phospholipids (lecithin and phosphoric ester) in the microbial mats. Cultivation experiments, furthermore, demonstrated that the cultivated bacteria were identified as sulfate reducing bacteria (for coccus typed bacteria) and oil decomposing bacteria (for filamentous bacteria). A schematic formation model is here suggested to better understand the interaction between bacterial cells and oil droplets under anaerobic conditions in the Tsukioka hot spring water. The biomineralization of covellite described in this study might have profound implications for bioremediation of oilcontaminated site.
