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doi:10.2204/iodp.proc.314315316.133.2009

Microbiology and biogeochemistry

Sample processing

To study microbiological and biogeochemical characteristics in sediments at Site C0004, samples were obtained from 71 different depth locations (Table T20). Almost all sample processing was carried out after X-ray CT scanning and was completed within 1 h after core recovery on deck.

Cell abundance

Using paraformaldehyde-fixed slurry samples, preliminary microbial cell abundances were enumerated by visual inspection using the SYBR Green I staining method (Lunau et al., 2005; see “Microbiology and biogeochemistry” in the “Expedition 316 methods” chapter). Under epifluorescent microscopy, very high numbers of microbial cells were observed in sediments at Site C0004 (Fig. F47). In the near-seafloor core samples, relatively large coccoids and rod-shaped cells were observed with population size estimated to be 5.70 × 10⁹ ± 1.01 × 10⁹ cells/cm³ (N = 6). Some aggregate structures consisting of a few bright cells were observed in shallow sediment samples. Cell populations slightly decreased with depth to 150 m CSF and slightly increased at ~280 m CSF in the fault zone of the lithologic Unit III accretionary prism (Fig. F47). In the fault zone, 3.64 × 10⁹ ± 5.90 × 10⁸ cells/cm³ (N = 6) were present as the endolithic microbial community, of which some components produced brighter fluorescent signals than those in upper sediments (Fig. F48). Irregular coccoids as well as tiny coccoids were also observed. Interestingly, cell populations sharply decreased across the lithologic Unit III/IV boundary. Minimum cell abundance, 5.48 × 10⁸ ± 2.09 × 10⁸ cells/cm³ (N = 6), was observed at 320.8 m CSF. Fluorescent signals below the fault structure were recognizable as cells but were generally dark, likely indicating low metabolic activity. Average cell abundance in the sediments at Site C0004 was estimated to be 1.83 × 10⁹ ± 4.25 × 10⁸ cells/cm³ (N = 25).

The average detected microbial population at this site is notably higher than at other drilling sites that typically harbor 10⁶–10⁹ microbial cells/cm³ (Parkes et al., 2000). The population size generally decreases with increasing depth because of the limitation of available nutrients (carbon and energy sources) and habitable space during burial. One exception was observed during the Chikyu shakedown cruise CK06-06 offshore the Shimokita Peninsula in Japan, where the sediments harbor >10¹⁰ cells/cm³ throughout the core to 365 m CSF, showing little or no decrease in cell number with depth (Inagaki et al., submitted). At the Shimokita site, the strong fluid and gas flow from underlying coal layers as well as high photosynthetic production in the water column are thought to support the large microbial population. Elevated cell numbers in deep subseafloor environments were also observed in sediment cores at Site 1301 on the Juan de Fuca Ridge flank, where the upward flow of hydrothermal fluids from the basaltic aquifer supports microbial life (Cowen et al., 2002; Nakagawa et al., 2006; Engelen et al., 2008). Similarly, this environment may be affected by fluid and dissolved gas flow, in which dissolved nutrients (probably generated by physical degradation of subducted organic matter and/or reaction between sedimentary rocks and fluids) may support the abundant microbial life. The adaptation and ecological functioning of these cells in the subseafloor seep ecosystem will be intensively investigated on shore by cultivation, molecular (deoxyribonucleic acid/​ribonucleic acid/​enzyme), and lipid biomarker analyses including carbon and nitrogen isotopic compositions.

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