Automatic Slide-Loader Fluorescence Microscope for Discriminative Enumeration of Subseafloor Life

Abstract

The marine subsurface environment is considered the potentially largest ecosystem on Earth, harboring one-tenth of all living biota (Whitman et al., 1998) and comprising diverse microbial components (Inagaki et al., 2003, 2006; Teske, 2006; Inagaki and Nakagawa, 2008). In deep marine sediments, the discrimination of life is significantly more difficult than in surface sediments and terrestrial soils because buried cells generally have extremely low metabolic activities (D’Hondt et al., 2002, 2004), and a highly consolidated sediment matrix produces auto-fluorescence fromdiatomaceous spicules and other mineral particles (Kallmeyer et al., 2008). The cell abundance in marine subsurface sediments has conventionally been evaluated by acridine orange direct count (AODC; Cragg et al., 1995; Parkes et al., 2000) down to 1613 meters below the seafloor (mbsf) (Roussel et al., 2008). Since the cell-derived AOsignals often fade out in a short exposure time, recognizing and counting cells require special training. Hence, such efforts to enumerate AO-stained cells from the subseafloor on photographic images have been difficult, and a verification of counts by other methods has been impossible. In addition, providing mean statistical values from low biomass sedimentary habitats has been complicated byphysical and time limitations, yet these habitats are considered critical for understanding the Earth’s biosphere close to the limits of habitable zones (Hoehler, 2004; D’Hondt et al., 2007).

Keywords

• subseafloor
• microscope