Microbial Community in Black Rust Exposed to Hot Ridge Flank Crustal Fluids

Abstract

During Integrated Ocean Drilling Program Expedition 301, we obtained a sample of black rust from a circulation obviation retrofit kit (CORK) observatory at a borehole on the eastern flank of Juan de Fuca Ridge. Due to overpressure, the CORK had failed to seal the borehole. Hot fluids from oceanic crust had discharged to the overlying bottom seawater and resulted in the formation of black rust analogous to a hydrothermal chimney deposit. Both culture-dependent and culture-independent analyses indicated that the black-rustassociated community differed from communities reported from other microbial habitats, including hydrothermal vents at seafloor spreading centers, while it shared phylotypes with communities previously detected in crustal fluids from the same borehole. The most frequently retrieved sequences of bacterial and archaeal 16S rRNA genes were related to the genera Ammonifex and Methanothermococcus, respectively. Most phylotypes, including phylotypes previously detected in crustal fluids, were isolated in pure culture, and their metabolic traits were determined. Quantification of the dissimilatory sulfite reductase (dsrAB) genes, together with stable sulfur isotopic and electron microscopic analyses, strongly suggested the prevalence of sulfate reduction, potentially by the Ammonifex group of bacteria. Stable carbon isotopic analyses suggested that the bulk of the microbial community was trophically reliant upon photosynthesis-derived organic matter. This report provides important insights into the phylogenetic, physiological, and trophic characteristics of subseafloor microbial ecosystems in warm ridge flank crusts.

Microbial Ecology of the Dark Ocean above, at, and below the Seafloor

“…Subsequent studies in this environment confirmed this observation (392,425). The function of these Firmicutes organisms is currently unclear, as the species observed do not group closely with cultivated microorganisms, although it has been suggested that the microorganisms may be involved in nitrogen or sulfur cycling (93,392). More work is necessary to define the range of microorganisms that exist in subsurface oceanic crust and to determine their metabolic processes.…”

“…A set of studies from vent sites on the eastern flank of Juan de Fuca Ridge, centered around the Baby Bare basalt outcrop diffuse hydrothermal (ϳ25°C) vents and an instrumented observatory that targets deeply buried (200 to 300 m) basement hydrothermal (ϳ65°C) fluid flow a few kilometers away (93,232,392,425), found some similar patterns in microbial ecology in basalt-hosted hydrothermal fluid. As before, thermophilic bacteria related to Desulfurobacterium and Epsilon-and Deltaproteobacteria were abundant in 16S rRNA gene clone libraries, although Thermotogales and candidate phylum OP8 were also detected (232).…”

Archaeoglobus sulfaticallidus sp. nov., a thermophilic and facultatively lithoautotrophic sulfate-reducer isolated from black rust exposed to hot ridge flank crustal fluids

“…Strain PM70-1 T was enriched and isolated from black rust formations collected from the steel surface of a seafloor borehole observatory (CORK 1026B) retrieved during IODP Expedition 301 on the eastern flank of Juan de Fuca Ridge, eastern Pacific Ocean (Fisher et al, 2005;Nakagawa et al, 2006). The black rust was located just above the parts of the CORK designed to seal the top of a 295 m-deep borehole that was reaching through sediments and 48 m into basement.…”

“…The black rust was located just above the parts of the CORK designed to seal the top of a 295 m-deep borehole that was reaching through sediments and 48 m into basement. The rust had probably formed by the mixing of the cold surrounding seawater and hot (~64 u C) crustal fluids leaking through the seal (Nakagawa et al, 2006). Fresh samples of the black rust were crushed in a sterile mortar in an anaerobic glove box under an N 2 /CO 2 /H 2 (90 : 5 : 5, by vol.)…”

“…The bottles were sealed with butyl rubber stoppers and aluminium crimp caps and brought back to the laboratory at the University of Bergen without temperature control. The primary enrichment of PM70-1 T was performed in a 25 ml bottle filled with 10 ml DSMZ 63-2 medium (Nakagawa et al, 2006) adjusted to pH 6.4 at room temperature. The bottle was inoculated with 1 ml of the black rust slurry and incubated at 70 u C without shaking.…”