Collaborative Research: ETBC: Deep Crustal Biosphere: Microbial Cycling of Carbon

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Intellectual Merit. For the past two decades studies from around the world in a wide variety of geological environments in both continental and marine settings have revealed the extent and variety of subsurface microbial ecosystems. From polar permafrost and ice sheets to equatorial marine sediments, from permeable aquifers ~200 meters deep to fractured rock 4 km deep, from deep vadose zones to oil reservoirs, diverse assemblages of bacteria and archaea, including many novel species, have been discovered. As depth and temperature increase in both the continental and oceanic crust, the effects of changing surface climate and photospheric processes on the subsurface microbial ecosystem diminish while Fisher-Tropsch-like and radiolytic processes, which enrich the crust with a mixture of organic and inorganic nutrients, become increasingly important. Regardless of the origin of the organic carbon, the rules that govern its biodegradation to CO2 and CH4 should be similar to those for petroleum reservoirs. The goals of this project are to determine those rules, determine the rates and limits of the biodegradation of organic matter by subsurface microbial ecosystems, and determine who is responsible. To accomplish these goals, investigators will utilize the readily accessible boreholes present in the mines of the Witwatersrand Basin, South Africa, which range in depth from 0.8 to 3.8 km, to examine the extent of and the inter-species interactions involved in biodegradation of organic photosynthate in shallow depths and abiogenic hydrocarbons at great depth. By employing a combination of 1) GCMS, HPLC, and NMR characterization of the DOC, 2) 13C analyses of DOC, DIC, hydrocarbons, lipids, and organic acids, 3) H isotope analyses of hydrocarbons, 4) 14C analyses of CH4, DIC, DOC, and lipids, and 5) stable isotope probes (SIP) of the 16S rRNA, we will delineate the relative importance of 1) chemoautotrophically produced organic acids, 2) abiogenically produced hydrocarbons, 3) thermogenically produced organic acids and 4) dissolved organic matter (DOC) from the surface photosphere, to the subsurface microbial community structure and its trophic interactions during carbon metabolism. These analyses will be performed on planktonic microbial communities collected from fracture water ranging in age from thousands of years to tens of millions of years. Broader Impacts. The program of study will engage undergraduates and graduates in hands-on interdisciplinary research where they will integrate isotope geochemistry and organic geochemical analyses with genetic analyses. Students from North America and South Africa will participate in the field and laboratory research as part of an ongoing collaboration with South African scientists. The RNA-SIP incubation experiments will be accompanied by isolation experiments that will further enlarge the subsurface culture collection and genetic database at the Univ. of Free State (UFS) and enhance their bioremediation and biotechnology efforts. The project will begin to integrate the genetic, taxonomic and functional diversity with respect to carbon metabolism. Investigators will work to assemble a web-based subsurface microbial database with a gallery of SEM/TEM images of subsurface microorganisms that can be downloaded for press releases; and as part of the gallery we will initiate a "Where in the world does this organism occur?" describing where related strains or sequences have been found. The web site will provide the beginnings of a microbial biogeographical database for the deep subsurface. When permitted by the mining companies videos of the sampling expeditions will be posted on the web site. The project will also provide fundamental scientific underpinnings for optimal exploitation of untapped natural gas reservoirs in the Witwatersrand Basin and for the development of CO2 sequestration projects in South Africa by private firms