RAPID: Collaborative Research: Carbon Cycling in Subsurface Hypersaline Environments Near the Abiotic Fringe

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Deep fractures within Precambrian Shield rock formations host fluids that range in age from tens of thousands of years for brackish fluid to more than a billion years for hypersaline fluids. Extensive reactions between the fracture fluid and surrounding rock lead to the production of helium, hydrogen, methane, short-chain hydrocarbons, oxidants and likely other compounds and to hypersalinity as non-hydrous silicates alter to clay in closed systems over time. Whether these deep Precambrian crust brines began as trapped ocean water and whether they contain microbial communities that have adapted to the increasing salinity over time is not known. Although such deep-seated, ancient brines are believed to be widespread, they have only been accessed in a few locations for scientific study. These principal investigators recently discovered hypersaline brines at 3 km depth in Moab Khotsong gold and uranium mine as part of an NSF/ICDP-supported project investigating a recent magnitude 5.5 earthquake. The brine salinity is 10x greater than any they have previously sampled in South Africa. Salinity also exceeds the known growth limit for methane-producing microbes. The methane-bearing gas, isotopic data, and fluid inclusions trapped in minerals suggest the brine could be at least 2 billion years old. This site is unique in the world in that it provides access to an ancient brine at high temperature (55 degree Centigrade). Researchers will analyze the chemistry of these unique subsurface brines, including the sources and amounts of gaseous energy sources and they will characterize the microbial communities in these ancient brines. If any extremophiles exist in these fractures, they will likely possess enzymes that will prove useful for remediating toxic metal- and radionuclide-contaminated water in the surface tailings ponds that are widespread in the mining districts of South Africa and elsewhere. If microbial life is absent in these fractures then they may provide a window into the abiotic processes responsible for the building blocks of life in the early Earth. The investigators have characterized the microbial communities of deep brackish fracture water at nearby mines, which have revealed multi-tier syntrophic microbial communities that recycle biogenically produced CH4. Halophilic methanogens and methanotrophs are not known and so either (1) other types of microbial C cycling must prevail here or (2) C cycling is limited to abiotic reactions. They successfully installed a packer/inconel U-tube into one hypersaline brine-filled fracture at 3.2 kmbls. and a steel valve on another at 3 kmbls., but increasing pressure and corrosion are causing both to leak. With the loss of key personnel at the end of the NSF/ICDP project, the collapse of the boreholes, and the loss of brine given their finite volume, researchers face losing an opportunity to sample a world class biogeochemical site. They therefore propose to return to Moab Khotsong Mine and with help from their South African colleagues to undertake a comprehensive survey and sampling of these brines and to take steps to secure longer-term access for future international researchers. They propose collecting sufficient biomass for metagenomic, metatranscriptomic and lipidomic analyses, gas for isotopic analyses and dating, and water for dissolved organic carbon characterization, as well as sealing and instrumenting other boreholes. The data provided by these brine samples will test the following hypotheses: 1) Life in the deeper, hotter, hypersaline fracture fluids is constrained by the energetic cost of combating intracellular water loss and an inventory of abiotic organic compounds formed by water/rock reactions over the past billions of years as well as biologically generated organic matter will be present and characterized for the first time; 2) Microbial communities present in cooler hypersaline fractures will be phylogenetically similar to phyla present in brackish fracture water and not related to the halophiles of existing surface saltpans; and 3) Biogenic CH4-supported communities found in brackish fluids will be replaced by communities supported by abiogenic hydrocarbons in hypersaline fluids, with greater expression of genes involved in osmotic regulation and with lipids whose composition provide lower membrane permeability.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.