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Methyl-compound use and slow growth characterize microbial life in 2-km-deep subseafloor coal and shale beds

  1. Victoria J. Orphana,1
  1. aDivision of Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125;
  2. bGeomicrobiology Group, Kochi Institute for Core Sample Research, Japan Agency for Marine-Earth Science and Technology (JAMSTEC), Monobe B200, Nankoku, Kochi 783-8502, Japan;
  3. cGeobiotechnology Group, Research and Development Center for Submarine Resources, JAMSTEC, Monobe B200, Nankoku, Kochi 783-8502, Japan;
  4. dResearch and Development Center for Ocean Drilling Science, JAMSTEC, Yokohama, Kanagawa 236-0001, Japan
  1. Edited by David M. Karl, University of Hawaii, Honolulu, HI, and approved September 6, 2017 (received for review May 5, 2017)

  1. Fig. S1.

    SYBR-stained cells after separation and FACS concentration on ITO-coated 0.2-μm polycarbonate NanoSIMS membranes from the two highest cell abundance conditions in 15R3 coal and mixed lithology. SYBR and SEM images highlight some distinctions in cell morphology between the two methanol amended samples. White arrows indicate region of SYBR image on larger membrane target area. The 15R3 coal incubation amended with methylamine (A), SEM image of A (B), 15R3 coal amended with methanol (C), C magnified with false color (D), mixed lithology amended with methanol + H2 + ammonium (E), SEM image of E (F), mixed lithology amended with methanol (G), and mixed lithology amended with methanol magnified with false color (H; Inset, the same sample filtered before cell sorting to demonstrate cell density without concentration). (Scale bar: H, Inset, 20 μm.)

  2. Fig. 2.

    1H ion (A), 2H ion (B), 14N12C ion (C), and 15N12C ion (D) counts of ROIs from 15R3 coal with 15N-methylamine and 2H2O. The pink arrow indicates biomass that is only present in 1H and 14N12C ion maps and connected to biomass present in all four ion maps, indicative of new biomass growth. Large circles are used for nitrogen background standard correction. There are also six other ROIs drawn on the 14N12C map that do not show 15N12C or 2H enrichment for comparison.

  3. Fig. 3.

    2H biomass generation time versus 15N biomass generation time, in years, calculated from ROIs where the 15N12N ion counts were above the Poisson error (Materials and Methods). The dashed line is where 2H and 15N generation time is at parity.

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