Microbial ecology of the deep biosphere

Exploring the deep subseafloor biosphere

Covering 70% of the globe, the oceans represent Earth’s largest biome, supporting diverse microbial life that is integral to many globally important biogeochemical cycles. Growing evidence suggests that the number of microbial cells in subseafloor sediments may equal that in the overlying ocean, yet we know very little about their diversity, activities, and function. Through our participation in C-DEBI and the NASA Astrobiology Institute ‘Life Underground’ team, members in the Orphan lab are investigating the metabolic diversity, activity, and interactions of indigenous microorganisms within deep subseafloor habitats, including bacteria, archaea, and viruses. Specific environments currently under investigation include subseafloor sediments (Santa Barbara Basin and South Pacific Gyre), deep coal seam and shale beds (Shimokita Peninsula, IODP Expedition 337), and subduction zones (Nankai Trough, IODP Expedition 370) as well as surface habitats (mud volcanoes, hydrothermal systems, and methane vents) that can serve as conduits for deep sourced fluids and microbial life.

On Earth, microorganisms appear to inhabit all physical space that provides the minimum requirements for life. These include the availability of water, carbon, nutrients, and light or chemical energy. While these are generally abundant in surface or near-surface environments, their mode and distribution in the subsurface are poorly constrained. Nevertheless, it has now been shown unequivocally that archaea and bacteria inhabit deeply buried rocks and sediments where they contribute to biogeochemical cycles. All evidence suggests that these subsurface ecosystems are spatially enormous, extraordinarily diverse, and critically undersampled. On other planets, at least in our solar system, putative extant or extinct life would most likely reside underground or in massive ice shells where damaging radiation is less abundant and temperatures more moderate. NASA has declared that searching for these extraterrestrials a priority, and has placed special emphasis on near-by planets where landed missions are more than just a possibility. Robust strategies for subsurface life detection on Mars, Europa, or other potential targets are poorly developed, in part because these techniques are scientifically and technologically extremely difficult. Our work focuses on using the deep biosphere on Earth as an analog to the extraterrestrial subsurface to develop and test new, non-invasive in situ life detection technologies.

NASA Astrobiology Institute

Our lab is part of the larger NASA Astrobiology Institute’s Life Underground team and we focus on in situ life detection and characterization and cultivation of deep biosphere organisms.

Our main research questions:

  • How do we search for microbes in the subsurface? And once detected, how do we understand their ecophysiology?
  • What is the distribution of microbial biomass and activity in the subsurface? What physicochemical factors influence their distributions?
  • What biosignatures or evidence of subsurface environments stand the test of time? Are there novel preservation possibilities in the subsurface?
  • Can these organisms be cultured in the lab using inventive new approaches, such as “Down-flow hanging sponge” reactors to investigate attached communities in porous media, or optically accessible diffusion chambers with established electrochemical gradients?

Deep Biosphere Project BLM-1 (NASA NAI)

Center for Dark Energy Biosphere Investigations (C-DEBI)

The overarching scientific goal of the C-DEBI community is to develop an integrated understanding of microbial subseafloor life, connecting the molecular, cellular, and ecosystem scales.

Research falls under 3 broadly defined themes:

Theme 1: Fluxes, Connectivity, and Energy—centering on subseafloor environmental conditions.
Theme 2: Activities, Communities, and Ecosystems—emphasizing resident microbial communities
Theme 3: Metabolism, Survival, and Adaptation—concentrating on the actions and traits of individual microbial species.

As part of Theme 2, we have been developing and applying single cell activity assays that enable tracking of biosynthetic activity and physiological potential of microorganisms within deep subseafloor environmental samples. These environments are challenging to study, with low cell abundance and slow rates of growth, requiring ultraclean sample handling and sensitive methods for microbial activity detection. Using stable isotope tracers coupled to nanoSIMS or FISH-nanoSIMS, we can quantify the amount of carbon-13, nitrogen-15, sulfur-33 or deuterium in single cell biomass from isotopically labeled substrates (for example 15N or 13C-methylamine, 33S-sulfate, and D2O). The distribution of isotopic enrichment among individual cells recovered after incubation enables the quantification of the proportion of anabolically active cells and can provide information regarding the identity of microorganisms metabolizing specific substrates. In addition to stable isotope probing, we are also exploring the use of BONCAT to visualize biosynthetically active microorganisms from the deep biosphere under defined physico-chemical conditions.

(Above) Fig. IIIc.3 from the C-DEBI proposal Overview of Theme 2 research approaches to be carried out using subseafloor samples and in situ experiments. Left column: representative core. Center (top to bottom): population structure (protein families and organisms); subseafloor metabolite concentration profile and production rates; SIMS image of subseaflor microbes. Right column (top to bottom): metagenomic analysis combined with PCA of environmental data; stable isotype experiments; pathway map of functional potential.

IOPD Expedition 370: T-Limit of the Deep Biosphere off Muroto

IOPD Expedition 370 T-limit of the Deep Biosphere off Muroto

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IOPD Expedition 337: Deep Coalbed Biosphere off Shimokita

IOPD Expedition 370 T-limit of the Deep Biosphere off Muroto

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