The ISO-BOSS: Quantifying and Resolving Metabolic Rates and Metabolic Pathways in the Deep Subsurface Biosphere
EMSL Project ID
39915
Abstract
Title: The ISO-BOSS: Quantifying and resolving metabolic rates and metabolic pathways in the deep subsurface biospherePNNL ISO-BOSS abstract
Recent evidence has shown that an extensive microbial community is flourishing in the deep terrestrial and marine subsurface biosphere. Indeed, studies of cell densities suggest that between one and two thirds of all known life exists within these environments. There is great speculation about the metabolic capacity of these microbes, and ongoing metagenomic sequencing and cultivation efforts are aimed at furthering our understanding of these communities' physiologies.
Due to the difficulties associated with conducting experiments in the deep subsurface -especially in marine systems- molecular and cultivation-based approaches are commonly used to study deep subsurface microbial communities. While metagenomic sequencing is a powerful tool for studying the functional potential of microbial communities, genomic data does not provide any information on metabolic rate. Cultivation affords the opportunity to study an organism under ex situ conditions, but again provide limited insight into the metabolic rates of microbes in situ. Studies have clearly shown that microbial metabolic rates are different between pure culture and mixed communities due to metabolic and ecological interactions such as metabolite exchange and antimicrobial warfare. Thus, our understanding of the rates of microbial metabolism in situ is extremely limited, especially in the deep subsurface. While geochemical measurements and thermodynamic / kinetic modeling can provide us with constraints on metabolic activity, such data do not tell us about the degree to which each particular species (ribotype) contributes to the net condition. Furthermore, geochemical data cannot tell us about the sensitivities of each ribotype to environmental perturbations, including anthropogenic perturbations. Understanding of the role that deep subsurface communities play in global biogeochemical cycles requires a far better understanding of their metabolic rate and physiological capacity. To date, no technologies exist that enable us to effectively measure a range of metabolic rates in situ.
Here we propose to leverage of ongoing advances in osmotic sampling technology to develop a microbial sampling system (termed BOSS) that can be used to conduct tracer studies in situ to A) calculate microbial metabolic rate via 13C or 15N isotopic incorporation into peptides and B) characterize metabolic intermediates and proteins containing 13C enriched carbon via solid-state NMR and mass spectrometric capabilities. In collaboration with scientists as EMSL, we have developed the first osmotically powered biological sampling system (the BOSS) that preserves a continuous series of microbial samples for molecular analyses such as qPCR, metagenomics, 'pyrotag' sequencing and peptide sequencing. The proposed research builds upon that development by incorporating stable isotopically labeled tracers (which are harmless) to measure metabolic rate and enable downstream analyses of metabolites and proteins to better understand biochemical capacity. Specifically, we will conduct proof of concept tests using Shewanella odeinensis grown in laboratory chemostat under aerobic and anaerobic conditions. By growing them on select isotopically labeled substrates, we will be able to validate the efficacy of using mass spectrometric analyses to calculate metabolic rate (while providing data on the peptide representation). We will also assess whether solid-state NMR or mass spectrometry are most appropriate to characterize metabolic intermediates from BOSS samples. Based on our results here, we will modify and deploy a BOSS (the ISO-BOSS) containing isotopically labeled substrates into a deep subsurface serpentinizing system in the Cazadero, California. The resulting data will be among the first metabolic rate measurements conducted in situ. Moreover, isotopically labeled intermediates or proteins will indicate which particular metabolic processes are occurring in situ. The temporal resolution of the ISO-BOSS will provide insight into whether microbial metabolic rate exhibits changes in response to geochemical perturbations over time.
This project is well aligned with the Geochemisty/Biogeochemistry science theme in that it seeks to investigate and quantify the role that deep subsurface microbial communities play in global carbon cycling. The proposed research is highly in line with the EMSL's overarching mission to support 'world-class research in the biological, chemical, and environmental sciences to provide innovative solutions to the nation's environmental challenges as well as those related to energy production.' The proposed research represents a fundamentally new approach to studying microbial metabolism, which integrates the scientific and technical expertise of PNNL scientists. This development could not occur without the inclusion of the PNNL, as their expertise is essential to the success of this project. Upon completion, the scientific community will have the tools to develop a more quantitative understanding of the role of key microbial ribotypes in biogeochemical carbon cycling. Our open access policy will facilitate this dissemination, as we will make our findings and specifications readily available to the public as soon as is practical. Upon completion, investigators will be better equipped to constrain microbial metabolic rate in the deep subsurface, and better understand the role they play in maintaining our surficial biosphere.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2010-10-01
End Date
2012-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
Robidart JC, SJ Callister, PF Song, CD Nicora, CG Wheat, and PR Girguis. 2013. "Characterizing Microbial Community and Geochemical Dynamics at Hydrothermal Vents Using Osmotically Driven Continuous Fluid Samplers." Environmental Science & Technology 47(9):4399-4407. doi:10.1021/es3037302