LIFE ON THE FRINGE: SURVEYING THE ECOPHYSIOLOGICAL TENACITY OF METHANOGENS AND ANAEROBIC METHANOTROPHS IN THE OLIGOTROPHIC DEEP SUBSURFACE BIOSPHERE

Abstract

Methanogenesis coupled to the Wood-Ljungdahl pathway and its reversal, the anaerobic oxidation of methane, AOM, are believed to be among the most ancient metabolisms known to life on Earth. Recent advances in cultivation-independent techniques utilizing high-throughput sequencing, cellular imaging, geochemical modeling and experimentation, and bioinformatics are rapidly altering the ways in which we investigate complex microbial communities in the environment. In particular, the application of these methods to the exploration of the deep biosphere has ignited a renaissance in the study of methane biogeochemistry, as mining efforts into these environments – both literally and computationally – continue to reveal that methanogens and anaerobic methanotrophs (ANMEs) are more diverse, pervasive, and resilient than ever previously thought.

In the spirit of continuing to “move the goalposts” that define the extent of microbial methane metabolisms, this dissertation explores the ecophysiologies of methanogens and ANMEs surviving under extreme conditions which characterize the deep biosphere of Earth and Mars. We develop a novel fluorescent in situ hybridization method, FISH-TAMB, to visualize mRNA in living methanogens and ANMEs, discussing its potential to characterize microbial dark matter and identify horizontal gene transfer based on metabolic function. We then investigate oligotrophic continental fracture fluid from South Africa, characterizing the potential of AOM in a novel species of phylum Candidatus “Bathyarchaeota”. We then explore the high-pressure, high-temperature sub-seafloor sediments of the Nankai Trough, employing high-pressure cultivation, stable isotope probing, 16S rDNA sequencing, and metagenomics to hunt for the first experimental evidence of thermo-piezotolerant AOM. Finally, we subject axenic cultures of the methanogen Methanosarcina barkeri to controlled incubations simulating freezing and highly oxidizing conditions of a perchlorate-riddled Mars Special Region, utilizing transcriptomics to better understand the potential of biological methanogenesis on the Red Planet. Collectively, these results demonstrate the remarkable tenacity of methanogens and ANMEs to survive on the biotic fringe.