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786 1. Introduction 786 2. Organization of the Ideas Presented and Discussed at the Conference 787 2.1. Definition of key terms 787 3. Martian Environments Deemed to Be Most Prospective for Extant Martian Life 788 3.1. Caves 790 3.2. Deep subsurface 790 3.3. Ice 793 3.4. Salts 796 4. Methodologies: A Summary of Possible Approaches and Strategies to Search for Extant Life on Mars 797 4.1. Geologically guided search strategies 797 4.2. Possible detection methods for extant martian life 799 4.3. Possible constraints relevant to the search for extant martian life derived from laboratory experiments 802 5. Discussion 804 Acknowledgments 805 References 805

Sources and sinks of methane (CH ) are critical for understanding global biogeochemical cycles and their role in climate change. A growing number of studies have reported that CH concentrations in cave ecosystems are depleted, leading to the notion that these subterranean environments may act as sinks for atmospheric CH . Recently, it was hypothesized that this CH depletion may be caused by radiolysis, an abiotic process whereby CH is oxidized via interactions with ionizing radiation derived from radioactive decay. An alternate explanation is that the depletion of CH concentrations in caves could be due to biological processes, specifically oxidation by methanotrophic bacteria. We theoretically explored the radiolysis hypothesis and conclude that it is a kinetically constrained process that is unlikely to lead to the rapid loss of CH in subterranean environments. We present results from a controlled laboratory experiment to support this claim. We then tested the microbial oxidation hypothesis with a set of mesocosm experiments that were conducted in two Vietnamese caves. Our results reveal that methanotrophic bacteria associated with cave rocks consume CH at a rate of 1.3-2.7 mg CH · m · d . These CH oxidation rates equal or exceed what has been reported in other habitats, including agricultural systems, grasslands, deciduous forests, and Arctic tundra. Together, our results suggest that depleted concentrations of CH in caves are most likely due to microbial activity, not radiolysis as has been recently claimed. Microbial methanotrophy has the potential to oxidize CH not only in caves, but also in smaller-size open subterranean spaces, such as cracks, fissures, and other pores that are connected to and rapidly exchange with the atmosphere. Future studies are needed to understand how subterranean CH oxidation scales up to affect local, regional, and global CH cycling.

Recent observations suggest that karst landscapes may be an unaccounted sink for atmospheric CH 4 , but questions remain about the processes contributing to sub-atmospheric CH 4 mole fractions in caves. The CH 4 dynamics associated with karst environments were studied over 18 months at 6 locations in Buckner Cave, Southern Indiana by measuring the mole fractions and stable isotopic composition of CH 4 and carbon dioxide (CO 2). CO 2 mole fractions were used to infer seasonal changes in airflow. Samples were obtained on a monthly basis. CH 4 mole fractions ranged from 1.9 ± 0.1 ppm near the cave entrance to 0.1 ± 0.1 ppm in the more remote parts of the cave. The carbon and hydrogen stable isotopic compositions of CH 4 in the cave ranged from-58.7 to +7 ‰ (VPDB) and-170 to +10 ‰ (VSMOW), respectively. The isotopic data suggest that the CH 4 dynamics of Buckner Cave can be

The Amazon rainforest is a biodiversity hotspot and large terrestrial carbon sink threatened by agricultural conversion. Rainforest-to-pasture conversion stimulates the release of methane, a potent greenhouse gas. The biotic methane cycle is driven by microorganisms; therefore, this study focused on active methane-cycling microorganisms and their functions across land-use types. We collected intact soil cores from three land use types (primary rainforest, pasture, and secondary rainforest) of two geographically distinct areas of the Brazilian Amazon (Santarém, Pará and Ariquemes, Rondônia) and performed DNA stable-isotope probing coupled with metagenomics to identify the active methanotrophs and methanogens. At both locations, we observed a significant change in the composition of the isotope-labeled methane-cycling microbial community across land use types, specifically an increase in the abundance and diversity of active methanogens in pastures. We conclude that a significant increase in the abundance and activity of methanogens in pasture soils could drive increased soil methane emissions. Furthermore, we found that secondary rainforests had decreased methanogenic activity similar to primary rainforests, and thus a potential to recover as methane sinks, making it conceivable for forest restoration to offset greenhouse gas emissions in the tropics. These findings are critical for informing land management practices and global tropical rainforest conservation.