Searching for life in extreme environments relevant to Jovian's Europa: Lessons from subglacial ice studies at Lake Vostok (East Antarctica)

The objective was to estimate the genuine microbial content of ice samples from refrozen water (accretion ice) from the subglacial Lake Vostok (Antarctica) buried beneath the 4-km thick East Antarctic ice sheet. The samples were extracted by heavy deep ice drilling from 3659 m below the surface. High pressure, a low carbon and chemical content, isolation, complete darkness and the probable excess of oxygen in water for millions of years characterize this extreme environment. A decontamination protocol was first applied to samples selected for the absence of cracks to remove the outer part contaminated by handling and drilling fluid. Preliminary indications showed the accretion ice samples to be almost gas free with a low impurity content. Flow cytometry showed the very low unevenly distributed biomass while repeated microscopic observations were unsuccessful.We used strategies of Ancient DNA research that include establishing contaminant databases and criteria to validate the amplification results. To date, positive results that passed the artifacts and contaminant databases have been obtained for a pair of bacterial phylotypes only in accretion ice samples featured by some bedrock sediments. The phylotypes included the chemolithoautotrophic thermophile Hydrogenophilus thermoluteolus and one unclassified phylotype. Combined with geochemical and geophysical considerations, our results suggest the presence of a deep biosphere, possibly thriving within some active faults of the bedrock encircling the subglacial lake, where the temperature is as high as 50 °C and in situ hydrogen is probably present.Our approach indicates that the search for life in the subglacial Lake Vostok is constrained by a high probability of forward-contamination. Our strategy includes strict decontamination procedures, thorough tracking of contaminants at each step of the analysis and validation of the results along with geophysical and ecological considerations for the lake setting. This may serve to establish a guideline protocol for studying extraterrestrial ice samples.