‘Follow the Water’: Hydrogeochemical Constraints on Microbial Investigations 2.4 km Below Surface at the Kidd Creek Deep Fluid and Deep Life Observatory

Microbiological and geochemical data are presented to characterize the hydrogeochemistry and to investigate extant microbial life in fracture waters 2.4 km below surface, at the Kidd Creek Observatory in Canada. Previous studies identified the world’s oldest groundwaters with mean residence times on the order of millions to billions of years trapped in fractures in Precambrian host rock here. In this study, major ion chemistry, δ¹⁸O and δ²H isotopic signatures and dissolved gases in the fracture waters are shown to be distinct from potential contamination end-members, demonstrating the fracture waters are not impacted by waters used in mining operations. A previous work on sulfur isotope signatures suggested a longstanding indigenous population of sulfate-reducing bacteria in these highly reducing fluids and sufficient sulfate to support microbial activity. Here, we report the first evidence for extant visible and cultivable microbial life at this location. Anaerobic metabolisms were investigated using the Most Probable Number (MPN) method. The fracture fluids contained extant cells at low biomass density (∼10³–10⁴ cells/mL) and showed a strong response from autotrophic sulfate-reducers and alkane-oxidizing sulfate reducers. These lines of evidence provide the interpretational framework (chemical, hydrogeologic, and microbiologic) essential to the on-going genomic and metagenomic investigations at the Kidd Creek Observatory–the world’s most longstanding location for investigation of subsurface fluids and deep life at such profound depth.