Genome-Resolved Metagenomic Insights Into Aquifer Microbiome Adaptations Following Two Decades of Neutral In Situ Leaching

Neutral in situ leaching (ISL) is a prominent technique for extracting uranium (U) from sandstone-type U deposits. However, the intricate biogeochemical processes and associated microbial communities in environments subjected to prolonged neutral in situ U leaching remain insufficiently understood. Here, capitalizing on groundwater samples collected from an area subjected to two decades of ISL (Undergoing-ISL) and a background aquifer (Non-ISL) at the Qianjiadian U mining site (the first CO₂ + O₂ ISL site located in the Songliao Basin of northern China), we investigate the responses of aquifer microbiomes to CO₂ + O₂ ISL using genome-resolved metagenomics, resulting in a total of 315 medium- and high-quality bacterial metagenome-assembled genomes (MAGs) and 5 archeal genomes. Seven putative keystone MAGs were identified based on their high connection degree and low betweenness centrality within the co-occurrence network of reconstructed MAGs, notably highlighting the prevalence of microorganisms involved in sulfur, ammonium, and Fe(II) oxidation, which are integral to facilitating U(IV) oxidation during CO₂ + O₂ ISL. In particular, sulfur-oxidizing bacteria and Fe(II)-oxidizing bacteria (Rhodoferax spp. and SHZL01 spp.) played a key role in U mobilization during long-term neutral ISL. These findings enhance our understanding of subsurface microbial ecology and hold significant implications for bioremediation strategies and groundwater management in regions affected by mining.