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

Abstract 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 CO 2 + O 2 ISL site located in the Songliao Basin of northern China), we investigate the responses of aquifer microbiomes to CO 2 + O 2 ISL using genome‐resolved metagenomics, resulting in a total of 315 medium‐ and high‐quality bacterial metagenome‐assembled genomes (MAGs) and 5 archaeal 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 CO 2 + O 2 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.