Engineering microbiome for carbonate precipitation in heavy oil produced water: Hot-zone identification of ion accumulation and crystallization

Microbially induced carbonate precipitation (MICP) presents a promising strategy for the softening and purification of produced water. However, produced water from heavy oil reservoirs exhibits high salinity, refractory organics, particularly with hardness ions such as Ca²⁺ and Mg²⁺, all of which substantially inhibit microbial mineralization activity. Industrially viable continuous-flow operational strategies remain underdeveloped, and the underlying biomineralization mechanisms are not yet fully elucidated. Here, we report the successful construction of an engineering microbiome through substrate gradient acclimation, achieving continuous and stable precipitation of Ca²⁺ (87.28 %) and Mg²⁺ (84.16 %). The process also revealed the sequential transformation of organic functional groups under high salinity perturbation. Hydroxyl groups (−OH) in extracellular polymeric substances preferentially bound divalent cations under neutral conditions, whereas carboxyl groups (−COO⁻) served as nucleation sites for carbonate formation under alkaline conditions. Extracellular carbonate precipitation predominated, while a minor fraction of amorphous magnesium carbonate was accumulated intracellularly. The engineering microbiome, dominated by urease-positive and hydrocarbon-degrading taxa, tolerated extreme salinity and hardness through metabolic complementarity and coordinated gene regulation. These findings demonstrate a robust, continuous-flow MICP process for HPW treatment, offering a foundation for industrial-scale integration with improved stability, efficiency, and microbiome resilience in complex environments.