Biotic Acid Mine Drainage (AMD) Suppression Using Biogas Slurry: Identification of Key Inhibitory Components and Molecular Inactivation Mechanism of Acidithiobacillus ferrooxidans

Acid mine drainage (AMD) is a significant environmental challenge, and its treatment can be expensive and complicated. Acidithiobacillus could accelerate the rate of AMD formation by 5–6 orders of magnitude. Acidithiobacillus ferrooxidans (A. ferrooxidans) is the model species of Acidithiobacillus. We initially tried to use the biogas slurry as an organic additive to prevent AMD formation. We determined the essential inhibitory components of the biogas slurry as organic acids (acetic acid (AA), a typical example). The results revealed that AA (≥50 ppm) prevented A. ferrooxidans from forming AMD. The transcriptomic and untargeted metabolomic evaluation found 324 differentially expressed genes and 35 significantly transformed metabolites. Combinatorial omics analysis showed that the presence of AA significantly inhibited the membrane biogenesis, Fe²⁺, and RISC metabolism pathways, reducing energy metabolites such as Fe³⁺ and SO₄^2–. Furthermore, AA treatment induced A. ferrooxidans defense mechanisms and overconsumed its internal carbon sources. These findings proved that biogas slurry had a significant inhibitory effect on key microorganisms in highly acidified mineral soils and provided a scientific foundation for the prevention of acidification and the ecological restoration of newly mined areas.