Magnetite-enhanced humic substance-driven anaerobic oxidation of methane

Redox-active natural organic matter (NOM, e.g., humic substances) is ubiquitous in organic-rich environments, which are the major sources of global methane emissions. Extracellular electron transfer (EET)-mediated anaerobic oxidation of methane (AOM) using NOM as electron acceptors is critical for mitigating methane emissions from these systems. However, the effects of magnetite on NOM-AOM and its underlying mechanisms are still not well understood. Batch experiments coupled with ¹³CH₄ isotope labeling showed that magnetite significantly enhanced AOM driven by humic acid (HA), a representative NOM. The methane oxidation rate in magnetite-amended systems (0.54 ± 0.04 mmol/g VSS/d) was 1.3-fold higher than that in the system without magnetite (0.41 ± 0.09 mmol/g VSS/d), with acetate identified as a key intermediate metabolite. Microbial community analysis revealed that Methanosarcina likely plays a pivotal role in HA-driven AOM. Its relative abundance in magnetite-amended systems was 1.5-fold higher than in controls (11.0 % vs. 7.2 %). Additionally, the electroactive acetate-oxidizing bacterium norank_Anaerolineaceae was also detected. Electrochemical characterization further demonstrated that magnetite enhanced microbial EET capacity, as evidenced by increased electron storage capacity, reduced electron transfer resistance, and enhanced redox activity of outer membrane proteins. Functional gene prediction analysis revealed that magnetite upregulated genes encoding electrically conductive pili and the membrane-bound electron transfer complex Rnf, which may underlie the promotion of EET by magnetite. These findings provide new insights into carbon cycling and highlight magnetite as a potential mediator for mitigating methane emissions from wetlands and paddy soils, which are rich in NOM and iron minerals.