| Summary: | The application of N fertilizer is one of the most critical soil acidification factors in China, and soil acidification significantly alters biogeochemical processes such as N loss. Anaerobic ammonium oxidation coupled with iron reduction (Feammox) is an important biological process for N loss in natural environments, with the end-products of N<sub>2</sub>, NO<sub>2</sub><sup>−</sup> and NO<sub>3</sub><sup>−</sup>. However, the response of Feammox pathways to soil pH fluctuation has not been thoroughly studied. In the current study, Feammox pathways and microbial communities were explored through a slurry culture experiment with an artificially adjusted pH combined with a <sup>15</sup>N isotope tracing technique and molecular biotechnology. Results showed significant differences in the gaseous N loss through Feammox (0.42–0.97 mg N kg<sup>−1</sup> d<sup>−1</sup>) under different pH conditions. The gaseous N loss pathways were significantly affected by the pH, and Feammox to N<sub>2</sub> was the predominant pathway in low-pH incubations. The proportion of N loss caused by Feammox coupled with denitrification increased as the soil pH increased. The gaseous N loss through Feammox increased by 43.9% when the soil pH decreased from 6.5 to 5.0. Fe-reducing bacteria, such as <i>Ochrobactrum</i>, <i>Sphingomonas</i>, and <i>Clostridium</i> increased significantly in lower pH incubations. Overall, this study demonstrated the effects of soil pH on Feammox pathways and extended the understanding of the N biogeochemical cycle in acidic soil.
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