Supplementary Data for Manuscript: Changes in salinity impact nitrogen removal and carbon preservation in coastal wetlands sediment

Coastal wetlands naturally remediate nitrogen (N) pollution through microbial pathways that either remove reactive N via denitrification and anammox, or retain it via dissimilatory nitrate reduction to ammonium (DNRA). The balance among three processes is closely linked to the carbon (C) cycle, as both heterotrophic denitrification and DNRA consume organic C and release alkalinity. While salinity fluctuations can disrupt these processes through direct ionic stress or sulfur (S) cycling, their net impact on N removal and C preservation services remains unclear. Here, we deployed microcosm experiments using mangrove sediments under a large salinity gradient (0-30 psu). We quantified N transformation rates using ¹⁵N isotope tracing technique, combined with geochemical analysis, and functional genes quantification. Freshening from ambient 30 psu to 10 psu decreased N removal efficiency by ~20%. This decline was caused by reduced denitrification, whereas anammox and DNRA were unaffected. Meanwhile, lower salinity appears to have stimulated C decomposition via reduced ionic stress. The reduced sulfate input diminished total alkalinity (TA) generation relative to dissolved inorganic carbon (DIC). The stoichiometric shift of TA:DIC ratio could further contribute to acidification in adjacent coastal waters. Additionally, the S-mediated regulation of N partitioning appears to be nitrate-dependent: under nitrate limitation, higher sulfate favored N retention; conversely, with enriched nitrate, it potentially favored N removal. Integrating the coupling effect of salinity on interaction between N, C and S cycles, our study demonstrates that coastal water freshening may weaken wetlands’ ability to remove N and preserve C.