Ammonium enrichment destabilizes the deep-sea recalcitrant carbon pool via direct nutrient relief and synergistic priming

Seawater samples (50 L each) were collected in December 2021 from the South China Sea (115.9961 E, 18.0097 N) at depths of 500 m and 2000 m (Fig. 1). Samples were filtered through pre-combusted (450 degrees C for 4 h) 0.7 um glass-fiber filters (GF/F, Whatman, UK) to remove plankton and particulate matter. Immediately following filtration on board the research vessel, the seawater was transferred into 1 L sterile screw-cap brown borosilicate glass bottles. The incubation was initiated immediately by placing the bottles in a temperature-controlled incubator at 24 +/- 0.5 degrees C in the dark. Upon disembarking, the microcosms were immediately transferred to a laboratory incubator and maintained under identical conditions. For each depth, two experimental groups were established: a treatment group (Ammonium-amended group) created by spiking each 1 L microcosm with 50 uL of a 1 M sterile ammonium chloride (NH4Cl) stock solution to achieve a final nominal concentration of 50 uM, and an unamended control group (Control group) which received no additions. Both groups were incubated in 1 L autoclaved amber borosilicate glass bottles under static, dark conditions at 24 degrees C. Samples for biogeochemical and microbial analyses were collected on days 1, 28, 60, and 90, with three biological replicates per treatment and time point.