The distinct phases of fresh-seawater mixing intricately regulate the nitrogen transformation processes in a high...

Excessive anthropogenic nitrogen inputs lead to the accumulation of nitrogen, and significantly impact the nitrogen transformation processes in estuaries. However, the governing of nitrogen during its transport from terrestrial to estuary under the influence of diverse human activities and hydrodynamic environments, particularly in the fresh-seawater mixing zone, remains insufficient researched and lack of basis. To address this gap, we employed multi-isotopes, including δ¹⁵N-NO₃^-, δ¹⁸O-NO₃^-, δ¹⁵N-NH₄⁺, and δ¹⁵N-PN, as well as microbial function analysis, to investigate the nitrogen transformation processes in the Pearl River Estuary (PRE), a highly anthropogenic and terrestrial estuary. Principle component analysis (PCA) confirmed that the PRE could clearly partitioned into three zone, e.g., terrestrial area (T zone), mixing area (M zone) and seawater area (S zone), in terms of nitrogen transportation and transformation processes. The δ¹⁵N-NO₃^- (3.38±0.60‰) and δ¹⁸O-NO₃^- (6.35±2.45‰) results in the inner estuary (T area) indicate that NO₃^-attributed to the domestic sewage and groundwater discharge in the river outlets lead to a higher nitrification rate in the outlets of the Pearl River than in the reaching and seawater intrusion areas, although nitrate is rapidly diluted by seawater after entering the estuary. The transformation of nitrogen in the T zone was under significant nitrogen fixation (0.61 ± 0.22 %) and nitrification processes (0.0043 ± 0.0032 %) (presumably driven by Exiguobacterium sp. (14.1 %) and Cyanobium_PCC-6307 (8.1 %)). In contrast, relatively low δ¹⁵N-NO₃^- (6.83 ± 1.24‰) and high δ¹⁸O-NO₃^- (22.13±6.01‰) imply that atmospheric deposition has increased its contribution to seawater nitrate and denitrification (0.53±0.13 %) was enhanced by phytoplankton/bacterial (such as Psychrobacter sp. and Rhodococcus) in the S zone. The assimilation of NH₄ results from the ammonification of NO₃^- reduces δ¹⁵N-NH₄⁺ (5.36 ± 1.49‰) and is then absorbed by particulate nitrogen (PN). The retention of nitrogen when fresh-seawater mixing enhances the elevation of δ¹⁵N-NH₄⁺ (8.19 ± 2.19‰) and assimilation of NH₄⁺, leading to an increase in PN and δ¹⁵N-PN (6.91 ± 1.52‰) from biological biomass (mainly Psychrobacter sp. and Rhodococcus). The results of this research demonstrate a clear and comprehensive characterization of the nitrogen transformation process in an anthropogenic dominated estuary, highlighting its importance for regulating the nitrogen dissipation in the fresh-seawater mixing process in estuarine ecosystems.