The long-term retention of nitrogen in sediment of a tidal freshwater marsh in the Scheldt estuary (Belgium) was investigated by an in situ 15N-labeling experiment. Sediment of an unvegetated creek bank and sediment vegetated by common reed (Phragmites australis) were labeled with 15NN-enriched NH4+ after which 15N was traced into pore-water dissolved NH4+, NO3-, and N2; plant roots and leaves; and bulk sediment over a 1-yr period. Label retention in the sediment organic matter was further characterized by analyzing KCl-extracted sediment and hydrolyzable amino acids (including the bacterial biomarker D-alanine). Within weeks all added 15NH4+ was transformed and/or assimilated by the biota. Between 42% and 48% of the added label was recovered in plants, root, and sediment (collectively) after 3–6 months, whereas 24% remained after 1 yr. Transfer to plants and roots was rapid and although retention in leaves was transient, the 15N assimilated by roots was retained for the entire 1-yr period. Root assimilation could account for 12–23% of the 15N retained in vegetated plots. The majority of 15N was retained in the organic matter pool within the sediment (represented by 15N in the KCl extracted sediment and total hydrolyzable amino acids), primarily through efficient recycling of the 15N within the microbial community. 15N incorporation into D-alanine confirmed that bacteria were the major group of microorganisms responsible for the strong retention of 15N. This study shows the strong potential of reed roots and especially bacteria to retain nitrogen in tidal freshwater sediment over longer periods of time.