Dissimilatory nitrate reduction to ammonium (DNRA) and denitrification are two nitrate respiration pathways in the microbial nitrogen cycle. Diversity and abundance of denitrifying bacteria have been extensively examined in various ecosystems. However, studies on DNRA bacterial diversity are limited, and the linkage between structure and activity of DNRA communities has yet to be discovered. We examined the composition, abundance and activities of DNRA communities at five sites along salinity gradients in the New River Estuary, North Carolina, USA, a shallow eutrophic system. Sediment slurry incubation experiments with 15N-nitrate were conducted to measure potential DNRA rates while abundance of DNRA communities was estimated using quantitative PCR of nrfA genes encoding cytochrome C nitrite reductase, which is unique to DNRA. A pyrosequencing method targeting nrfA genes was developed using an Ion Torrent sequencer to examine diversity and composition of DNRA communities within the estuarine sediment communities. We found a positive correlation between nrfA gene abundance and DNRA rates. Among the environmental variables, %organic and sulfide in sediments might support higher activities of DNRA communities. Pyrosequencing analysis of nrfA genes revealed spatial variation of DNRA communities along the saline gradient of New River Estuary. Relative abundance of dominant populations was found to have significant influences on overall activities and abundance of DNRA communities. Thus, we found that abundance of dominant DNRA bacteria was an important regulator of DNRA activities in the eutrophic New River Estuary.