Salinization is a growing problem throughout the world and poses a threat especially to freshwater ecosystems. However, much remains to be learned about the mechanisms by which salinity impacts microbially mediated biogeochemical processes. Elevated nitrogen (N) concentrations in estuarine ecosystems have led to their eutrophication, but the relationship between N transformation and the functional genes involved in the response to saltwater intrusion is poorly understood. Here, using the Minjiang River, a tidal river in southeastern China as an easily accessible natural laboratory, we conducted a 2-year field survey to investigate N speciation during ebb and flood tides. Then, in a laboratory experiment we simulated the varying degrees of salt intrusion that occur in natural tidal reaches. The microcosm study allowed quantitative assessments of N transformation and functional gene responses. The field surveys showed that concentrations of NH4+ rose during flood tides, while the concentrations of NO3- and total N fluctuated. In the microcosms, NO3- concentrations decreased in response to salt pulses, due to simultaneous declines in the abundance of genes responsible for nitrification and increases in the abundance of those involved in dissimilatory nitrate reduction to ammonium (DNRA). The elevated salinity led to increased yields of NH4+, a response that correlated positively with the abundance of nrfA genes, involved in DNRA. Furthermore, an increase in salinity promoted N2O accumulation during the denitrification process. Altogether, our study suggests that saltwater intrusion leads to a decrease in nitrification while favoring N transformation via denitrification and DNRA and that N2O accumulation in the water is dependent on the strength of the salt pulse.