Estuarine sediment-seawater microcosms were established to evaluate the influence of salinity on the population, transcriptional activity, and diversity of ammonia-oxidizing archaea (AOA) and bacteria (AOB). AOA was found to show the most abundant and the highest transcriptional activity under moderate salinity; on the other hand, AOB abundance was not sensitive to salinity variation but showed the highest transcriptional activity in the low-salinity microcosms. AOA exhibited more advantages than AOB on growth and ammonia-oxidizing activity under moderate- and high-salinity environments. The highest richness and diversity of active AOA were found under salinity of 15 psu. All the active AOA detected under the salinities studied were clustered into Nitrosopumilus maritimus linage, with the composition shifted from N. maritimus C12 cluster, N. maritimus like 1.1 cluster, N. maritimus SCM1 cluster, and N. maritimus like 1.2 cluster to N. maritimus C12 and N. maritimus A10 clusters when salinity was increased from 5 to 30 psu.