Complete ammonia oxidizers (comammox), which convert ammonia to nitrate through nitrite, are a newly discovered nitrifying group. In recent years, comammox Nitrospira have been discovered in various natural and engineered ecosystems. However, little is known about the distribution dynamics of comammox Nitrospira in estuary tidal flat sediments. Chongming eastern tidal flat, the largest tidal flat in the Yangtze River Estuary, was selected as the research area. Through a combination of molecular biology assays, phylogenetic analysis based on functional gene sequences and statistical correlation with physicochemical properties, we determined the distribution and diversity of comammox Nitrospira in Chongming eastern tidal flat and analyzed the potential influencing environmental factors. The results indicated comammox Nitrospira were widely distributed in Chongming eastern tidal flat, while with lower abundance than the ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA). Clade A1 comammox Nitrospira showed adaptation to relatively high salinity, and was more distributed in middle and low tidal flats, while clade A2 and clade A3 were mostly distributed in high tidal flats with low salinity. The abundance and community structure of comammox Nitrospira were mainly affected by salinity, ammonia concentration, and temperature. This study showed the general existence of comammox Nitrospira in Chongming eastern intertidal sediments and indicated that the differences of tidal locations, which lead to a gradient in the physicochemical properties of the sediments, in turn affects the spatial distribution of comammox Nitrospira in estuary tidal flats.