Macroalgae has bloomed in the brackish lake of Shenzhen Bay, China continuously from 2010 to 2014. Gracilaria tenuistipitata was identified as the causative macroalgal species. The aim of this study was to explore the outbreak mechanism of G. tenuistipitata, by studying the effects of salinity and nitrogen sources on growth, and the different nitrogen sources uptake characteristic. Our experimental design was based on environmental conditions observed in the bloom areas, and these main factors were simulated in the laboratory. Results showed that salinity 12 to 20 ‰ was suitable for G. tenuistipitata growth. When the nitrogen sources' (NH4+, NO3−) concentrations reached 40 µM or above, the growth rate of G. tenuistipitata was significantly higher. Algal biomass was higher (approximately 1.4 times) when cultured with NH4+ than that with NO3− addition. Coincidentally, macroalgal bloom formed during times of moderate salinity (~12 ‰) and high nitrogen conditions. The NH4+ and NO3− uptake characteristic was studied to understand the potential mechanism of G. tenuistipitata bloom. NH4+ uptake was best described by a linear, rate-unsaturated response, with the slope decreasing with time intervals. In contrast, NO3− uptake followed a rate-saturating mechanism best described by the Michaelis-Menten model, with kinetic parameters Vmax = 37.2 µM g−1 DM h−1 and Ks = 61.5 µM. Further, based on the isotope 15N tracer method, we found that 15N from NH4+ accumulated faster and reached an atom% twice than that of 15N from NO3−, suggesting when both NH4+ and NO3− were available, NH4+ was assimilated more rapidly. The results of the present study indicate that in the estuarine environment, the combination of moderate salinity with high ammonium may stimulate bloom formation.