Increasing anthropogenic activities has caused intensive environmental issues and undesirable ecological impacts on coastal bay ecosystems. Bacterioplankton play critical roles in ecological functioning of the bay ecosystems, but much remains to be learned regarding the response of bacterioplankton community assembly to heterogeneous environmental issues in the bay ecosystems and its underlying mechanisms. In this study, we used the phylogenetic framework to analyze the bacterioplankton community assembly in the subtropical Daya Bay, where the habitats are connected by water flows and tides and their trophic status are subject to intensive environmental stress induced by human activities (e.g., thermal discharge and nutrient release from aquaculture). We found bacterioplankton community compositions (BCCs) among the sampling habitats in the Daya Bay showed obvious spatial heterogeneity. The spatial distributions of BCCs were significantly shaped by phosphate concentration among the examined environmental factors. We observed that between pairwise sampling habitats, the BCC dissimilarity significantly increased with the differences in seawater phosphate concentration. Compared with stochastic processes, phosphate enrichment imposed stronger effects of environmental filtering in determining bacterioplankton community assembly in the subtropical bay, and bacterioplankton communities tended to be higher phylogenetically clustered in more phosphate-enriched habitats. In summary, we propose that phosphate is a major environmental determinant in the subtropical Daya Bay impacted by thermal discharge and regulates the interplay between deterministic and stochastic processes underlying bacterioplankton community assembly.