Phytoplankton communities in the proposed water source area of the Western Route of the South-to-North Water Diversion Project showed multi-level responses across monitoring-period groups and diversion areas. Based on 64 valid samples, total biomass ranged from 0.027 to 5.659 mg L⁻¹ and showed no consistent differences between monitoring-period groups or diversion areas, indicating site- and sampling-period-scale patchiness. Among the dominant biomass-contributing taxa, most were diatom taxa, and the relative contributions of the top ten dominant taxa and Other taxa were reorganized among monitoring-period–area combinations. NMDS, PERMANOVA, and PERMDISP showed that monitoring period was significantly associated with community structure, whereas diversion-area effects were not significant. dbRDA indicated significant environmental–spatial constraints on community composition, with an adjusted explanatory power of 28.2%; T, NH₄⁺–N, TN, NO₃⁻–N, EC, pH, DO, and DTN were significant predictors. VPA showed stronger pure environmental than pure spatial effects, while DDR and EDR revealed significant geographic and environmental distance relationships. Taxonomic Bray–Curtis null models suggested a predominance of stochastic-like taxonomic turnover signatures, with stronger deterministic-like deviations in the upper-line diversion area. GAM identified NH₄⁺–N, DO, and EC as significant biomass predictors. These findings support integrating biomass, community composition, measured physicochemical variables, and taxonomic assembly signatures into sustainability-oriented phytoplankton monitoring for high-elevation riverine water source areas, thereby providing ecological evidence for sustainable water source protection and adaptive management.