Apparent pH-TAN operational breakpoint links duckweed-microbiome destabilization and nitrogen fate shift in ammonia-rich swine digestate

Ammonia-rich agricultural wastewaters challenge reliable biological treatment because treatment failure can leave reactive nitrogen in the dissolved phase. Duckweed-based systems offer a low-energy approach for polishing nutrient-rich wastewaters, but their stability under ammonia stress remains poorly resolved. Here, we integrated physiological, physicochemical, microbiome, nitrogen-cycling gene, and 15 N-tracing analyses with bacterial–microalgal intervention to examine treatment responses in a duckweed-based swine digestate system. Duckweed responses showed a threshold-like transition, with an apparent pH–TAN operational breakpoint corresponding to a calculated FA concentration of approximately 8.5 mg NH₃-N/L under the tested conditions. Beyond this operational breakpoint, duckweed performance and the associated O₂/pH conditions declined, bacterial diversity and community composition shifted, and selected nitrogen-cycling genes decreased in DNA-level abundance. High-FA conditions were associated with slower NH₄⁺-N removal, reduced NO₃⁻-N formation, and redistribution of tracer N from plant- and microbial-associated pools toward the residual dissolved pool. The combined bacterial-microalgal intervention produced the strongest partial recovery of treatment performance and biological nitrogen retention, although microalgal effects cannot be separated from the associated aeration. These findings identify a condition-specific pH–TAN operational breakpoint and highlight pH, ammonia speciation, and aeration as important co-determining factors in ammonia-rich wastewater treatment.