ABSTRACT: Despite increasing application of microalgae for antibiotic-contaminated aquaculture wastewater treatment, how salinity regulates antibiotic transformation depth and cellular adaptation remains unclear. This study combined salinity-gradient cultivation, process partitioning, transformation-product profiling, physiological assays, transcriptomics, and metabolomics to elucidate salinity-dependent norfloxacin (NFX) transformation by the halotolerant Chlorella sp. AMC-2. AMC-2 maintained growth at 1–15 ppt, whereas 25–35 ppt inhibited biomass accumulation. Compared with 1 ppt, the algal biomass at 15 ppt decreased by 23.2% on day 9, while NFX removal declined from 52.0% to 46.8%. Biotransformation became dominant earlier at 15 ppt, contributing 58.4% of NFX removal on day 5 versus 26.0% at 1 ppt. Transformation-product profiling showed comparable primary reactions at both salinities but restricted downstream ring cleavage and scaffold disruption at 15 ppt. At 15 ppt, ETRmax decreased by approximately 40%, while antioxidant defenses were enhanced and bound extracellular polymeric substances increased by approximately 38%. Genes related to proline biosynthesis (ALDH18A1, P5CS, proB, and proC) and antioxidant defense (SOD2, PRDX5, and GST) were upregulated, whereas genes associated with photosynthetic electron transport and carbon fixation (petA, petF, rbcL, and rbcS) were downregulated. Proline, ornithine, cadaverine, trehalose, and ADP accumulated, while TCA-cycle intermediates decreased. Together, physiological adaptation sustained primary NFX biotransformation under moderate salinity, whereas constrained electron transport, carbon fixation, and energy metabolism limited deeper structural degradation. These findings reveal a salinity-dependent divergence between parent NFX removal and transformation depth, providing a mechanistic basis for assessing microalgal treatment performance and transformation-product risks across freshwater, brackish, and marine aquaculture wastewater.
