Symbiosis modulates pharmaceutical toxicity and contaminant fate in the Azolla-Nostoc system

Pharmaceutical contamination in aquatic environments frequently occurs as complex mixtures; however, the extent to which biological interactions modify the toxicity and fate of contaminants remains poorly understood. Here, we investigated the physiological, biochemical, ecotoxicological, and phytoremediation responses of Azolla filiculoides cultivated in the presence (A+N) or absence (A-N) of Nostoc flagelliforme exposed to hydroxychloroquine (HCQ), ivermectin (IVE), and azithromycin (AZI), applied individually across environmentally relevant to elevated exposure concentrations (0-200 μg L^-1) and in fixed-ratio mixtures (2 μg L^-1 per compound). IVE induced the strongest physiological disruption, impairing photosynthetic performance, increasing oxidative stress, and destabilizing nitrogen metabolism, whereas HCQ produced intermediate effects and AZI caused comparatively weaker responses. Symbiosis was associated with higher ecotoxicological thresholds for several physiological endpoints, with threshold values differing by up to two orders of magnitude between A-N and A+N plants. Compared with A-N plants, A+N plants generally maintained higher photosynthetic performance, lower oxidative damage, and higher nitrogen-related metabolite concentrations, including elevated NH₄⁺, glutamate, glutamine, and total-N concentrations. Symbiosis was also associated with differences in pharmaceutical fate in a compound-specific manner. For AZI and IVE, enhanced removal efficiency coincided with lower tissue accumulation, whereas HCQ exhibited both greater removal and higher internal retention under symbiotic conditions. Collectively, these findings indicate that symbiosis modulates pharmaceutical responses, ecotoxicological sensitivity, and contaminant fate in a compound-specific manner, highlighting the importance of biological interactions in shaping ecotoxicological responses and pharmaceutical attenuation in aquatic plant-microbe systems.