Individual and mixture effects of the antibiotics sulfamethazine and tetracycline on freshwater ecosystems

Antibiotics frequently co-occur in aquatic environments, yet their combined ecological effects remain insufficiently understood under realistic mixture scenarios. To address this knowledge gap, we established 36 replicated freshwater microcosms and exposed them to two commonly detected antibiotics, sulfamethazine (SMT) and tetracycline (TC), both individually and in combination, at environmentally realistic concentrations over a 28-day period. At the primary producer level, both antibiotics reduced green algal abundance at intermediate and high concentrations alone, whereas their combination exposure did not consistently inhibit growth, suggesting altered ecological effects under mixture exposure. Zooplankton community structure was significantly altered under both individual and mixture treatments, with stimulatory as well as adverse effects observed at different dose levels. Microbial communities responded in a compartment-specific manner. Single-compound exposures significantly shifted community structure in both water column and sediment. In contrast, no significant changes were detected in water-column microbial communities under mixture treatments, suggesting antagonistic interactions between SMT and TC, likely related to their distinct modes of action and compensatory microbial regulation. At the functional level, leaf-litter decomposition was consistently accelerated across multiple treatments, indicating enhanced microbial activity and functional stimulation. Collectively, these findings provide experimental evidence that antibiotic mixtures can exert non-additive effects across trophic levels and ecosystem processes, highlighting the importance of accounting for such interactions in future research and risk assessment frameworks.