Wastewater treatment plants (WWTP) are regarded as the last barriers for the release of incompletely separated and recycled nanoparticles (NPs) into the environment. Despite the importance and ubiquity of microbial extracellular polymeric substances (EPSs) in the complex wastewater matrix, the interaction between NPs and EPSs of anaerobic microflora involved in wastewater treatment and the resultant impact on the biomass metabolomics are unclear. Thus, the impacts of different metal oxide (TiO2, ZnO, and CuO) NPs on functional bacteria in anaerobic granular sludge (AGS) and the possible toxicity mechanisms were investigated. In particular, the binding quality, enhanced resistance mechanism, and chemical fractional contribution of EPSs from AGS against the nanotoxicity of different NPs was assessed. The results showed that CuO NPs caused the most severe inhibition towards acetoclastic and hydrogenotrophic methanogens, followed by ZnO NPs, whereas TiO2 NPs caused no inhibition to methanogenesis. Excessive EPS production, especially the protein-like substances, was an effective strategy for reducing certain NPs’ toxicity by immobilizing NPs away from AGS cells, whereas the metabolism restriction on inner microorganisms of AGS induced by CuO NPs can deteriorate the protective role of EPS, indicating that the roles of EPS may not be amenable to generalizations. Further investigations with lactate dehydrogenase (LDH) and reactive oxygen species (ROS) assays indicated that there are greatly essential differences between the toxicity mechanisms of metal NPs to AGS, which varied depending on the NPs’ type and dosage. In addition, dynamic changes in the responses of EPS content to different NPs can result in a significant shift in methanogenic and acidogenic microbial communities. Thus, the production and composition of EPSs will be a key factor in determining the fate and potential effect of NPs in the complex biological matrix. In conclusion, this study broadens the understanding of the inhibition mechanisms of metal oxide NPs on the AGS process, and the influence of EPSs on the fate, behavior, and toxicity of NPs.

Antibiotics in aqueous environments can have extremely adverse effects on non-targeted organisms. However, many research projects have only focused on the toxicological evaluation of individual antibiotics in various environments. In the present work, individual and binary mixture toxicity experiments have been conducted with the model organism Raphidocelis subcapitata (R. subcapitata), and a mixture concentration-response curve was established and contrasted with the estimated effects on the basis of both the concentration addition (CA) and the independent action (IA) models. In addition, different risk assessment methods were used and compared to evaluate the environmental risk of binary mixtures. The toxic ranking of the selected antibiotics to R. subcapitata was erythromycin (ERY) > sulfamethoxazole (SMX) > sulfamethazine (SMZ). In general, the conclusion of this study is that the adverse effects of binary mixtures are higher than the individual antibiotics. The CA model and RQSTU are more suitable for toxicity prediction and risk assessment of binary mixtures. This study reveals the potential ecological risks that antibiotics and their mixtures may pose to water ecosystems, thus providing scientific information for environmental quality regulation.