High-throughput screening of combined toxicity of nanoplastics and coexisting pollutants using luminescent bacterium

Micro/nanoplastics (MNPs) commonly coexist with contaminants such as heavy metals and antibiotics in aquatic environments, potentially inducing complex joint toxicity. However, conventional approaches for MNP-pollutant combined toxicity assessment are typically challenged by inefficiencies, high costs, and labor-intensive procedures. In this study, a high-throughput platform using microplate-based inhibition assays with a luminescent bacterium (Vibrio qinghaiensis sp.-Q67), was applied to systematically evaluate the combined toxicity of four polystyrene nanoplastics (PS-NPs), varying in size and surface functionalization, with seven heavy metals and five antibiotics. Results showed that metal cations (Cu²⁺, Zn²⁺, Pb²⁺, Cd²⁺, and Cr³⁺) and chlortetracycline combined with PS-NPs primarily induced antagonistic or additive toxicity, whereas anions (Cr₂O₇^2- and AsO₂^-) and antibiotics (tetracycline, oxytetracycline, norfloxacin, and ciprofloxacin) exhibited synergistic or additive toxicity. Smaller PS-NPs (50 nm) amplified the toxicity interactions compared to 100 nm PS-NPs. Carboxyl-modified PS-NPs exhibited the most pronounced antagonistic effects with cations, while amino-modified PS-NPs favored amplified synergistic effects with Cr₂O₇^2-. The combined effects were intensified at higher PS-NP concentrations and at moderate co-contaminant concentrations (i.e., 1/5 or 1 × the median effect concentration). Antibiotics with higher water solubility exhibited more pronounced synergistic effects with PS-NPs, and the antagonism intensities for metal cations followed an order of Pb²⁺ > Cu²⁺ > Cr³⁺ > Zn²⁺ > Cd²⁺. This high-throughput strategy efficiently assessed multifactorial impacts on combined toxicity, providing systematic insights into the toxic interaction patterns between MNPs and pollutants.