Combined thermal and 6PPD-quinone stressors alter biological endpoints, oxidative stress responses, and transcriptomic profiles in the rotifer Brachionus plicatilis

In this study, we investigated the single and combined effects of 6PPD-quinone (50 µg/L) and elevated temperature (30 °C) on fecundity, lifespan, oxidative stress, antioxidant enzyme activities (CAT, SOD, and GST), and transcriptomic responses in the euryhaline rotifer Brachionus plicatilis. Exposure to 6PPD-quinone alone reduced fecundity, while elevated temperature alone reduced lifespan. Combined exposure decreased both fecundity and lifespan, although no additive effects were observed for these life-history endpoints. In contrast, combined exposure significantly increased reactive oxygen species (ROS) production and antioxidant enzyme activities compared with the control. Two-way ANOVA and the stress addition model (SAM) identified synergistic interactions between elevated temperature and 6PPD-quinone for ROS production and CAT activity, whereas GST and SOD exhibited additive responses. Transcriptomic analysis revealed that combined exposures induced the greatest number of differentially expressed genes, followed by elevated temperature alone and 6PPD-quinone alone. RNA-seq pathway analysis identified mitochondrial dysfunction and NRF2-mediated oxidative stress response as the most significantly dysregulated pathways, indicating pronounced oxidative stress under combined exposure. These findings demonstrate that warming can amplify the molecular and physiological toxicity of 6PPD-quinone and highlight the importance of evaluating multiple environmental stressors to improve ecological risk assessments under future climate change scenarios.