Non-Invasive Characterization of Locomotor and Ventilatory Responses in Rainbow Trout Under Acute Ammonia Nitrogen Stress.

Ammonia nitrogen is one of the most common environmental stressors in aquaculture water environments, and its accumulation can induce physiological disturbance, altered ventilation regulation, and abnormal behavioral responses in fish. To achieve non-invasive quantitative characterization of rainbow trout responses to ammonia nitrogen stress, this study developed a computer-vision-based framework for the integrated analysis of locomotor behavior and ventilation activity. Rainbow trout were exposed to four ammonia nitrogen concentrations: 0, 15, 30, and 60 mg/L. A total of 16 rainbow trout were used in this study, with an average body length of 14.0 ± 1.0 cm and an average body weight of 38.65 ± 2.42 g. The fish were assigned to four experimental aquaria, with four fish maintained in one aquarium for each TAN treatment. Stereo videos for locomotor behavior analysis and monocular mouth-region videos for ventilation analysis were simultaneously collected, and the final 5 min of each recording was analyzed. YOLOv11n, multi-object tracking, and stereo vision were used to extract three-dimensional position sequences of rainbow trout and calculate the amount of exercise, average swimming speed, and spatial distribution. Meanwhile, optical-flow analysis was applied to quantify mouth opening-closing motion and estimate ventilation frequency. The results showed that with increasing ammonia nitrogen concentration, rainbow trout locomotor behavior tended to be suppressed, with average swimming speed showing the clearest decrease, whereas ventilation frequency continuously increased. Average swimming speed decreased from 3.83 cm/s in the 0 mg/L group to 1.03 cm/s in the 60 mg/L group, while ventilation frequency increased from 84.91 breaths/min to 133.43 breaths/min. Compared with locomotor indicators, ventilation frequency showed a more stable response to changes in ammonia nitrogen concentration. This study achieved the synchronous quantification of rainbow trout locomotor behavior and ventilation activity, revealing a differentiated response pattern characterized by enhanced ventilation and suppressed locomotor behavior under acute ammonia nitrogen stress. These findings provide a methodological reference for fish stress assessment and risk warning in aquaculture environments.