• • At 1.0 μg·L−1 acetamiprid, head swing frequency increased by 23.86% vs. control, indicating neurobehavioral disruption at environmentally relevant concentrations, a sensitive endpoint for soil nematode risk assessment.
• • F0 exposure elevated ROS by 30.42%–48.28% across 1–100 μg·L−1, linking oxidative stress to reproductive, developmental, and intestinal toxicity, providing a mechanistic basis for biomarker selection.
• • Body width inhibition persisted to T2 generation at 1.0 μg·L−1, demonstrating transgenerational growth effects at low doses, critical for chronic soil exposure scenarios.
• • SOD activity exhibited the highest sensitivity and transgenerational persistence among oxidative stress biomarkers, suggesting SOD as a robust early-warning indicator for neonicotinoid soil contamination.