D3337: Exploring mitochondrial disruptions in Tribolium castaneum under carbonyl sulfide fumigation

Carbonyl sulfide (COS) has recently emerged as a promising alternative to phosphine for managing stored-product insect pests, yet its subcellular mechanisms of toxicity remain poorly understood. In this study, we performed transmission electron microscopy (TEM), mitochondrial complexes and antioxidant enzyme activity assays to investigate the mitochondrial toxicity induced by COS fumigation in T. castaneum. COS exposure significantly altered mitochondrial ultrastructure, as demonstrated by matrix rarefaction, and vacuolization. These structural disruptions were accompanied by a decline in ATP content and enhanced reactive oxygen species (ROS) production. Notably, the NAD+ levels increased without significant changes in NADH and NAD+/ NADH ratio, indicating a potential redox imbalance under COS-induced stress. Biochemical analysis revealed inhibition of Complex III activity, with compensatory upregulation of Complex II (succinate dehydrogenase, SDH) and Complex IV (cytochrome c oxidase, COX), suggesting a disrupted but adaptive mitochondrial response. Furthermore, COS exposure disrupted antioxidant enzyme activities, where superoxide dismutase (SOD) remained unchanged, while peroxidase (POD) was elevated and catalase (CAT) activity was significantly suppressed, thereby highlighting impaired hydrogen peroxide detoxification. Additionally, we analyzed the impact of acetazolamide, a CA inhibitor, on the survival rates of T. castaneum following COS exposure, suggesting that acetazolamide may reduce COS toxicity by inhibiting CA activity. Furthermore, RNAi was employed to silence the CA3 gene, exploring its role in CA-mediated COS metabolism in T. castaneum. These findings are the first comprehensive study of COS toxicity in T. castaneum, offering novel insights into its toxic mode of action and supporting its potential as a novel fumigant alternative.