Ammonia-induced oxidative stress triggered apoptosis in the razor clam (Sinonovacula constricta)

Abstract

As one of the most significant contaminants and stressors in aquaculture systems, ammonia adversely jeopardizes the health of aquatic animals. Ammonia exposure affects the development, metabolism, and survival of shellfish. However, the responses of the innate immune and antioxidant systems and apoptosis in shellfish under ammonia stress have rarely been reported. In this study, razor clams (Sinonovacula constricta) were exposed to different concentrations of non-ion ammonia (0.25 mg/L, 2.5 mg/L) for 72 h and then placed in ammonia-free seawater for 72 h for recovery. The immune responses induced by ammonia stress on razor clams were investigated by antioxidant enzyme activities and degree of apoptosis in digestive gland and gill tissues at different time points. The results showed that exposure to a high concentration of ammonia greatly disrupted the antioxidant system of the razor clam by exacerbating the accumulation of reactive oxygen species ( $O_2^{-}$ , H₂O₂) and disordering the activities of antioxidant enzymes (superoxide dismutase, catalase, and glutathione peroxidase), and the level of activity remained at a significantly high level after recovering for 72 h (P < 0.05). In addition, there were significant differences (P < 0.05) in the expression of key genes (Caspase 7, Cyt-c, Bcl-2, and Bax) in the mitochondrial apoptotic pathway in the digestive glands and gills of razor clams as a result of ammonia stress and were unable to return to normal levels after 72 h of recovery. TUNEL staining indicated that apoptosis was more pronounced in gills, showing a dose and time-dependent pattern. As to the results, ammonia exposure leads to the activation of innate immunity in razor clams, disrupts the antioxidant system, and activates the mitochondrial pathway of apoptosis. This is important for comprehending the mechanism underlying the aquatic toxicity resulting from ammonia in shellfish.

Keywords: Sinonovacula constricta; Ammonia; Antioxidant system; Apoptosis; Reactive oxygen species.