Ammonia Stress Disturbs Moult Signaling in Juvenile Swimming Crab Portunus trituberculatus

SIMPLE SUMMARY: Ammonia is the most common contaminant in aquaculture systems, including intensive hatchery systems for crustaceans. Although it has been demonstrated that ammonia can result in abortive moulting and massive mortality of juvenile crustaceans, the underlying mechanisms are still unknown. The present work is aimed at examining the effect of ammonia exposure on the moulting process as well as molt signaling in the swimming crab Portunus trituberculatus, an important aquaculture species in China. We examined the survival rate and moulting rate of the juvenile crabs (C2) and analyzed the expression pattern of the genes in key components of molt signaling during a complete moulting cycle under different concentrations of ammonia. Our results revealed that ammonia has dose-dependent biphasic effects on moulting in juvenile swimming crab. Specifically, low levels of ammonia (5 mg/L) stimulated moulting, while high levels of ammonia (20 mg/L) suppressed the moulting process and caused moulting death syndrome (MDS). The gene expression analysis indicated that low levels of ammonia can reduce the expression of MIH, which encodes the key negative regulator of moulting, and trigger ecdysteroid biosynthesis and ecdysteroid signaling in the juvenile crabs. In contrast, though the high level of ammonia increased MIH expression, it still resulted in excessive ecdysteroids and over-activation of ecdysteroid signaling, which may contribute to the depressed moulting and MDS in the juvenile crabs. The novel findings of this study improve the understanding of ammonia toxicity in brachyura and provide valuable information for hatchery management of P. trituberculatus. ABSTRACT: Ammonia is a significant concern during hatchery culture in brachyuran species, and its accumulation may lead to abortive moulting and large-scale deaths of the early juveniles. To date, the underlying mechanism for ammonia-induced alteration of the moulting process is still unknown. In this study, we aimed to investigate the effects of ammonia on the moulting as well as the potential mechanisms in early juveniles of the swimming crab Portunus trituberculatus, an important aquaculture species in China. We evaluated the survival rate and moulting rate of the juvenile crabs (C2) and analyzed the expression pattern of the genes in key components of molt signaling during a complete moulting cycle under different concentrations of ammonia nitrogen (the control group: <0.1 mg/L; the LA group: 5 mg/L; and the HA group: 20 mg/L). The results showed that: (1) the survival rate in the LA and HA groups was lower than that in the control group at the end of the experiment, and moulting death syndrome (MDS) was only observed in the HA group; (2) the moulting rate was higher in the LA group and lower in the HA group compared to the control group; (3) consistent with the results of the moulting experiment, MIH showed decreased expression, and genes related to ecdysteroid synthesis, ecdysteroid receptors, and responsive effectors exhibited increased expression in the LA group compared to the control group; and (4) although MIH expression was upregulated, increased expression of the genes associated with ecdysteroid synthesis, ecdysteroid receptors and downstream effectors still observed in the HA group. Our results indicated that low levels of ammonia can promote moulting in juvenile swimming crabs by inhibiting the expression of MIH and activating moult signaling, whereas high levels of ammonia inhibit moulting and lead to MDS through impairing moult signaling.