BackgroundSea cucumbers (Apostichopus japonicus) are an imperiled fauna exposed to a variety of environmental condition such as salinity and studies are urgently needed to assess their effects to guide aquaculture efforts. The effects of acute salinity stress on coelomic fluid osmotic pressure, ion concentrations, the activity of Na+/K+-ATPase in respiratory trees and the histological variations were measured to evaluate the salinity tolerance of sea cucumbers.ResultsSignificant correlations in osmotic pressure were observed between coelomic fluid and ambient environmental salinity. In coelomic fluid, Na+ concentration was observed fluctuated during salinity 18 psu and the inflection point presented at the 6 h. The Na+/K+-ATPase activity in respiratory trees indicated the “U-shaped” fluctuant change and the change trend was opposite with the Na+ concentration. The ions (K+, Cl−) concentration decreased and showed the same tendency at salinity 40 psu with salinity 18 psu. The total coelomocytes counts and phagocytosis of coelomic fluid Na+/K+-ATPase activity indicated fluctuating changes under different salinity stress. Histological variation revealed a negative relation between decreasing salt concentration and tissue integrity. Tissue damages were significantly observed in intestines, muscles and tube feet under low salinity environment (18, 23 and 27 psu). The connective tissue in intestines of A. japonicus exposed to 18 and 23 psu damaged and partly separated from the mucosal epithelium. The significant variations occurred in tube feet, which presented the swelling in connective tissue and a fracture in longitudinal muscles under low salinity (18 psu). The morphological change of tube feet showed the shrinkage of connective tissue under high salinity (40 psu). The amount of infusoria in the respiratory trees decreased or even disappeared in salinity treatment groups (18 and 23 psu).ConclusionThe results inferred that osmoconformity and ionoregulation were seen in sea cucumbers, which contributed to understand the salinity regulatory mechanisms of A. japonicus under acute salinity stress.

The lysozyme gene was silenced using RNA interference (RNAi) to analyze the function of lysozyme in sea cucumber under salt stress. The interfering efficiency of four lysozyme RNAi oligos ranged from 0.55 to 0.70. From the four oligos, p-miR-L245 was used for further interfering experiments because it had the best silencing efficiency. Peristomial film injection of p-miR-L245 (10 μg) was used for further interfering experiments. The lowest gene expression, determined by RT-PCR assay, in muscle, coelomic fluid, and parapodium occurred 48 h after p-miR-L245 injection, while that of body wall and tube foot was 96 h and 24 h, respectively. Lysozyme activity in muscle and body wall was significantly lower than the controls. The lowest lysozyme activity in muscle, body wall and parapodium, was found at 48, 72, and 48 h, respectively, which was consistent with the transcription expression of lysozyme. The lowest point of lysozyme activity was at 96 h in coelomic fluid and tube foot, which was laid behind lysozyme expression in transcription level. The expression profile of the lysozyme transcription level and lysozyme activity in the body wall and tube foot increased at 12 h after p-miR-L245 injection before the interference effect appeared. NKA gene expression was expressed at a high level in the positive control (PC) and negative control (NC) groups at 12, 24, and 48 h, while NKA was expressed at low levels in the lysozyme RNAi injection group at 12 and 24 h. The level of NKA gene expression recovered to the level of the PC and NC group at 48, 72, and 96 h after the lysozyme RNAi injection. NKCC1 gene expression was high in the PC and NC groups at 96 h, while the NKCC1 was expressed at a low level 96 h after lysozyme RNAi injection. The results suggest that lysozyme decay involves NKA and NKCC1 gene expression under salinity 18 psμ. The K and Cl concentration after lysozyme RNAi injection was lower than in the PC and NC group.