Effects of diets supplemented with either individual or combined probiotics, Bacillus subtilis E20 and Lactobacillus plantarum 7-40, on the immune response and disease resistance of the mud crab, Scylla paramamosain (Estampador) Abstract Two trials, including firstly, diets incorporating individual or combined species of probiotics, Bacillus subtilis E20 and Lactobacillus plantarum 7-40, were fed to the mud crab, Scylla paramamosian, for 28 days to evaluate the growth, immunity and disease resistance, and secondly, the optimal level of B. subtilis E20 in the diet by tracking the growth, immunity and disease resistance after 28 days of feeding were analysed. No significant differences in growth, total haemocyte counts, respiratory bursts, superoxide dismutase, or glutathione peroxidase were found in the two trials. Crabs fed the B. subtilis E20-containing diet (D20) had significantly increased phenoloxidase (PO) activity, phagocytic activity (PA) and disease resistance compared with crabs fed the control and the mixed probiotics diet (MD). The mortality of crabs fed the D20 was significantly lower than that of crabs fed the L. plantarum 7-40-containing diet. L. plantarum 7-40 had a great inhibitory effect on the growth of B. subtilis E20, which may have led to the decreased probiotic effect of the MD. An analysis of the optimal level of B. subtilis E20 in the diet showed that crabs fed the B. subtilis E20-containing diet at a level of 10 9-10 cfu kg À1 had better immune response of PO and PA, and disease resistance in the second trial.

Six fish meal basal diets supplied with 0 (control), 0.25, 0.5, 1, 2.5 and 5 g kg À1 of choline chloride, resulting in choline levels of 2.57, 2.67, 2.94, 3.84, 4.99 and 7.71 g kg À1 , respectively, were fed to giant grouper, Epinephelus lanceolatus, for 56 and 30 days to evaluate the growth and lipid metabolism, and stress tolerance respectively. In the first trial, fish fed different levels of cholinecontaining diets for 56 days had no significant difference in weight gain, survival and feeding efficiency. Fish fed increased levels of dietary choline, however, tended to have decreases in the hepatic somatic index; lipids, triglycerides, and cholesterol in liver; and triglycerides and cholesterol in serum. A decrease of lipid content in dorsal muscle was recorded in fish fed the diets containing choline >2.94 g kg À1 . Additionally, dietary choline improved the reactions of fish to ammonia stress, including survival and behavioural responses, in fish fed diets containing choline levels >2.94 g kg À1 . These findings indicate that choline plays important roles in lipid metabolism and stress tolerance in giant grouper.

The purpose of this study was to use an electrolytic oxidizing (EO) water system in a prawn hatchery. The results of the EO water generation efficiency showed that available chlorine in the EO water increased as the salinity and electric power increased, but was reduced as the water flow rate increased. A bactericidal activity assay showed that the growth of six aquatic pathogens was well inhibited by incubation with EO water containing 1 mg L−1 of available chlorine. The removal of NH3‐N and NO2‐N by electrolysis at 15‰ and 5 W had significant effects, and the removal rate of NO2‐N was much faster than that of NH3‐N. Prawn zoea (Z) reared in EO water with different levels of available chlorine at 0 (control), 0.2, 0.3 and 0.4 mg L−1 had mortalities of 0%, 12%, 27% and 42%, respectively, after 72 h. However, Z were able to well survive in activated carbon‐treated EO water for 1 week. For the on‐farm use of the EO water system in a prawn hatchery, larvae in the EO water group had faster development and higher survival than those in the activated carbon group. From these results, we believe that the EO water group had excellent water quality and lower bacterial counts resulting in increased larval development and survival.

Bacillus subtilis E20, an isolate from natto – a Japanese fermented soy product – is used in a wide variety of marine and freshwater aquatic species to enhance growth and immunity. This paper describes the cloning, expression and characterization of xylanase isolated from B. subtilis E20 in B. subtilis RIK1285. The BsXynE20 gene was identified to encode a 213‐amino‐acid protein for a glycoside hydrolase family 11 xylanase containing a 28‐residue signal peptide whose cleavage yields a 185‐residue mature protein with a predicted molecular weight of 20.25 kDa. The full length of the BsXynE20 gene was subcloned into the pBE‐S expression vector for secretory production of recombinant BsXynE20 in B. subtilis RIK1285. Western blot and zymogram analysis, respectively, revealed that recombinant BsXynE20 can be secreted into culture medium and is a functional xylanase. The xylanase exhibited optimal activity at pH 5.0–6.0 and 60°C. Using recombinant BsXynE20‐pretreated duckweed meal for feed preparation improved the growth performance of tilapia. Finally, we achieved secretive expression of functional BsXynE20, which can improve the nutrient utilization of a plant‐derived diet. Thus, the use of BsXynE20 may be beneficial for tilapia aquaculture.