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Amylase‐producing bacteria could improve water quality contaminated by waste from feed residue and fish metabolism, thereby increasing the efficiency of aquaculture systems. The objective of this research was to screen and optimize fermentation conditions of a high amylase‐producing strain. Four amylase‐producing bacterial strains (named S458‐1, G05, H38 and B09) were isolated from a grass carp pond, and the strain S458‐1 showed the highest amylase‐producing ability, with 19.58 ± 0.38 mm hydrolysis circle diameter. The strain S458‐1 was identified as Bacillus cereus based on morphological identification, biochemical identification and 16S rDNA sequence analysis. The optimal culture medium formula included (in g/L) Ca2+ 0.8, Mg2+ 0.2, Mn2+ 0.4, Fe2+ 0.6, Al3+ 0.2, 1% soluble starch and 1% peptone. The optimal fermentation conditions were determined as initial pH 9, culture temperature 37°C, fermentation time of 60 hr and 2% inoculum. Under the optimal formula and condition, its enzyme activity increased from 32 U/ml to 173.01 U/ml, a 5.41‐fold increase. Surprisingly, our research found that the strain S458‐1 also had phosphorus degradation capabilities. Its phosphorus‐dissolving ability was both time‐ and concentration‐dependent. Thus, this study will make a contribution to the bacterial amylase based on the fermentation process and provide a theoretical basis for further research of aquatic probiotics.

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Aeromonas hydrophila is a common pathogen and causes huge economic losses in aquaculture; to date, we still have no efficient methods to control fish diseases caused by A. hydrophila. Our previous research found that a strain of Bacillus methylotrophicus WM-1 could effectively inhibit A. hydrophila KS02. In the current study, the safety of WM-1 and its protective effects against A. hydrophila in the aquaculture system were examined. Grass carp were selected and fed with WM-1 supplemented feed (0, 10 3 , 10 5 and 10 7 CFU/g feed, as CG, LG, MG and HG) for a period of 90 days. On day 30, grass carp were challenged with KS02 (WM-1+KS02 group) or saline (WM-1 group) as control. The growth performance and enzyme activities were measured; blood routine tests were conducted on day 90. In the WM-1 group, AKP activity in the serum was significantly reduced and SOD activity in the liver was significantly enhanced by WM-1 supplemented feed, while other monitored indices had no significant changes. In the WM-1+KS02 group, GPT activity in the serum and SOD activity in the liver were significantly enhanced, while AKP activity in the serum had no significant change except for the MG group. Survival rates of grass carp were advanced by WM-1 supplemented feed, reaching 70% after challenge with KS02. Overall, WM-1 could maintain the homeostasis of grass carp and protected them against A. hydrophila infection efficiently, which indicated WM-1 would be a probiotic candidate for use in aquaculture.

Microbial agents have been widely used in the Chinese aquaculture industry due to their advantages of enhancing the immunity of aquatic animal, improving disease resistance and improving the aquaculture environment. In previous studies, we found a highly antagonistic Aeromonas hydrophila strain, Bacillus methylotrophicus WM‐1, isolated from grass carp (Ctenopharyngodon idella) culture systems. The purpose of this study was to investigate the impact of WM‐1 on cultured water and fish. Strain WM‐1 was mixed into the feed and fed to the grass carp for 30 days, set control group (CG, 0 CFU/g feed), low concentration group (LG, 103 CFU/g feed), medium concentration group (MG, 105 CFU/g feed) and high concentration group (HG, 107 CFU/g feed). Water quality and physiological indicators of grass carp were tested. Compared with CG, the concentrations of total phosphorus (TP) and soluble reactive phosphorus (SRP) were significantly increased in the MG in the early stage (before day 18, p < 0.05), whereas decreased in the long stage. After the 12th day, treatment groups showed a significant decrease in chemical oxygen demand (COD) (p < 0.05). Total nitrogen (TN), ammonia nitrogen (NH4+‐N), nitrite (NO2‐‐N) and nitrate (NO3‐‐N) had no significant difference (p > 0.05). In grass carp serum, GPT activity in MG and HG was significantly reduced, SOD activity was significantly increased, and POD in MG was significantly higher than LG and HG (p < 0.05). In the liver, the GPT activity of the LG was significantly increased (p < 0.05). Other indicators (the specific growth rate and condition factor of grass carp, serum GOT, AKP and POD activity, total liver protein content, liver SOD, POD, AKP and GPT activity, haemocyte counts) have no significant difference between CG and treatment groups (p > 0.05). The results show that WM‐1 is safe for water quality and culture objects, and has the effects of improving water quality and the immunity of grass carp. The corrent results combined with WM‐1 effectively inhibit A. hydrophila shown that WM‐1 can be applied to aquaculture production as a probiotics.