| In recent years, the farming yield of Litopenaeus vannamei has been steadily increasing year by year, positioning it as the highest-producing economic crustacean in global aquaculture. However, with the ongoing depletion of fishmeal resources and the fluctuation of its prices, the need for alternative protein sources has become more pressing. As a result, plant protein has emerged as a promising and sustainable replacement for fishmeal, becoming the primary protein source in aquaculture feeds. Despite its potential, the absorption and utilization of plant protein by aquatic animals, including shrimp, has been a significant challenge, as they are not naturally adapted to efficiently digest plant-based proteins. To overcome this limitation, probiotic fermentation of plant protein has become the leading method to enhance the digestibility and bioavailability of plant-based protein for aquatic species, ultimately improving the overall feed efficiency and growth performance of aquaculture animals. In this study, a strain of Bacillus subtilis was isolated and identified from the intestine of L. vannamei, with a similarity of 99.20% to Bacillus subtilis subsp. subtilis strain 168. It was named SQVGB8. This strain is a Gram-positive bacterium, non-hemolytic, and sensitive to the majority of antibiotics. The characteristic analysis results showed that the strain can efficiently produce protease, lipase, amylase, cellulase, and phytase; inhibit Vibrio harveyi, Vibrio alginolyticus, and Vibrio parahaemolyticus; secrete active substances such as SCFAs and EPS; and also exhibit high intestinal adhesion properties.
The whole genome analysis results showed that the strain SQVGB8 possesses the ability to produce digestive enzymes, phytase, SCFAs, and EPS. Additionally, SQVGB8 can secrete various substances with antibacterial and antimicrobial potential, including surfactin, fengycin, and bacillibactin. These active substances have significant application prospects in antibacterial and pathogen control. The suitable fermentation conditions for the strain SQVGB8 fermenting plant meals (soybean meal: cottonseed meal = 3:1) were determined through a single-factor experiment. The optimal range of conditions is as follows: inoculum amount 3%~12%, fermentation temperature 35~45 ℃, fermentation time 36~72 h, and solid-to-liquid ratio 1:1.2~1:0.6 g/mL. Therefore, a response surface methodology (RSM) with a 4-factor, 3-level analysis was used, and the optimal fermentation conditions for SQVGB8 fermenting plant meals were determined to be an inoculum amount of 7.70%, a fermentation temperature of 35.70 ℃, a fermentation time of 55.00 h, and a solid-to-liquid ratio of 1:0.78 g/mL. The acid soluble protein and acidic polysaccharide contents were 10.30% and 12 mg/mL, respectively. Compared to the unfermented plant meals, the content of antinutritional factors (ANFs) was significantly reduced. The trypsin inhibitor (TI) content was reduced by 91.67%, soybean globulin decreased by 61.57%, β-conglycinin decreased by 76.75%, gossypol content decreased by 59.00%, and phytic acid content decreased by 48.72%. It can be concluded that the strain SQVGB8 is an efficient enzyme-producing, active substance-secreting, and gut-colonizing probiotic, with high efficacy in fermenting plant meals. It has great potential for development and application in aquaculture feed. |
