Evaluation of Potential Probiotics Bacillus subtilis WB60, Pediococcus pentosaceus, and Lactococcus lactis on Growth Performance, Immune Response, Gut Histology and Immune-Related Genes in Whiteleg Shrimp, Litopenaeus vannamei

Abstract

An eight-week feeding trial was conducted to evaluate the effects of different dietary probiotic supplements in juvenile whiteleg shrimp, Litopenaeus vannamei. A basal control diet without probiotics (CON), and five other diets by supplementing Bacillus subtilis at 10⁷ CFU/g diet (BS₇), B. subtilis (BS₈), Pediococcus pentosaceus (PP₈), and Lactococcus lactis (LL₈) at 10⁸ CFU/g diet, and oxytetracycline (OTC) at 4 g/kg diet were used. Whiteleg shrimp with initial body weights of 1.41 ± 0.05 g (mean ± SD) were fed with these diets. Growth of shrimp fed BS₈ and LL₈ diets was significantly higher than those of shrimp fed the CON diet (p < 0.05). Superoxide dismutase activity in shrimp fed PP₈ and LL₈ diets was significantly higher than that of shrimp fed the CON diet (p < 0.05). Lysozyme activity of shrimp fed probiotics and OTC diets significantly improved compared to those on the CON diet (p < 0.05). The intestinal histology showed healthier guts for shrimp fed the probiotic diets (p < 0.05). Immune-related gene expression in shrimp fed BS₈, PP₈ and LL₈ diets was recorded as significantly higher than that of shrimp fed CON and OTC diets (p < 0.05). Also, results of the challenge test for 7 days and the digestive enzyme activity of shrimp fed BS₈, PP₈, and LL₈ were significantly improved compared to those on the CON diet (p < 0.05). Therefore, these results indicated that L. lactis at 10⁸ CFU/g could be an ideal probiotic for whiteleg shrimp, and also B. subtilis WB60 and P. pentosaceus at 10⁸ CFU/g could improve the growth, immunity, histology, gene expression, digestive enzyme activity, and disease resistance, while replacing antibiotics.

Keywords: Bacillus; Lactococcus; Pediococcus; Probiotic; feed additive; whiteleg shrimp.

Figure 1

Histological sections of juvenile whiteleg shrimp intestine fed the experimental diets for 8 weeks. A-CON, basal diet; B-BS₇, Bacillus subtilis at 1 × 10⁷ CFU/g; C-BS₈, Bacillus subtilis at 1 × 10⁸ CFU/g; D-PP₈, Pediococcus pentosaceus at 1 × 10⁸ CFU/g; E-LL₇, Lactococcus lactis at 1 × 10⁸ CFU/g; F-OTC, oxytetracycline at 4 g/kg. (Scale bar = 100 µm; Original magnification × 4). ML = muscular layer thickness VH = villi height.

Figure 2

Intestinal gene expression levels of serine protease, peroxinectin and prophenoloxidase were evaluated in juvenile whiteleg shrimp fed the experimental diets for 8 weeks. Bars with range represent mean ± SD of triplicate samples, and diets refer to Figure 1.

Figure 3

Cumulative survival rate of juvenile whiteleg shrimp fed the experimental diets with different probiotics for 8 weeks and experimentally challenged with V. parahaemolyticus for 7 days. Each value represents mean of triplicate groups. Significant differences among means are indicated by different superscripts (p < 0.05), and diets refer to Figure 1.

Figure 4

Specific enzyme activities of 1. Trypsin, 2. Amylase and 3. Lipase measured in the intestines of juvenile whiteleg shrimp fed the experimental diets with different probiotics for 8 weeks, and diets refer to Figure 1.

Figure 4

Specific enzyme activities of 1. Trypsin, 2. Amylase and 3. Lipase measured in the intestines of juvenile whiteleg shrimp fed the experimental diets with different probiotics for 8 weeks, and diets refer to Figure 1.