Abstract：
This study investigated the effects of two distinct probiotics, Leuconostoc mesenteroides B4 (B4) and Bacillus pumilus D5 (D5), along with their combination, on the diet of white shrimp (Litopenaeus vannamei) during an eight-week feeding trial. The diets tested included B4 + dextran at 107 CFU/g feed (the B4 group), D5 alone at 107 CFU/g feed (the D5 group), and a combination of B4 + dextran and D5 at 5 × 106 CFU/g feed each (the B4+dextran + D5 group). Relative to the control group, those administered probiotics exhibited moderate enhancements in growth. By the eighth week, the weight gain for the B4, D5, and B4+D5 groups was 696.50 ± 78.15%, 718.53 ± 130.73%, and 693.05 ± 93.79%, respectively, outperforming the control group\'s 691.66 ± 31.10% gain. The feed conversion ratio was most efficient in the B4 group (2.16 ± 0.06), closely followed by B4+D5 (2.21 ± 0.03) and D5 (2.22 ± 0.06), with the control group having the highest ratio (2.27 ± 0.03). While phenoloxidase activity was somewhat elevated in the B4 and D5 groups, no significant differences were noted in respiratory burst activity or total hemocyte count across all groups. Challenge tests at weeks 4 and 8 showed that the B4 + D5 combination offered superior protection against AHPND-causing Vibrio parahaemolyticus. The 4-week cumulative survival rate was highest in shrimp treated with B4 + dextran + D5 (56.25%), followed by B4 + dextran (31.25%), control (18.75%), and lowest in D5 (12.5%). By week 8, the B4 + dextran + D5 (43.75%) and B4 + dextran (37.5%) groups significantly outperformed the control group (6.25%, p < 0.05), with no significant difference observed between the D5 group (37.5%) and the control group at day 56. Analysis of the shrimp\'s foregut microbiota revealed an increase in unique OTUs in the B4 and B4 + D5 groups. Compared to the control, Proteobacteria abundance was reduced in all probiotic groups. Potential pathogens like Vibrio, Bacteroides, Neisseria, Botrytis, Clostridioides, and Deltaentomopoxvirus were detected in the control but were reduced or absent in probiotic groups. Beneficial microbes such as Methanobrevibacter and Dictyostelium in the B4+D5 group, and Sugiyamaella in the B4 group, showed significant increases. Probiotics also led to higher transcript levels of nitric oxide synthase in the hemocytes, and lysozyme and transglutaminase in the midgut, along with lysozyme and α2-macroglobulin in the foregut. Notably, the combined B4 + D5 probiotics synergistically enhanced the expression of superoxide dismutase and prophenoloxidase in the foregut, indicating an improved immune response. In summary, this study demonstrates that the probiotics evaluated, especially when used in combination, significantly boost the expression of specific immune-related genes, enhance the bacterial diversity and richness of the intestine, and thus prevent the colonization and proliferation of Vibrio spp. in L. vannamei.
摘要：
这项研究调查了两种不同的益生菌的作用，中肠明串珠菌B4(B4)和短小芽孢杆菌D5(D5),连同他们的组合，在为期八周的喂养试验中，对虾（凡纳滨对虾）的饮食。测试的饮食包括B4+葡聚糖107CFU/g饲料（B4组），以107CFU/g饲料单独使用D5（D5组），和B4+葡聚糖和D5的组合,各5×106CFU/g饲料(B4+葡聚糖+D5组)。相对于对照组，那些服用益生菌的人表现出适度的生长增强。到第八周,B4，D5和B4+D5组的体重增加为696.50±78.15%，718.53±130.73%，693.05±93.79%，分别,优于对照组的691.66±31.10%的增益。B4组的饲料转化率最高（2.16±0.06），紧随其后的是B4+D5(2.21±0.03)和D5(2.22±0.06)，与对照组的比例最高(2.27±0.03)。虽然B4和D5组的酚氧化酶活性有所升高，所有组的呼吸爆发活动或总血细胞计数均无显著差异.在第4周和第8周的攻击测试表明，B4D5组合对引起AHPND的副溶血性弧菌具有出色的保护作用。B4+葡聚糖+D5处理的虾4周累积存活率最高(56.25%),其次是B4+葡聚糖(31.25%)，控制（18.75%），D5最低（12.5%）。到第8周，B4+葡聚糖+D5（43.75%）和B4+葡聚糖（37.5%）组显著优于对照组（6.25%，p<0.05)，在第56天，D5组（37.5％）和对照组之间没有显着差异。对虾的前肠微生物群的分析显示，B4和B4D5组中独特的OTU增加。与对照相比，在所有益生菌组中,变形杆菌丰度降低。潜在的病原体，如弧菌，拟杆菌,奈瑟菌,葡萄孢菌,梭菌,和Deltaentomopoxvirus在对照组中检测到，但在益生菌组中减少或不存在。B4+D5组的有益菌，如甲那巴杆菌和网菌属，B4组的Sugiyamaella,显着增加。益生菌还导致血细胞中一氧化氮合酶的转录水平更高，中肠中的溶菌酶和转谷氨酰胺酶，与溶菌酶和α2-巨球蛋白一起在前肠中。值得注意的是,B4+D5益生菌联合协同增强前肠超氧化物歧化酶和酚氧化酶原的表达,表明免疫反应改善。总之,这项研究表明，益生菌评估，特别是当组合使用时，显著促进特定免疫相关基因的表达，增强肠道的细菌多样性和丰富度，从而防止弧菌的定殖和增殖。在南美白对虾。
