DataSheet_1_Low fish meal diet supplemented with probiotics ameliorates intestinal barrier and immunological function of Macrobrachium rosenbergii via the targeted modulation of gut microbes and derived secondary metabolites.zip

The unsuitable substitution ratio of fish meal by plant protein will reshape the intestinal microbial composition and intestine immunity. However, previous studies were mostly limited to investigating how different feed or probiotics characterized the microbial composition but ignored the biological interactions between bacteria and host physiology through secondary metabolites. Therefore, this study integrates the apparent indicators monitoring, 16S rDNA sequencing, and metabonomics to systematically investigate the effects of cottonseed protein concentrate (CPC) substitution of fish meal and Bacillus coagulans intervention on gut microbes, secondary metabolites, and intestinal immunity of Macrobrachium rosenbergii. Prawns were fed with three diets for 70 days: HF diets contained 25% fish meal, CPC in LF diets were replaced with 10% fish meal, and LF diets supplemented with 2 × 108 CFU/g diet B. coagulans were designated as BC diets. Results showed that CPC substitution induced a significant decrease in digestive enzyme activities (trypsin and lipase) and gut barrier protein PT-1 expression and a significant increase in γ-GT enzyme activity and inflammatory-related factors (Relish and Toll) expression. B. coagulans treatment mitigated the negative changes of the above indicators. Meanwhile, it significantly improved the expression levels of the barrier factor PT-1, the reparative cytokine IL-22, and Cu/Zn-SOD. CPC substitution resulted in a remarkable downregulated abundance of Firmicutes phyla, Flavobacterium spp., and Bacillus spp. B. coagulans treatment induced the callback of Firmicutes abundance and improved the relative abundance of Sphingomonas, Bacillus, and Ralstonia. Functional prediction indicated that CPC substitution resulted in elevated potential pathogenicity of microbial flora, and B. coagulans reduces the pathogenesis risk. Pearson’s correlation analysis established a significant positive correlation between differential genera (Sphingomonas, Bacillus, and Ralstonia) and secondary metabolites (including sphingosine, dehydrophytosphingosine, amino acid metabolites, etc.). Meanwhile, the latter were significantly associated with intestinal immunoregulation-related genes (Cu/Zn-SOD, IL-22, PT-1, Toll, and Relish). This study indicated that B. coagulans could mediate specific gut microbes and the combined action of multiple functional secondary metabolites to affect intestinal barrier function, digestion, and inflammation. Our study revealed the decisive role of gut microbes and derived secondary metabolites in the model of dietary composition-induced intestinal injury and probiotic treatment from a new perspective.

植物蛋白替代鱼粉的比例不当，会重塑肠道微生物组成与肠道免疫功能。然而，既往研究多聚焦于探究不同饲料或益生菌对微生物组成的调控作用，却忽视了细菌通过次级代谢产物与宿主生理之间的相互作用。因此，本研究整合表观指标监测、16S rDNA测序（16S rDNA sequencing）与代谢组学技术，系统探究棉籽蛋白浓缩物（cottonseed protein concentrate, CPC）替代鱼粉以及凝结芽孢杆菌（Bacillus coagulans）干预对罗氏沼虾（Macrobrachium rosenbergii）肠道微生物、次级代谢产物及肠道免疫的影响。
本研究设置三组饲料饲喂罗氏沼虾70天：HF组饲料含25%鱼粉；LF组饲料中10%鱼粉被CPC替代；LF组中添加2×10^8 CFU/g饲料的凝结芽孢杆菌，记为BC组。
结果显示，CPC替代会显著降低消化酶活性（胰蛋白酶、脂肪酶）与肠道屏障蛋白PT-1的表达水平，并显著升高γ-谷氨酰转移酶（γ-GT）活性以及炎症相关因子（Relish、Toll）的表达量。凝结芽孢杆菌干预可缓解上述指标的负面变化，同时显著提升屏障因子PT-1、修复性细胞因子IL-22以及铜锌超氧化物歧化酶（Cu/Zn-SOD）的表达水平。CPC替代会显著下调厚壁菌门（Firmicutes）、黄杆菌属以及芽孢杆菌属的相对丰度；而凝结芽孢杆菌干预可逆转厚壁菌门丰度的下降，并提升鞘氨醇单胞菌属、芽孢杆菌属以及罗尔斯通氏菌属的相对丰度。
功能预测结果表明，CPC替代会提升微生物菌群的潜在致病性，而凝结芽孢杆菌可降低该致病风险。Pearson相关性分析显示，差异菌群（鞘氨醇单胞菌属、芽孢杆菌属与罗尔斯通氏菌属）与次级代谢产物（包括神经鞘氨醇、脱氢植物鞘氨醇、氨基酸代谢物等）之间存在显著正相关；同时，这些代谢产物与肠道免疫调控相关基因（Cu/Zn-SOD、IL-22、PT-1、Toll及Relish）显著关联。
本研究表明，凝结芽孢杆菌可通过调控特定肠道菌群与多种功能性次级代谢产物的协同作用，影响肠道屏障功能、消化能力与炎症状态。本研究从全新视角揭示了肠道微生物及其衍生次级代谢产物在饲料组成诱导的肠道损伤模型与益生菌治疗中的关键作用。