低聚木糖调控断奶仔猪后肠黏膜双歧杆菌黏附及其利用机制

The growth and adhesion of probiotics promote intestinal heath of weaned piglets. Molecular mechanism for xylooligosaccharides (XOS) selectively increase Bifidobacterium growth in the intestinal mucosa of weaned piglets is still unclear. According to the scientific problem of microbial growth in the intestinal mucosa depended on the adhesion and carbon utilization of microbiota, the program will employ the integrated methods of genomics, proteomics and microbiology in a weaned piglet model and include three main contents: 1) determining the effect of inclusion of XOS to a diet fed to weaned piglets for 14 and 28 days on microbial population in the cecal and colonic mucosa using a high-throughput 16S rRNA gene sequencing, establishing the protein-sequence database of microbiota and analyzing the change for Bifidobacterium population; 2) isolating and screening the Bifidobacterium strains with high adhesive ability and extracting the surface and intracellular protein for the strains; 3) identifying the differently expressed surface and intracellular protein of the Bifidobacterium strains by the inclusion of XOS using a quantitative proteomics technology. The program will be achieved by establishing the signal network of the effect of XOS on adhesion of Bifidobacterium in the hindgut mucosa and the utilization model of XOS mediated an ATP binding cassette (ABC) transporter. The results could reveal the reason that inclusion of XOS increases the Bifidobacterium growth and enrich the regulation theory of growth and adhesion of Bifidobacterium in the intestinal mucosa by oligosaccharides, in order to provide the theoretical basis for nutrition regulation of intestinal health in weaned piglets.

有益菌的黏附与增殖是仔猪肠道健康的关键，低聚木糖选择性刺激肠黏膜双歧杆菌增殖，但其机制尚不明晰。本项目围绕肠道微生物黏附和碳源利用的能力决定肠黏膜微生物增殖这一线索，以断奶仔猪为对象，采用基因组学、蛋白组学、微生物学的方法，依次开展三方面研究：①使用16S rRNA基因高通量测序技术研究低聚木糖对断奶后14和28天仔猪盲肠和结肠微生物菌群结构多样性，建立肠内微生物蛋白质序列库，分析肠黏膜双歧杆菌种群变化规律；②分离和筛选低聚木糖作用下高黏附性双歧杆菌菌株，提取高黏附性双歧杆菌的表面蛋白质和胞内蛋白质；③采用定量蛋白质组学技术，鉴定低聚木糖作用下双歧杆菌差异表达的菌体蛋白质，构建低聚木糖调控双歧杆菌黏附后肠黏膜以及ABC转运子介导下双歧杆菌利用低聚木糖的信号网络。揭示低聚木糖促进双歧杆菌增殖的机制，丰富双歧杆菌黏附肠黏膜与增殖的寡糖调控理论，为断奶仔猪肠道健康营养调控提供理论依据。

断奶仔猪肠道正常菌群数量的维持和肠道屏障功能的健全是肠道发育和健康的重要保障。本项目针对低聚木糖促进断奶仔猪肠道有益菌增殖的作用机制尚不清楚，以断奶仔猪为对象，采用基因组学、蛋白组学、分子生物学的方法，研究发现低聚木糖在断奶仔猪生长和肠道健康中剂量效应，分析低聚木糖作用下断奶仔猪肠道不同生态位微生物菌群结构的变化规律，筛选低聚木糖作用下有益菌群，分析低聚木糖调控优势菌群及黏膜细胞蛋白质信号通路，初步揭示了低聚木糖促进断奶仔猪肠内有益菌增殖和增强黏膜免疫的分子机制。本项目研究表明，不同低聚木糖添加剂量对断奶仔猪生长、生产性能、血液指标和肠道形态结构存在明显的剂量效应，研究发现断奶仔猪生长和肠道健康最适的低聚木糖剂量（500mg/kg）；分析低聚木糖作用下断奶仔猪回肠、盲肠微生物菌群结构的变化规律，研究筛选出低聚木糖作用下断奶仔猪肠道高黏附的Lactobacillus有益菌，显著降低Clostridium_sensu_stricto_1和Terrisporobacter菌含量；低聚木糖促进断奶仔猪肠道乳酸杆菌增殖、改变黏膜蛋白糖基化的修饰和抗原递呈途径，添加低聚木糖调控粘膜内质网内系列糖基化修饰酶（POFUT1、B4GALT5、FUT11等）的表达，影响蛋白折叠（DNAJC5、ST13、DNAJA3）以及糖蛋白运输（KIF2C、AP3M1、TRAPPC10等）相关蛋白表达；低聚木糖可通过改变肠黏膜糖基化合成、转运以及代谢系列过程，通过为Lactobacillus等有益微生物提供黏附位点和底物，改善微生物的菌群结构，增强抗原递呈和处理进程，增强肠道黏膜免疫功能。项目初步揭示了低聚木糖促进断奶仔猪肠内有益菌增殖和增强黏膜免疫的分子机制，项目结果可丰富肠黏膜屏障的寡糖调控理论，为断奶仔猪肠道健康营养调控提供理论依据。