罗伊氏乳酸杆菌SL001对AD模型小鼠和C57BL/6小鼠肠道微生物的影响

Influence of Lactobacillus reuteri SL001 on intestinal microbiota in AD model mice and C57BL/6 mice

摘要:

本研究旨在调查阿尔茨海默症模型小鼠 (APP/PS1双转基因小鼠，AD小鼠) 与野生型小鼠 (C57BL/6小鼠) 肠道菌群的多样性差异，以及添食罗伊氏乳酸杆菌SL001 (Lactobacillus reuteri SL001) 对两种小鼠肠道微生物菌群结构组成的影响，探讨其是否可对AD小鼠起到积极的改善作用。首先将雄性AD模型和野生型小鼠分别分成添食组和未添食组 (每组5只)，添食组小鼠每天用0.2 mL浓度为5×1011 CFU/mL的L. reuteri SL001菌悬液灌胃，未添食组则每天接受等量的无菌PBS，通过口服管饲法进行给物，周期为45 d。实验结束后，采集小鼠的粪便样本，通过高通量测序技术测定了16S rRNA V3–V4区序列，并对群落结构和多样性进行比较分析。结果显示4个处理组小鼠的肠道微生物OTUs总共包括19个门、40个纲、64个目、104个科和175个属。AD模型小鼠的α多样性大于野生型小鼠，但是两者之间的差异不显著。添食L. reuteri SL001后，两种小鼠的α多样性均增加，AD模型小鼠的增幅较小。在门水平上，4组小鼠的优势菌门均为拟杆菌门 (Bacteroidetes)、厚壁菌门 (Firmicutes) 和变形菌门 (Proteobacteria)，AD模型小鼠中的拟杆菌门丰度低于野生型，厚壁菌门的丰度高于野生型，添食L. reuteri SL001会降低小鼠的厚壁菌门/拟杆菌门的比例 (F/B)。在目水平上，AD模型小鼠中的拟杆菌目 (Bacteroidales)、乳酸杆菌目 (Lactobacillales)、芽胞杆菌目 (Bacillales) 和双歧杆菌目 (Bifidobacteriales) 的相对丰度低于野生型小鼠。在属水平上，丰度较高的属有Allobaculum、毛螺旋菌属 (Lachnospiraceae_NK4A136_group)、拟杆菌属 (Bacteroides)、乳酸菌属 (Lactobacillus)。AD模型小鼠的拟杆菌属、乳酸菌属、双歧杆菌属 (Bifidobacterium)、和拟普雷沃菌属 (Alloprevotella) 的相对丰度低于野生型小鼠，添食L. reuteri SL001会增加这些属以及拟杆菌目、乳酸杆菌目、芽胞杆菌目和双歧杆菌目在AD模型小鼠中的丰度。AD模型小鼠的丁酸弧菌属 (Butyrivibrio) 相对丰度也低于野生型，添食L. reuteri SL001后，其相对丰度在两种小鼠中均出现降低。野生型小鼠的优势菌种为杆菌纲 (Bacilli) 中的乳酸杆菌科 (Lactobacillaceae)，AD模型小鼠中未发现优势菌群，这一结果与不同比例物种分类树结果相符。文中结果揭示了AD模型小鼠与野生型小鼠肠道微生物多样性差异，也表明添食L. reuteri SL001可以改善小鼠肠道细菌群落结构组成，对AD模型小鼠能够起到一定程度的积极调整作用。

Abstract:

We studied the influence of Lactobacillus reuteri SL001 (L. reuteri SL001) on the gut microbial community in Alzheimer’s disease model mice (APP/PS1 double transgenic mice, ADmice) and wild type (C57BL/6) mice. The AD model mice and wild type mice were divided into test and control groups (4 in total), with 5 mice each and only male mice. The test group was fed with 0.2 mL suspension of L. reuteri SL001 at 5×1011 CFU/mL. The control group received the same amount of sterile PBS daily for 45 days. Fecal samples were collected to compare and analyze the community structure and diversity of microbiota through high-throughput sequencing of the V3–V4 region of 16S rRNA gene. By sequence alignment and classification, the intestinal microbial OTUs of the 4 groups including 19 phyla, 40 classes, 64 orders, 104 families and 175 genera. The α diversity of AD model mice was greater than that of wild type mice, but the difference between the two was small. After adding L. reuteri SL001, the α diversity of both mice increased, and the increase in AD model mice was smaller. At the phyla level, the dominant phyla of the four groups of mice were Bacteroidetes, Firmicutes and Proteobacteria. The abundance of Bacteroides phylum in AD model mice was lower than that of wild type, and the abundance of chlamydomonas was higher than that of wild type. Feeding L. reuteri SL001 reduced the proportion of Firmicutes/Bacteroidetes (F/B) in mice. At the order level, the relative abundance of Bacteroidales, Lactobacillales, Bacillales and Bifidobacteriales in AD model mice was lower than that of wild type mice. At the genus level, the abundant genera were Allobaculum, Lachnospiraceae_NK4A136_group, Bacteroides and Lactobacillus. The relative abundance of Bacteroides, Lactobacillus, Bifidobacterium and Alloprevotella in AD model mice was lower than that in wild type mice. Adding L. reuteri SL001 increased the abundance of these genera and Bacteroides, Lactobacillus, Bacillus and Bifidobacteria in AD model mice. The relative abundance of Butyrivibrio in AD model mice was also lower than that in wild type, but after the feeding of L. reuteri SL001, the relative abundance was reduced in both mice. The dominant strains of wild-type mice were Lactobacillus, and no dominant flora was found in AD model mice. The results in this article provide valuable data for revealing the difference in intestinal microbial diversity between AD model mice and C57BL/6 mice, and feeding L. reuteri SL001 play positive roles in adjusting the intestinal bacterial community structure of AD model mice.