目的 通过16SrRNA基因测序方法探索1型糖尿病(T1DM)儿童肠道菌群结构变化的特征,并比较T1DM儿童与健康儿童间肠道菌群的差异,为临床应用益生菌进行TIDM早期干预提供理论基础和实验依据。方法 选取2018年10月-2019年10月在昆明市儿童医院内分泌遗传代谢科住院并初诊为T1DM的5~14岁儿童18例,同时选取19例性别、年龄相近的健康儿童为研究对象,收集两组儿童粪便标本后进行16SrRNA基因测序实验,并采用QIIME 2分析流程进行生物信息学分析,比较两组间肠道菌群的差异。结果 1)利用QIIME2软件将相似度100%的序列聚类分析后共获得3 248个Feature数;2)经物种鉴定及注释,绝大部分菌群都分类到属级和种级;3)Alpha多样性分析说明本次研究测序深度充分,并且T1DM儿童肠道菌群的丰富度及多样性较健康儿童降低 (P<0.05);4)Beta多样性分析中,PCoA图说明T1DM儿童和健康儿童间肠道菌群结构存在明显差异;5)肠道菌群组成差异分析中,在门水平上,T1DM儿童放线菌门、拟杆菌门和蓝细菌门丰度显著增高,而变形菌门、杆菌门的丰度降低(P<0.05);在属水平上,T1DM儿童粪杆菌属、双歧杆菌属和拟杆菌属的丰度较健康儿童增高,而埃希氏杆菌属-志贺氏杆菌、肠球菌属、Blautia菌属丰度降低(P<0.05)。结论 T1DM儿童存在肠道菌群生态失衡,并且菌群物种的丰富度及多样性降低。此外,T1DM儿童与健康儿童肠道菌群结构分布存在明显差异。
Abstract
Objective To explore the characteristics of intestinal flora in children with type 1 diabetes mellitus (T1DM) by 16SrRNA gene sequencing method, and to compare the differences on gut microbiota between T1DM patients and healthy controls, so as to provide theoretical basis and evidence for clinical application of probiotics in early intervention for T1DM children. Methods A total of 18 children aged 5 to 14 years who were newly diagnosed with T1DM in the Department of Endocrinology and Metabolism, Kunming Children′s Hospital were enrolled in this study from October 2018 to October 2019.At the same time, 19 healthy children with similar gender and age were selected as controls.After collecting fresh stool samples from two groups, 16SrRNA gene sequencing experiments were performed, and QIIME 2 analysis processes were used for bioinformatics analysis.Then the marked differences of gut microbiota among two groups were compared. Results 1) Totally 3 248 Features with 100% similarity were obtained by using QIIME2 software.2) Most of intestinal bacteria were classified into genus and species after identification and annotation.3) Alpha diversity analysis showed that the sequencing depth was sufficient in this study.Moreover, the richness and diversity of gut microbiota in T1DM children were lower than those in healthy controls (P<0.05).4) In the analysis of beta diversity, the PCoA image indicated that the gut microbiota structure was different between T1DM group and healthy controls.5) At the level of phylum, the abundance of Actinobacteria, Bacteroidetes and Cyanobacteria was increased while the abundance of Proteobacteria and Patescibacteria was decreased in T1DM group(P<0.05).At the genus level, the abundance of Faecalibacterium, Bifidobacterium and Bacteroides was increased in T1DM group, but the abundance of Escherichia_Shigella, Enterococcus and Blautia was reduced (P<0.05). Conclusions The ecology of gut microbiota is imbalanced in T1DM children.Compared with healthy children, the richness and diversity are notably lower in T1DM children as well.Moreover, there are obvious differences on the structure of gut microbiota between T1DM children and healthy children.
关键词
肠道菌群 /
1型糖尿病 /
儿童 /
16SrRNA测序
Key words
gut microbiota /
type 1 diabetes mellitus /
children /
16SrRNA sequencing
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