基于高通量测序技术分析食物蛋白诱导性直肠结肠炎患儿肠道菌群特征

Features of intestinal flora in children with food protein-induced proctocolitis based on high-throughput sequencing

摘要:

目的 通过高通量测序分析食物蛋白诱导性直肠结肠炎（food protein-induced proctocolitis，FPIP）患儿肠道菌群特征。 方法 选取2018年10月至2021年2月遵义医科大学第三附属医院门诊就诊的小于6月龄纯母乳喂养发生FPIP患儿31例纳入FPIP组，正常健康婴儿31例纳入健康对照组；采集两组粪便样本提取DNA并通过PCR扩增，采用高通量测序分析粪便样品中的16S rDNA V3~V4片段并进行相关生物信息学分析。 结果 肠道菌群多样性分析示FPIP组菌群多样性Shannon指数低于健康对照组，但差异无统计学意义（P>0.05）；菌群丰富度Chao指数高于健康对照组（P<0.01）。在门水平上，两组菌群组成均主要由厚壁菌门、放线菌门、变形菌门、拟杆菌门4个菌门构成；与健康对照组相比，FPIP组中放线菌门构成比显著降低（P<0.001），变形菌门构成比明显增加（P<0.05）。在属水平上，FPIP组主要由大肠埃希氏菌属、梭状芽孢杆菌属、肠球菌属、克雷伯氏菌属和双歧杆菌属组成；健康对照组主要由双歧杆菌属、链球菌属组成；与健康对照组相比，FPIP组中双歧杆菌属和瘤胃球菌属构成比明显降低（P<0.05），梭状芽孢杆菌属和志贺氏菌属构成比明显增加（P<0.05）。 结论 FPIP组与健康对照组相比，肠道菌群多样性下降，丰富度升高；在菌群组成结构上某些细菌菌属存在差异。提示在FPIP的发生发展过程中，肠道微生物群在属水平上的构成改变可能起到了重要作用。

Abstract:

Objective To study the features of intestinal flora in children with food protein-induced proctocolitis (FPIP) by high-throughput sequencing. Methods A total of 31 children, aged <6 months, who experienced FPIP after exclusive breastfeeding and attended the outpatient service of the Third Affiliated Hospital of Zunyi Medical University from October 2018 to February 2021 were enrolled as the FPIP group. Thirty-one healthy infants were enrolled as the control group. Fecal samples were collected to extract DNA for PCR amplification. High-throughput sequencing was used to perform a bioinformatics analysis of 16S rDNA V3-V4 fragments in fecal samples. Results The diversity analysis of intestinal flora showed that compared with the control group, the FPIP group had a lower Shannon index for diversity (P>0.05) and a significantly higher Chao index for abundance (P<0.01). At the phylum level, the intestinal flora in both groups were composed of Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes. Compared with the control group, the FPIP group had a significant reduction in the composition ratio of Actinobacteria (P<0.001) and a significant increase in the composition ratio of Proteobacteria (P<0.05). At the genus level, the intestinal flora in the FPIP group were mainly composed of Escherichia, Clostridium, Enterococcus, Klebsiella, and Bifidobacterium, and the intestinal flora in the control group were mainly composed of Bifidobacterium and Streptococcus. Compared with the control group, the FPIP group had a significant reduction in the composition ratio of Bifidobacterium and Ruminococcus (P<0.05) and significant increases in the composition ratios of Clostridium and Shigella (P<0.05). Conclusions Compared with the control group, the FPIP group has a reduction in the diversity of intestinal flora and an increase in their abundance, and there are certain differences in several bacterial genera. These results suggest that changes in the composition of intestinal flora at genus level may play an important role in the development and progression of FPIP. Citation:Chinese Journal of Contemporary Pediatrics, 2022, 24(5): 536-542