基于高通量测序技术分析慢性鼻窦炎患者的肠道菌群结构特点

Analysis of intestinal flora in patients with chronic rhinosinusitis based on highthroughput sequencing

目的采用高通量测序技术,检测和比较慢性鼻窦炎伴鼻息肉(CRSwNP)患者和正常人群的肠道菌群多样性、相对丰度及分布的改变,发现与慢性鼻窦炎伴鼻息肉显著相关的菌属结构,旨在探究慢性鼻窦炎发生发展过程中可能具有重要作用的菌群变化及预测相关代谢通路。方法选取在广东省人民医院耳鼻喉头颈外科住院的慢性鼻窦炎伴鼻息肉(CRSwNP)患者10例作为观察组,另随机选取同期健康体检者10例作对照,对粪便样本相关细菌采用粪便基因组DNA提取试剂盒抽提粪便样本中DNA,并进行DNA片段长度测定和定量,用一系列PCR引物扩增原核生物16S rDNA基因的V3、V4高度可变区,再进行文库构建、Illumina MiSeq测序、序列比对和物种鉴定分析。分析肠道菌群的相对丰度、多样性及分布特点,进而预测相关代谢通路。结果慢性鼻窦炎伴鼻息肉患者的肠道菌群整体结构与健康对照人群具有显著的差异(P<0.05),主要表现为糖多孢菌属(Saccharopolyspora)丰度增加,而瘤胃菌科(Ruminococcaceae)、粪球菌属(Coprococcus)、柯林斯氏菌属(Collinsella)、杆菌属(Dialister)含量降低。其中在代谢通路预测中,9条代谢通路存在显著差异(P<0.05)。结论慢性鼻窦炎组与健康对照组对比,肠道菌群结构特点发生了明显改变,预测其与多个代谢通路水平变化相关。在慢性鼻窦炎的发生发展中,肠道菌群微生态的改变可能起到了重要作用。

Objective To investigate the changes in diversity,relative abundance and distribution of intestinal flora in patients with chronic rhinosinusitis and nasal polyps(CRSwNP)using high-throughput sequencing technology identify the intestinal flora significantly related to pathogenesis and progression of CRSwNP.Methods Ten patients with CRSwNP hospitalized in the Department of Otolaryngology-Head and Neck Surgery of Guangdong Provincial People's Hospital were selected as the case group with 10 healthy volunteers recruited in the same period as the control group.Fecal genomic DNA extraction kit was used to extract the DNA in the fecal samples,and the DNA fragment length was measured and quantified.The V3 and V4 highly variable regions of the 16S rDNA gene of prokaryotes were amplified followed by library construction,Illumina MiSeq sequencing,sequence alignment and species identification analysis.The relative abundance,diversity and distribution characteristics of the intestinal flora were analyzed,and the relevant metabolic pathways were predicted.Results Compared with the control group,the patients with CRSwNP had significant changes in the overall structure of the intestinal flora,highlighted by increased abundance of Saccharopolyspora and decreased contents of Ruminococcae,Coprococcus,Collinsella and Dialister.Among the metabolic pathways predicted to be associated with CRSwNP,9 showed significant changes in patients with CRSwNP as compared with the control group(P<0.05).Conclusion Patients with CRSwNP have significant changes in the structural characteristics of intestinal flora related with multiple metabolic pathways,and these changes may play an important role in the development of chronic rhinosinusitis.