宏基因组测序技术在发热患者咽部微生物菌群研究中的应用

Application of metagenomic sequencing technology in the study of pharyngeal microflora in fever patients

目的 通过宏基因组高通量测序分析,研究不同发热患者咽部微生物菌群多样性。方法 采用随机法选取2020年1—4月石家庄市五所医院发热门诊就诊21例发热患者作为实验组,选取同期10例健康人作为对照组,采集咽拭子样本并提取核酸,应用Illumina测序平台进行宏基因组测序并进行微生物菌群分析。结果 实验组共获得1 716 802 816条优质序列,鉴定出4 079个物种,归属于97个门,951个属。实验组和对照组咽部微生物菌群分析显示,在门水平上,梭杆菌门和厚壁菌门在对照组中升高,拟杆菌门在实验组中升高;在纲水平上,梭杆菌纲和梭菌纲在对照组中升高,拟杆菌纲在实验组中升高;在目水平上,拟杆菌目在实验组中升高;在科水平上,瘤胃菌科在对照组中升高,拟杆菌科、理研菌科、阿克曼氏菌科、红细菌科和火色杆菌科在实验组中升高;在属水平上,拟普雷沃菌属、拟杆菌属、理研菌属、副拟杆菌属等在实验组中升高,瘤胃球菌属在对照组中升高;在种水平上,紫单胞菌科细菌(Porphyromonadaceae bacterium KA00676)、两个拟杆菌属(Bcteroidales bacterium Barb7, Bcteroidales SP. CAG:702)、五个普雷沃氏菌(CAG:617、CAG:755、CAG:732、CAG:924、CAG:1031)、卟啉单胞菌KLE1280、甲烷囊菌属的PtaU1.Bin060等在实验组中升高。实验组差异物种有消化链球菌、拟杆菌、科马加泰杆菌、明串珠菌、支原体、假棒状杆菌等。耐药基因主要为β-内酰胺酶类。结论 发热患者咽部有复杂的微生物菌群结构,本研究表明咽部菌群结构会对机体发热有显著影响。

Objective This study aimed to investigate the diversity of pharyngeal microflora in patients with different fever by metagenomic high-throughput sequencing.Methods A total of 21 patients with fever were randomly selected as the experimental group, and 10 healthy people in the same period were selected as the control group. Pharyngeal swab samples were collected and nucleic acids were extracted. Metagenomic sequencing and microbiota analysis were carried out by Illumina sequencing platform.ResultsA total of 1 716 802 816 high-quality sequences were obtained in the experimental group, and4 079 species were identified, belonging to 97 phylums and 951 genera. The analysis of pharyngeal microbiota in the experimental group and the control group showed that at the phylum level, Clostridium and Firmicutes increased in the control group, and Bacteroidetes increased in the experimental group;at the class level, Fusobacteriia and Clostridia increased in the control group and Bacteroidea increased in the experimental group;at the order level, Bacteroides increased in the experimental group;at the family level, Ruminococcaceae increased in the control group, Bacillariophyceae, Rikenomycetes,Ackermanniaceae, Erythrobacteriaceae, and Pyrobacteriaceae increased in the experimental group;at the genus level,Prevotella spp., Mycobacterium spp., Rikenia spp., Paramecium spp. increased in the experimental group, and Rumenococcus spp. increased in the control group;at the species level, Porphyromonadaceae bacterium ka00676, two Bacteroides(Bcteroides Bacterium barb7, Bcteroides sp. CAG: 702), five Prevotella(CAG: 617, CAG: 755, CAG: 732, CAG: 924, CAG: 1031),Porphyromonas kle1280 and ptau1.bin060 of Methanogens increased in the experimental group. The different species in the experimental group were Peptostreptococcus, Bacteroides, Komagataeibacter, Leuconostoc, Mycoplasma, Pseudoclavibacter and so on. The drug resistance genes were mainly β-lactamases.Conclusion The pharynx of patients with fever has complex microbial flora structure, this study shows that the pharynx flora structure can have a significant effect on fever.