利用培养组学技术分离培养肺部微生物群研究  

Isolation and cultivation of lung microbiota with culturomics

作　　者：刘月姣 李俭杰 孙一凡 黄自然 李军峰 安彤同[2] 卓明磊[2] 迟雨佳 李生 杨晓丹 颊博 潘志远 杨瑞馥 王子平[2] 毕玉晶 LIU Yuejiao;LI Jianjie;SUN Yifan;HUANG Ziran;LI Junfeng;AN Tongtong;ZHUO Minglei;CHI Yujia;LI Sheng;YANG Xiaodan;JIA Bo;PAN Zhiyuan;YANG Ruifu;WANG Ziping;BI Yujing(State Key Laboratory of Pathogen and Biosecurity,Institute of Microbiology and Epidemiology,Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100071,China;Department of Thoracic Oncology,Peking University Cancer Hospital,Beijing 100142,China)

机构地区：[1]军事科学院军事医学研究院微生物流行病研究所,北京100071 [2]北京大学肿瘤医院胸部肿瘤内一科,北京100142

出　　处：《微生物学报》2022年第3期1110-1118,共9页Acta Microbiologica Sinica

基　　金：北京大学肿瘤医院科学基金(2017-18)。

摘　　要：【目的】使用培养组学技术分离培养肺部微生物群,初步了解人体肺部可培养细菌的组成特点,建立呼吸道微生物菌库,为以后进行重点菌株的功能研究提供菌株条件。【方法】针对6个临床肺泡灌洗液样本,使用补充不同添加剂的血培养瓶对样本进行预增菌,在预增菌不同的时间点对血培养瓶内细菌进行分离培养和保藏,使用MALDI-TOF质谱和16SrRNA基因测序鉴定分离菌株。【结果】共获得101种已鉴定细菌,6株潜在新菌,2株真菌。其中细菌包括5个门、14个纲、24个目、35个科和45个属。预增菌前期的时间点分离菌种数较多,厌氧环境分离菌种多于需氧条件,在预增菌时添加羊血和瘤胃液起到良好的分离效果,本实验分离的肺部微生物群与人体其他部位(尤其是肠道)有很大的交叉。【结论】利用培养组学技术可以实现对肺部微生物群的分离培养,继续探索对肺部微生物群培养的优化方法具有现实意义。[Objective] To understand the composition and characteristics of the human lung microbiota, and build a respiratory tract microbial bank which provides strains for further study, we isolated lung microbiota with culturomics. [Methods] Six clinical alveolar lavage fluid samples were collected, and blood culture bottles supplemented with different additives were used to pre-enrich the samples. The strains in the blood bottles were isolated, cultured, and preserved at different time points of the pre-enrichment. Isolated strains were identified with matrix-assisted laser desorption/ionization-time of flight(MALDI-TOF) mass spectrometry or 16 S rRNA gene sequencing. [Results] A total of 101 known bacterial species, 6 potential new bacterial species, and 2 fungal species were obtained. The bacteria belonged to 45 genera, 35 families, 24 orders, 14 classes of 5 phyla. More isolates were obtained from the early stage of pre-enrichment, and there were more isolates in the anaerobic environment than at aerobic conditions. The additional supplement of sheep blood and rumen fluid during pre-enrichment were beneficial to strain isolation. The lung bacteria isolated in this experiment overlapped to a large extent with those from other parts(especially the intestine) of the human body.[Conclusion] Culturomics realizes the isolation of lung microbiota, and it is essential to continue to optimize the methods for isolating more bacteria from lungs.

关 键 词：培养组学 肺部微生物群 肺泡灌洗液 方法优化 

分 类 号：R446.5[医药卫生—诊断学] R563[医药卫生—临床医学]