基于16S rRNA高通量测序分析9种中药饮片污染微生物群落特征  

Analysis of Microbial Community Characteristics in 9 Kinds of Chinese Herbal Pieces Based on 16S rRNA High Throughput Sequencing

作　　者：陈柯 陈敏 林黎 邹惠亮 CHEN Ke;CHEN Min;LIN Li;ZOU Huiliang(Huzhou Institute for Food and Drug Control,Huzhou Zhejiang 313000,China)

机构地区：[1]湖州市食品药品检验研究院,浙江湖州313000

出　　处：《中医药导报》2022年第9期53-56,61,共5页Guiding Journal of Traditional Chinese Medicine and Pharmacy

基　　金：浙江省市场监督管理局科研计划项目(20210147)。

摘　　要：目的:基于16S核糖体RNA(16S rRNA)高通量测序分析浙江省湖州市南太湖地区9种中药饮片的污染微生物群落特征。方法:根据《中华人民共和国药典》(2020年版)对柴胡、白术、山药、玄参、麦冬、焦栀子、砂仁、铁皮枫斗、金银花共9种中药饮片进行控制菌检查和分类鉴定,同时采用16S rRNA高通量测序法测定基因序列并将优化后的序列与数据库比对,进行多样性分析,分析微生物群落特征。结果:(1)90批中药饮片中有1批山药检测出沙门菌和大肠埃希菌,有41批(45.56%)检出耐胆盐革兰氏阴性菌。经菌落鉴定共获得4个科10个属共15种污染微生物,其中包括阴沟肠杆菌、铜绿假单胞菌、阪崎克罗诺杆菌、沙门氏菌、大肠埃希菌、肺炎克雷伯氏菌、鲍曼不动杆菌、奇异变形杆菌等致病菌或条件致病菌。(2)高通量测序共获得32个属、186种污染微生物,山药、白术、柴胡鉴定出的微生物种类最高,分别为112、95、88,焦栀子、铁皮枫斗和麦冬获得微生物种类最少,分别为20、26、34。经多样性分析,柴胡、白术和山药具有最高的Chaol指数,柴胡、山药和砂仁具有最高的Shannon指数,柴胡、山药和砂仁具有最高的Simpson指数。优势菌属为肠杆菌属、假单胞菌属、泛菌属、变形杆菌属、不动杆菌属、克罗诺杆菌属、克雷伯氏菌属、芽孢杆菌属、葡萄球菌属、链球菌属、肠球菌属。结论:上述中药饮片中存在微生物污染,有一定的致病风险。与传统平板培养相比较,高通量测序可以更加全面、快速获得中药饮片中的微生物信息和群落特征。Objective:To analyze the microbial community characteristics in 9 kinds of Chinese herbal pieces in South Tai lake area based on 16S rRNA high throughput sequencing.Methods:According to the Chinese Pharmacopoeia(2020 Edition),the control bacteria were examined,classified and identified in 9 kinds of Chinese herbal pieces,including Chaihu(Bupleurum),Baizhu(Atractylodes macrocephala),Shanyao(Yam),Xuanshen(Scrophularia),Maidong(Ophiopogon japonicas),Jaiozhizi(stir-baked gardenia),Sharen(Amomum villosum),Tiepi Fengdou(Dendrobium officinale)and Jinyinhua(Honeysuckle).The gene sequence was determined by 16S rRNA high throughput sequencing,and the optimized sequence was compared with the database to analyze the diversity and analyze the characteristics of microbial community.Results:(1)Salmonella and Escherichia coli were detected in one batch of Shanyao(Yam)among 90 batches of Chinese herbal pieces,and bile salt resistant gram-negative bacteria were detected in 41 batches(45.56%).A total of 15 kinds of polluting microorganisms were obtained by colony identification in 10 genera from 4 families,including Enterobacter cloacae,Pseudomonas aeruginosa,cronobacteria sakazakii,Salmonella,Escherichia coli,Klebsiella pneumoniae,Acinetobacter baumannii,Proteus mirabilis and other pathogenic or conditional pathogens.(2)A total of 186 contaminated microorganisms belonging to 32 genera were obtained by high throughput sequencing.The microbial species identified in Shanyao(Yam),Baizhu(Atractylodes macrocephala)and Chaihu(bupleurum)had the most microbial species,which were respectively 112,95 and 88.Jaiozhizi(stir-baked gardenia),Tiepi Fengdou(Dendrobium officinale)and Maidong(Ophiopogon japonicas)obtained the least microbial species,which were respectively 20,26 and 34.According to the diversity analysis,Chaihu(bupleurum),Baizhu(Atractylodes macrocephala)and Shanyao(Yam)showed the highest Chaol index,and Chaihu(bupleurum),Shanyao(Yam)and Sharen(Amomum villosum)showed the highest Shannon index.Chaihu(bupleurum),Shanyao(yam)and Sharen

关 键 词：高通量测序 中药饮片 微生物污染 群落特征 耐胆盐微生物 16S rRNA 

分 类 号：R282.4[医药卫生—中药学]