大环内酯类抗生素耐药肺炎支原体标准菌株的建立与评价  

Establishment and evaluation of standard strain of macrolide-resistant Mycoplasma pneumonia

作　　者：刘立雍 张倚玮 向才鑫 何利华[1] 孟凡亮[1] 刘洁[1,2] 张建中[1] 赵飞[1] Liu Liyong;Zhang Yiwei;Xiang Caixin;He Lihua;Meng Fanliang;Liu Jie;Zhang Jianzhong;Zhao Fei(State Key Laboratory for Infectious Disease Prevention and Control,National Institute for Communicable Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China;School of Public Health,China Medical University,Shenyang 110000,Liaoning,China)

机构地区：[1]中国疾病预防控制中心传染病预防控制所,传染病预防控制国家重点实验室,北京102206 [2]中国医科大学公共卫生学院,辽宁沈阳110000

出　　处：《疾病监测》2023年第12期1466-1472,共7页Disease Surveillance

基　　金：中国疾病预防控制中心传染病所自主课题(No.TS202313);鼠疫、霍乱、布病等传统传染病监测防治综合能力提升(No.102393230020020000002);细菌及真菌性传染病预防与干预(No.102393220020020000031)。

摘　　要：目的依据病原微生物菌(毒)种国家标准菌株评价技术标准(WS/T 812—2022)建立大环内酯类抗生素耐药肺炎支原体(Mp)标准菌株,加速我国Mp相关研究进展,推进国家病原微生物资源保藏体系建设。方法通过对菌株进行16S rDNA测序及特异性基因PCR鉴定,确定菌株为Mp菌株。通过对标准菌株菌落形态观测、基因分型、活性测定、最低抑菌浓度(MIC)值测定、耐药基因检测等明确其生物学特征,并进行菌株稳定性和活性评价。结果标准菌株经16S rDNA测序及特异性基因PCR鉴定为Mp菌种,其连续传代后菌落形态及各种分子生物学特征稳定。标准菌株对红霉素、克拉霉素和阿奇霉素体外MIC值范围分别为≥256μg/mL、128μg/mL~≥256μg/mL和8μg/mL~32μg/mL,在基因水平上表现为23S rDNA基因A2063G突变。标准菌株在−80℃条件下可稳定保存12个月,解冻后需尽快进行增菌培养,避免4℃或者室温保存,标准菌株使用时避免超过两次冻融。结论本研究建立并评价了大环内酯类抗生素耐药Mp标准菌株(CHPC 1.13907),为Mp疾病防控、科研以及药物和疫苗的研发等工作提供标准菌株。Objective To establish standard strain of macrolide-resistant Mycoplasma pneumoniae(Mp)according to the Technical Requirements for Evaluation of National Standard Strains of Pathogenic Microorganisms(WS/T 812—2022),accelerate the progress of Mp research,and promote the construction of a national pathogenic microbial resource conservation system.Methods The Mp standard strain was identified by 16S rDNA sequencing and specific gene PCR detection.The biological characteristics of standard strains were clarified through observation of colony morphology,genotyping,activity determination,MIC value determination and drug resistance gene detection,as well as strain stability and activity evaluation.Results The standard strain was identified by 16S rDNA sequencing and specific gene PCR as Mp strain,and its colony morphology and various molecular biological characteristics were stable after successive generations.Minimum inhibitory concentration values of the standard strain in vitro ranged from≥256μg/mL,128μg/mL to≥256μg/mL and 8μg/mL to 32μg/mL for erythromycin,clarithromycin and azithromycin,respectively.The standard strain exhibited the 23S rDNA gene A2063G mutation at the genetic level.The standard strain can be stored stably for 12 months at−80℃.After thawing,the standard strain should be incubated as soon as possible,avoiding storage at 4℃or room temperature and avoiding more than two freeze-thaws during use.Conclusion This study established and evaluated standard strain of macrolide-resistant Mp(CHPC 1.13907),providing an important material basis for Mp disease prevention and control,scientific research,drug development and vaccine development.

关 键 词：肺炎支原体 标准菌株 大环内酯类抗生素 

分 类 号：R211[医药卫生—中医学] R378[生物学—微生物学] Q939.99