结核分枝杆菌体外药敏试验标准株的建立与评价

Establishment of Chinese reference strains for in vitro drug sensitivity testing of Mycobacterium tuberculosis

目的按照《病原微生物菌(毒)种国家标准株评价技术标准》(WS/T 812—2022),对已有多年使用基础的5株中国临床来源结核分枝杆菌菌株进行系统鉴定评价,以此建立我国的结核分枝杆菌体外药敏试验标准株。方法建立标准的实验操作程序,对菌株进行培养特性、表型、生化特征、药敏试验及分子特征检测等系统评价,完成全基因组测序,建立主代及工作菌株库。结果菌株的表型、生化及分子生物学检测结果表明5株菌符合结核分枝杆菌特征,性状稳定;耐药类型为2株全敏感菌株,2株耐多药结核菌株和1株广泛耐药结核菌株;完成全基因组测序;已获得中国疾病预防控制中心病原微生物菌(毒)种保藏中心(CHPC)保藏编号。结论5个菌株符合结核分枝杆菌体外药敏试验质控菌株及抗结核药物筛选及评价测试菌株的要求,将为中国标准株的建立工作提供参考,为我国结核病耐药监测以及抗结核药物研发提供有力的支撑。

Objective To establish Chinese reference strains for in vitro antimicrobial susceptibility testing of Mycobacterium tuberculosis in China by systematically identification and evaluation of 5 Chinese clinical Mycobacterium tuberculosis strains that have been used for many years in accordance with the Technical Requirements for Evaluation of National Standard Strains of Pathogenic Microorganisms(WS/T 812—2022).Methods Standard operation procedure was established.Bacterial culture characteristics,phenotype characteristics,biochemical tests,antimicrobial susceptibility testing,and molecular feature were systematically evaluated.Whole genome sequencing was conducted,and library of main and working strains were established.Results The phenotypic,biochemical,and molecular biological results of the five strains showed that the characteristics of these bacteria were stable and consistent with that of Mycobacterium tuberculosis,which were composed of 2 pan-susceptible strains,2 multi-drug resistant tuberculosis strains,and 1 extensively drug resistant tuberculosis strains.Whole genome sequencings were completed for all strains.National preservation number and standard strain certificate were obtained from National Pathogen Resource Center.Conclusion The characteristics of the five strains meet the requirements of quality control strains for in vitro drug susceptibility testing of Mycobacterium tuberculosis and strains for anti-tuberculosis drug screening and evaluation.It will provide guideline for the establishment of Mycobacterium tuberculosis complex reference strains,and strong support for drug resistance monitoring of tuberculosis and anti-tuberculosis drug research in China.