采集3 家无菌制药企业的核心生产环境(A/B/C 级洁净车间) 的沉降菌、浮游菌和人员及设备表面菌，通过生化鉴定、 16S rRNA 序列分析和基因间隔序列(ITS) 鉴定，发现革兰阳性菌在微生物中占比最高，其中葡萄球菌属、微球菌、库克菌和芽孢杆菌等凝聚酶阴性葡萄球菌(CoNS) 属于优势菌种。CoNS 作为主要的污染微生物，通过选取6 个看家基因对其进行多位点序列分型(multi-locus sequence typing，MLST)，形成菌株聚类关系图。药敏试验表明，20 株CoNS 对庆大霉素、万古霉素、替加环素等7 种抗菌药的敏感率为100％，对头孢西丁、青霉素、红霉素等11 种抗菌药的敏感率为25％～ 95％，有2 株沃氏葡萄球菌分别对7 种抗菌药表现出耐受性。企业应持续进行环境监测，建立微生物菌种数据库，为微生物的溯源和污染控制提供科学依据。
Abstract
Settling bacteria, planktonic bacteria and bacteria on the surface of personnel and equipment in the core production environment of three sterile pharmaceutical companies(A/B/C grade clean room) were collected for biochemical identification, 16S rRNA sequence analysis, and internal transcribed spacer(ITS) identification. The results showed that gram-positive bacteria accounted for the highest proportion of microorganisms, and coagulasenegative Staphylococci(CoNS) such as Staphylococcus, Micrococcus, Kocuria and Bacillus were dominant species. As the main contaminating microorganisms, CoNS were used to select six house-keeping genes for multi-locus sequence typing(MLST) to form a strain clustering relationship map. The drug sensitivity test showed that the sensitivity of twenty CoNS strains to seven antibiotics(such as gentamicin, vancomycin and tigecycline) was 100％. The sensitivity to eleven antibiotics(such as cefoxitin, penicillin and erythromycin) was 25％ - 95％. And there were two strains of Staphylococcus warneri that were resistant to seven antibiotics. Enterprises should continue to conduct environmental monitoring and establish a microbial strain database to provide scientific basis for microbial traceability and pollution control.
关键词
洁净车间 /
微生物污染 /
菌种鉴定 /
多位点序列分型 /
耐药谱
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Key words
clean room /
microorganism contamination /
species identification /
multi-locus sequence typing /
antibiotic resistance profile
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