荧光素酶生物发光法在快速药敏试验中的研究与应用  

Research and application of luciferase bioluminescence in rapid antimicrobial susceptibility testing systems

作　　者：向杰 崔晓莉 李璐一 周同喜 方娟 郑业焕 Xiang Jie;Cui Xiaoi;Li Luyi;Zhou Tongxi;Fang Juan;Zheng Yehuan(Autobio Diagnostics Co.,Ltd,Zhengzhou 450016)

机构地区：[1]郑州安图生物工程股份有限公司,郑州450016

出　　处：《中国抗生素杂志》2023年第11期1261-1266,共6页Chinese Journal of Antibiotics

摘　　要：目的通过荧光素酶生物发光法与微量肉汤稀释法对质控菌株和临床收集菌株的最低抑菌浓度(minimum inhibitory concentration,MIC)进行测定,探讨荧光素酶生物发光法在病原微生物快速药物敏感性试验中的应用价值。方法以D-荧光素钠、Mg^(2+)、O_(2)和待测菌株为荧光素酶催化反应底物,连续检测待测菌与抗生素孵育过程中相对光强度(relative light unit,RLU)变化,计算发光率并确定MIC,分析与微量肉汤稀释法的基本一致性(essential agreement,EA)和分类一致性(categorical agreement,CA)。结果荧光素酶生物发光法快速检测大肠埃希菌、金黄色葡萄球菌等质控菌株MIC与微量肉汤稀释法检测MIC结果CA和EA值均为100%;并应用该检测体系对91株临床革兰阴性菌样本、93株革兰阳性菌样本进行MIC测定,革兰阴性菌检测结果中的6种药物EA和CA分别大于90%和84%,革兰阳性菌检测结果中的6种药物EA和CA分别大于90%和80%,并使检测周期由常规药敏实验的16~24 h缩短至5~6 h。结论荧光素酶生物发光法快速检测药敏结果与微量肉汤稀释法结果具有较高一致性,且更快速、敏感,可为今后的临床检验工作提供全新的快速药敏检测方法。Objective The minimum inhibitory concentration(MIC)of quality-control bacteria and clinical strains was determined by the luciferase bioluminescence method and the broth microdilution method to explore the application value of the luciferase bioluminescence method in rapid antimicrobial susceptibility tests of pathogenic microorganisms.Methods Using D-luciferin sodium,Mg^(2+),O_(2) and the tested strain as substrates of the luciferase catalytic reaction,continuously detect the change of relative light unit(RLU)during the incubation of the tested bacterium with antibiotics,calculate the luminosity factor and determine the MIC,compare the results that were obtained using the broth micro-dilution(BMD)method,and then evaluate the essential agreement and categorical agreement.Results The CA and EA values of rapid MIC detection of quality control strains,such as E.coli and S.aureus,by the luciferase bioluminescence method and the MIC detection by the broth microdilution method were 100%.In addition,the detection system was applied to perform MIC detection on 91 clinical Gram-negative bacterial samples and 93 Gram-positive bacterial samples.Among the Gram-negative bacterial detection results,the EA and CA values of the 6 drugs were greater than 90%and 84%,respectively.The EA and CA values of the 6 drugs in the Gram-positive bacterial detection results were greater than 90%and 80%,respectively,and the detection duration was shortened from 16~24 h in conventional drug sensitivity experiments to 5~6 h.Conclusion The results of luciferase bioluminescence rapid antimicrobial susceptibility testing(RAST)were highly consistent with those obtained by the broth microdilution method,which was more rapid and sensitive than traditional methods.It could provide a new rapid drug sensitivity detection method for clinical testing in the future.

关 键 词：荧光素酶 发光率 一致性 快速药敏 

分 类 号：R9[医药卫生—药学]