熒光素酶生物發(fā)光法在快速藥敏試驗中的研究與應(yīng)用

向杰?崔曉莉?李璐一?周同喜?方娟?鄭業(yè)煥

摘要：目的 通過熒光素酶生物發(fā)光法與微量肉湯稀釋法對質(zhì)控菌株和臨床收集菌株的最低抑菌濃度（minimum inhibitory concentration，MIC）進(jìn)行測定，探討熒光素酶生物發(fā)光法在病原微生物快速藥物敏感性試驗中的應(yīng)用價值。方法 以D-熒光素鈉、Mg2+、O2和待測菌株為熒光素酶催化反應(yīng)底物，連續(xù)檢測待測菌與抗生素孵育過程中相對光強(qiáng)度（relative light unit， RLU）變化，計算發(fā)光率并確定MIC，分析與微量肉湯稀釋法的基本一致性（essential agreement，EA）和分類一致性（categorical agreement，CA）。結(jié)果 熒光素酶生物發(fā)光法快速檢測大腸埃希菌、金黃色葡萄球菌等質(zhì)控菌株MIC與微量肉湯稀釋法檢測MIC結(jié)果CA和EA值均為100%；并應(yīng)用該檢測體系對91株臨床革蘭陰性菌樣本、93株革蘭陽性菌樣本進(jìn)行MIC測定，革蘭陰性菌檢測結(jié)果中的6種藥物EA和CA分別大于90%和84%，革蘭陽性菌檢測結(jié)果中的6種藥物EA和CA分別大于90%和80%，并使檢測周期由常規(guī)藥敏實驗的16～24 h縮短至5～6 h。結(jié)論 熒光素酶生物發(fā)光法快速檢測藥敏結(jié)果與微量肉湯稀釋法結(jié)果具有較高一致性，且更快速、敏感，可為今后的臨床檢驗工作提供全新的快速藥敏檢測方法。

關(guān)鍵詞：熒光素酶；發(fā)光率；一致性；快速藥敏

中圖分類號：R9文獻(xiàn)標(biāo)志碼：A

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

Xiang Jie， Cui Xiaoli， Li Luyi， Zhou Tongxi， Fang Juan， and Zheng Yehuan

（Autobio Diagnostics Co.， Ltd， Zhengzhou 450016）

Abstract 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， Mg2+， O2 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.

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