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首页> 美国卫生研究院文献>Antimicrobial Agents and Chemotherapy >Measurement of Effects of Antibiotics in Bioluminescent Staphylococcus aureus RN4220
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Measurement of Effects of Antibiotics in Bioluminescent Staphylococcus aureus RN4220

机译：抗生素对生物发光金黄色葡萄球菌RN4220影响的测量

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摘要

The spread of antibiotic resistance among pathogenic bacteria is a serious threat to humans and animals. Therefore, unnecessary use should be minimized, and new antimicrobial agents with novel mechanisms of action are needed. We have developed an efficient method for measuring the action of antibiotics which is applied to a gram-positive strain, Staphylococcus aureus RN4220. The method utilizes the firefly luciferase reporter gene coupled to the metal-inducible cadA promoter in a plasmid, pTOO24. Correctly timed induction by micromolar concentrations of antimonite rapidly triggers the luciferase gene transcription and translation. This sensitizes the detection system to the action of antibiotics, and especially for transcriptional and translational inhibitors. We show the results for 11 model antibiotics with the present approach and compare them to an analytical setup with a strain where luciferase expression is under the regulation of a constitutive promoter giving only a report of metabolic inhibition. The measurement of light emission from intact living cells is shown to correlate extremely well (r = 0.99) with the conventional overnight growth inhibition measurement. Four of the antibiotics were within a 20% concentration range and four were within a 60% concentration range of the drugs tested. This approach shortens the assay time needed, and it can be performed in 1 to 4 h, depending on the sensitivity needed. Furthermore, the assay can be automatized for high-throughput screening by the pharmaceutical industry.
机译：抗生素耐药性在病原细菌中的传播对人类和动物构成了严重威胁。因此，应尽量减少不必要的使用，并且需要具有新颖作用机理的新型抗菌剂。我们已经开发出一种有效的方法来测量抗生素的作用，该方法适用于革兰氏阳性菌株金黄色葡萄球菌RN4220。该方法利用与萤光虫萤光素酶报告基因偶联的质粒pTOO24中的金属诱导的cadA启动子。通过微摩尔浓度的锑矿进行正确定时的诱导会迅速触发荧光素酶基因的转录和翻译。这会使检测系统对抗生素的作用敏感，尤其是对于转录和翻译抑制剂。我们用本方法显示了11种模型抗生素的结果，并将它们与荧光素酶表达处于组成型启动子调控下的菌株的分析设置进行比较，仅给出了代谢抑制的报告。从完整的活细胞发出的光的测量结果显示与常规的过夜生长抑制测量值非常相关（r = 0.99）。其中四种抗生素在20％的浓度范围内，四种抗生素在60％的浓度范围内。这种方法缩短了所需的测定时间，取决于所需的灵敏度，可以在1-4小时内完成。此外，该测定可以自动化进行制药行业的高通量筛选。

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