Rapid detection of antibiotic resistance by MALDI-TOF mass spectrometry using a novel direct-on-target microdroplet growth assay
Rapid detection of antibiotic resistance by MALDI-TOF mass spectrometry using a novel direct-on-target microdroplet growth assay
复制标题
DOI：
10.1016/j.cmi.2017.10.016
复制
发表时间：
2018-07-01
影响因子：
14.2
通讯作者：
Becker, K.
中科院分区：
文献类型：
--
作者：
Idelevich, E. A.;Sparbier, K.;Becker, K.
目的：我们的目的是开发一种通用的表型方法，它允许简单，快速的抗菌药物敏感性测试独立的潜在的耐药mechanism.Methods：我们建立了一种新的直接靶向微滴生长测定抗生素耐药性的检测在几个小时内，这是基于基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF MS）。将微生物在有和没有美罗培南的情况下在营养肉汤中作为微滴直接在MALDI-TOF MS靶标上孵育。随后，通过使微滴与吸收性材料接触来将肉汤与微生物细胞分离。通过MALDI-TOF MS检测在抗生素存在下生长的微生物。共使用24株肺炎克雷伯菌和24株铜绿假单胞菌分离株来评估美罗培南耐药性检测的性能。研究的微滴体积为2，4，6，8和10 μ L。结果：使用6 μ L的微滴实现了最佳性能。应用该体积，成功检测到所有生长对照品（有效试验的定义），孵育18小时后，所有分离株均正确归类为敏感或非敏感。克雷pneumoniae，6 mL微滴孵育4 h后，有效试验率、敏感性和特异性均达100%。使用相同的微滴体积为铜绿假单胞菌，孵育5小时导致83.3%的有效测试与100%的灵敏度和100% specificity.Conclusions：我们表现出简单，快速和准确的耐药性检测，以碳青霉烯类耐药革兰氏阴性菌为例。我们的技术适用于自动化，并可扩展到其他应用，例如同时检测多种抗生素以及直接从临床样品中进行耐药性测定。(c)2017年欧洲临床微生物学和传染病学会。由爱思唯尔有限公司出版。保留所有权利。
Objectives: We aimed to develop a universal phenotypic method, which allows easy and rapid antimicrobial susceptibility testing independently of underlying resistance mechanisms.Methods: We established a novel direct-on-target microdroplet growth assay for the detection of antibiotic resistance within a few hours, which is based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The microorganisms were incubated with and without meropenem in nutrient broth as microdroplets directly on MALDI-TOF MS target. Subsequently, broth was separated from microbial cells by contacting the microdroplets with an absorptive material. The microorganisms grown in the presence of antibiotic were detected by MALDI-TOF MS. A total of 24 Klebsiella pneumoniae and 24 Pseudomonas aeruginosa isolates were used to assess performance for detection of meropenem resistance. The microdroplet volumes investigated were 2, 4, 6, 8 and 10 mu L.Results: The best performance was achieved using 6-mu L microdroplets. Applying this volume, all growth controls were successfully detected (definition of valid test), and all isolates were correctly categorized as susceptible or non-susceptible after an 18-h incubation. For K. pneumoniae, rate of valid tests, sensitivity and specificity all reached 100% after a 4-h incubation of 6-mL microdroplets. Using the same microdroplet volume for P. aeruginosa, incubation for 5 h resulted in 83.3% of valid tests with 100% sensitivity and 100% specificity.Conclusions: We demonstrated easy, rapid and accurate resistance detection using carbapenem-resistant Gram-negative bacteria as an example. Our technology is suitable for automatization and expandable to further applications, e.g. simultaneous testing of multiple antibiotics as well as resistance determination directly from clinical samples. (c) 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.