Use of PCR Coupled with Electrospray Ionization Mass Spectrometry for Rapid Identification of Bacterial and Yeast Bloodstream Pathogens from Blood Culture Bottles
Use of PCR Coupled with Electrospray Ionization Mass Spectrometry for Rapid Identification of Bacterial and Yeast Bloodstream Pathogens from Blood Culture Bottles
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DOI：
10.1128/jcm.00936-10
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发表时间：
2011-01-01
影响因子：
9.4
通讯作者：
Wolk, Donna M.
中科院分区：
文献类型：
--
作者：
Kaleta, Erin J.;Clark, Andrew E.;Wolk, Donna M.
在美国，脓毒症是导致死亡的十大原因之一。快速使用抗生素是患者存活的最重要因素之一，但目前只有有限的方法可以快速鉴定血液感染中培养的微生物，这些微生物可能导致败血症。虽然传统的单靶点分子方法已被证明能够使广谱抗生素快速降级，从而极大地提高败血症患者的存活率，但多重方法提供了更大的可能性。采用聚合酶链式反应-电喷雾电离-质谱法(PCR/ESI-MS)对引起人类血液感染的微生物进行了鉴定。直接从234个Bact-Alert血培养瓶中提取DNA，并与临床参考标准方法进行比较。结果表明，在属和种水平上的符合率分别为98.7%和96.6%。在29个血液培养瓶中发现了混合微生物，包括同一属的混合物种，以及含有革兰氏阳性和革兰氏阴性微生物的混合微生物，这证明了PCR/ESI-MS无需培养即可同时识别多种微生物的能力。与常规的血液培养瓶传代培养和微生物表型鉴定相比，这项研究显示出更高的分析准确性。在不预先知道可能存在的微生物的情况下，在自动血培养系统发出阳性警报后，PCR/ESI-MS方法可以在短短5到6小时内提供准确的结果；然而，目前的批量模式测试限制了该方法的临床应用。
Sepsis is among the top 10 causes of mortality in the United States. Rapid administration of antibiotics is one of the most important contributors to patient survival, yet only a limited number of methods exist for rapid identification of microbes cultivated from bloodstream infections, which can lead to sepsis. While traditional single-target molecular methods have been shown to greatly improve survival for septic patients by enabling rapid deescalation of broad-spectrum antibiotics, multiplex methods offer even greater possibilities. A novel multiplex method, PCR coupled to electrospray ionization mass spectrometry (PCR/ESI-MS), was used to identify the genus and species of microorganisms found to cause human bloodstream infections. DNA was directly extracted from 234 BacT-Alert blood culture bottles, and results were compared to those obtained by clinical reference standard methods. The study results demonstrated 98.7% and 96.6% concordance at the genus and species levels, respectively. Mixtures of microbes were identified in 29 blood culture bottles, including mixed species of the same genus, as well as mixtures containing Gram-positive and Gram-negative organisms, exemplifying the PCR/ESI-MS capability to identify multiple organisms simultaneously without the need for cultivation. This study demonstrates high analytical accuracy in comparison to routine subculture of blood culture bottles and phenotypic identification of microbes. Without foreknowledge of the microorganisms potentially present, the PCR/ESI-MS methods can deliver accurate results in as little as 5 to 6 h after a positive alarm from the automated blood culture system; however, current batch mode testing limits the method's clinical utility at this time.