Fraunhofer Institute for Cell Therapy and Immunology (IZI), Leipzig, Germany.
J Appl Microbiol. 2010 Oct;109(4):1150-8. doi: 10.1111/j.1365-2672.2010.04736.x. Epub 2010 Aug 19.
We established a real-time PCR assay for the detection and strain identification of Candida species and demonstrated the ability to differentiate between Candida albicans the most common species, and also Candida parapsilosis, Candida glabrata, Candida tropicalis and Candida dubliniensis by LightCycler PCR and melting curve analysis.
The DNA isolation from cultures and serum was established using the QIAmp Tissue Kit. The sensitivity of the assay was ≥ 2 genome equivalents/assay. It was possible to differentiate all investigated Candida species by melting curve analysis, and no cross-reaction to human DNA or Aspergillus species could be observed.
The established real-time PCR assay is a useful tool for the rapid identification of Candida species and a base technology for more complex PCR assays.
We carried out initial steps in validation of a PCR assay for the detection and differentiation of medically relevant Candida species. The PCR was improved by generating PCR standards, additional generation of melting curves for species identification and the possibility to investigate different specimens simultaneously.
我们建立了一种实时 PCR 检测方法，用于检测和鉴定念珠菌属，并通过 LightCycler PCR 和熔解曲线分析，展示了区分最常见的白色念珠菌以及近平滑念珠菌、光滑念珠菌、热带念珠菌和都柏林念珠菌的能力。
使用 QIAmp 组织试剂盒从培养物和血清中提取 DNA。该检测方法的灵敏度≥2 个基因组当量/检测。通过熔解曲线分析可以区分所有研究的念珠菌属，并且没有观察到与人类 DNA 或曲霉属的交叉反应。
建立的实时 PCR 检测方法是快速鉴定念珠菌属的有用工具，也是更复杂 PCR 检测的基础技术。
我们进行了用于检测和区分与医学相关的念珠菌属的 PCR 检测方法验证的初步步骤。通过生成 PCR 标准品、为物种鉴定额外生成熔解曲线以及同时研究不同样本的可能性，对 PCR 进行了改进。
