通过实时聚合酶链式反应检测类似高达干酪的奶酪上存活和死亡的单核细胞增生李斯特菌。
Detection of viable and dead Listeria monocytogenes on gouda-like cheeses by real-time PCR.
作者信息
Rudi K, Naterstad K, Drømtorp S M, Holo H
机构信息
MATFORSK, Norwegian Food Research Institute, Osloveien, As, Norway.
出版信息
Lett Appl Microbiol. 2005;40(4):301-6. doi: 10.1111/j.1472-765X.2005.01672.x.
AIMS
Surface contamination by Listeria monocytogenes of gouda-like cheeses during processing represents a potential public health problem. The aim of this work was to develop novel real-time PCR diagnostics to detect the presence of viable, dead or viable but not culturable (VBNC) cells on gouda-like cheeses.
METHODS AND RESULTS
We used ethidium monoazide bromide (EMA)-PCR for direct quantification of viable and dead cells, while semiquantitative detection of culturable cells below the PCR detection limit (c. 100 CFU g(-1)) was obtained by combining growth and real-time PCR. We were able to quantify the fraction of >0.5% viable cells in a background of dead cells by EMA-PCR, given that the viable cell concentration was above the PCR detection limit. The combined growth and real-time PCR complemented the EMA-PCR, and enabled semiquantitative detection of low levels of culturable cells (10 and 100 CFU g(-1)).
SIGNIFICANCE AND IMPACT OF THE STUDY
The significance of this work is that we have developed a novel concept for detection of viable and potentially infectious L. monocytogenes.
目的
在加工过程中，类高达干酪被单核细胞增生李斯特菌表面污染是一个潜在的公共卫生问题。本研究的目的是开发新型实时聚合酶链反应诊断方法，以检测类高达干酪上活的、死的或活的但不可培养（VBNC）细胞的存在。
方法与结果
我们使用溴化乙锭单叠氮化物（EMA）-PCR直接定量活细胞和死细胞，同时通过结合生长和实时PCR对低于PCR检测限（约100 CFU g⁻¹）的可培养细胞进行半定量检测。当活细胞浓度高于PCR检测限时，我们能够通过EMA-PCR在死细胞背景中定量>0.5%的活细胞比例。生长与实时PCR相结合补充了EMA-PCR，并能够对低水平的可培养细胞（10和100 CFU g⁻¹）进行半定量检测。
研究的意义和影响
这项工作的意义在于我们开发了一种检测活的和潜在感染性单核细胞增生李斯特菌的新概念。
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