Research Group Microbiology and Systems Biology, TNO Earth, Environmental and Life Sciences, Zeist, The Netherlands.
Appl Environ Microbiol. 2011 Sep;77(18):6433-40. doi: 10.1128/AEM.00142-11. Epub 2011 Jul 15.
A novel generic approach for stress profiling was applied to Listeria monocytogenes strain F2365. This food-borne pathogen was exposed to gradients of five different stresses of increasing intensity, typically ranging from moderate to lethal conditions. The stress factors included heat, acidic pH, a detergent disinfectant, an oxidant, and hyperosmotic conditions. In addition to CFU counts and lag time, five different molecular viability parameters were measured by fluorescence-based assays, including membrane integrity, membrane potential, esterase activity, redox activity, and intracellular pH stability. The last was measured by our recently invented real-time viability assay. Exposure to all stresses resulted in clear dose-response relationships for all viability parameters with the exception of hyperosmotic conditions. A statistical analysis showed strong correlations for (i) the growth parameters plate counts and lag times, (ii) the enzyme-associated functions redox and esterase activity, and (iii) the membrane-associated pH stability and membrane integrity. Results indicated a pronounced difference in the susceptibilities of the measured parameters depending on the stress factor applied. However, at relatively high stress intensities, all of the viability parameters became affected independent of the stress factor. Applications of the approach presented here include studies on the mechanism of action of unknown compounds with biocidal activity and a comparative analysis of the severities of the impact of stress conditions of interest. It appears that a meaningful evaluation of the impact of mild stress conditions can be obtained only through measurement of multiple viability parameters.
一种新颖的通用应激分析方法被应用于单核细胞增生李斯特菌 F2365 菌株。这种食源性病原体暴露于五种不同强度递增的应激梯度中，通常范围从温和到致死条件。应激因素包括热、酸性 pH 值、清洁剂消毒剂、氧化剂和高渗条件。除了 CFU 计数和滞后时间外，还通过荧光基测定法测量了五个不同的分子存活参数，包括膜完整性、膜电位、酯酶活性、氧化还原活性和细胞内 pH 值稳定性。最后一个参数是通过我们最近发明的实时存活测定法测量的。所有应激因素的暴露都导致所有存活参数的明显剂量-反应关系，除了高渗条件。统计分析表明，（i）生长参数平板计数和滞后时间，（ii）与酶相关的功能氧化还原和酯酶活性，以及（iii）与膜相关的 pH 值稳定性和膜完整性之间存在很强的相关性。结果表明，所测量的参数对应用的应激因素的敏感性存在明显差异。然而，在相对较高的应激强度下，所有存活参数都受到影响，而与应激因素无关。这里提出的方法的应用包括对具有杀菌活性的未知化合物的作用机制的研究以及对感兴趣的应激条件的严重程度的比较分析。似乎只有通过测量多个存活参数才能对轻度应激条件的影响进行有意义的评估。
