Rapid Identification of Pseudomonas spp. via Raman Spectroscopy Using Pyoverdine as Capture Probe,Analytical Chemistry

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Rapid Identification of Pseudomonas spp. via Raman Spectroscopy Using Pyoverdine as Capture Probe
Analytical Chemistry ( IF 6.7 ) Pub Date : 2016-01-08 00:00:00 , DOI: 10.1021/acs.analchem.5b02829
Susanne Pahlow _{1,

2} , Stephan Stöckel _{1,

2} , Sibyll Pollok ₃ , Dana Cialla-May _{1,

2,

4} , Petra Rösch _{1,

2} , Karina Weber _{1,

2,

4} , Jürgen Popp _{1,

2,

4}

Affiliation

Pyoverdine is a substance which is excreted by fluorescent pseudomonads in order to scavenge iron from their environment. Due to specific receptors of the bacterial cell wall, the iron loaded pyoverdine molecules are recognized and transported into the cell. This process can be exploited for developing efficient isolation and enrichment strategies for members of the Pseudomonas genus, which are capable of colonizing various environments and also include human pathogens like P. aeruginosa and the less virulent P. fluorescens. A significant advantage over antibody based systems is the fact that siderophores like pyoverdine can be considered as “immutable ligands,” since the probability for mutations within the siderophore uptake systems of bacteria is very low. While each species of Pseudomonas usually produces structurally unique pyoverdines, which can be utilized only by the producer strain, cross reactivity does occur. In order to achieve a reliable identification of the captured pathogens, further investigations of the isolated cells are necessary. In this proof of concept study, we combine the advantages of an isolation strategy relying on “immutable ligands” with the high specificity and speed of Raman microspectroscopy. In order to isolate the bacterial cells, pyoverdine was immobilized covalently on planar aluminum chip substrates. After capturing, single cell Raman spectra of the isolated species were acquired. Due to the specific spectroscopic fingerprint of each species, the bacteria can be identified. This approach allows a very rapid detection of potential pathogens, since time-consuming culturing steps are unnecessary. We could prove that pyoverdine based isolation of bacteria is fully Raman compatible and further investigated the capability of this approach by isolating and identifying P. aeruginosa and P. fluorescens from tap water samples, which are both opportunistic pathogens and can pose a threat for immunocompromised patients.

中文翻译：

快速鉴定假单胞菌。通过吡咯烷定为捕获探针的拉曼光谱

Pyoverdine是一种由荧光假单胞菌排泄的物质，目的是清除环境中的铁。由于细菌细胞壁的特定受体，铁负载的吡啶二酮分子被识别并转运到细胞中。可以利用该过程为假单胞菌属的成员开发有效的分离和富集策略，该策略能够在各种环境中定殖，还包括人类病原体，如铜绿假单胞菌和毒性较低的荧光假单胞菌。。与基于抗体的系统相比，一个显着的优势是，像吡夫定这样的铁载体可以被认为是“不可变的配体”，因为细菌在铁载体吸收系统中发生突变的可能性非常低。而每种假单胞菌通常生产结构独特的丙酮丁二酮，只有生产菌株才能利用，但确实会发生交叉反应。为了可靠地识别捕获的病原体，有必要对分离的细胞进行进一步的研究。在此概念验证研究中，我们结合了依靠“不变配体”的分离策略的优势以及拉曼光谱的高特异性和高速度。为了分离细菌细胞，将吡over定共价固定在平坦的铝芯片基底上。捕获后，获得分离物种的单细胞拉曼光谱。由于每种物种具有特定的光谱指纹，因此可以识别细菌。由于不需要费时的培养步骤，因此该方法可以非常快速地检测潜在的病原体。来自自来水样品的铜绿假单胞菌和荧光假单胞菌都是机会致病菌，可能对免疫功能低下的患者构成威胁。

更新日期：2016-01-08

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