Biomedical Spectroscopy (P 25), Robert-Koch-Institut, Nordufer 20, D-13353 Berlin, Germany.
Anal Chem. 2010 Oct 15;82(20):8464-75. doi: 10.1021/ac101036s.
Yersinia are Gram-negative, rod-shaped facultative anaerobes, and some of them, Yersinia enterocolitica, Yersinia pseudotuberculosis, and Yersinia pestis, are pathogenic in humans. Rapid and accurate identification of Yersinia strains is essential for appropriate therapeutic management and timely intervention for infection control. In the past decade matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) in combination with computer-aided pattern recognition has evolved as a rapid, objective, and reliable technique for microbial identification. In this comprehensive study a total of 146 strains of all currently known Yersinia species complemented by 35 strains of other relevant genera of the Enterobacteriaceae family were investigated by MALDI-TOF MS and chemometrics. Bacterial sample preparation included microbial inactivation according to a recently developed mass spectrometry compatible inactivation protocol. The mass spectral profiles were evaluated by supervised feature selection methods to identify family-, genus-, and species-specific biomarker proteins and--for classification purposes--by pattern recognition techniques. Unsupervised hierarchical cluster analysis revealed a high degree of correlation between bacterial taxonomy and subproteome-based MALDI-TOF MS classification. Furthermore, classification analysis by supervised artificial neural networks allowed identification of strains of Y. pestis with an accuracy of 100%. In-depth analysis of proteomic data demonstrated the existence of Yersinia-specific biomarkers at m/z 4350 and 6046. In addition, we could also identify species-specific biomarkers of Y. enterocolitica at m/z 7262, 9238, and 9608. For Y. pseudotuberculosis a combination of biomarkers at m/z 6474, 7274, and 9268 turned out to be specific, while a peak combination at m/z 3065, 6637, and 9659 was characteristic for strains of Y. pestis. Bioinformatic approaches and tandem mass spectrometry were employed to reveal the molecular identity of biomarker ions. In this way, the Y. pestis-specific biomarker at m/z 3065 could be identified as a fragment of the plasmid-encoded plasminogen activator, one of the major virulence factors in plague infections.
耶尔森氏菌是革兰氏阴性、杆状的兼性厌氧菌，其中一些，如小肠结肠炎耶尔森氏菌、假结核耶尔森氏菌和鼠疫耶尔森氏菌，对人类具有致病性。快速准确地鉴定耶尔森氏菌菌株对于适当的治疗管理和及时的感染控制干预至关重要。在过去的十年中，基质辅助激光解吸电离飞行时间（MALDI-TOF）质谱（MS）结合计算机辅助模式识别已发展成为一种快速、客观和可靠的微生物鉴定技术。在这项全面的研究中，总共研究了 146 株已知的所有耶尔森氏菌物种，同时还研究了 35 株其他相关肠杆菌科属的菌株，采用 MALDI-TOF MS 和化学计量学方法进行分析。细菌样品制备包括根据最近开发的与质谱兼容的失活动力学协议进行微生物失活。通过监督特征选择方法评估质谱图谱，以鉴定家族、属和种特异性生物标志物蛋白，并通过模式识别技术进行分类。非监督层次聚类分析显示细菌分类法与基于亚蛋白组的 MALDI-TOF MS 分类之间存在高度相关性。此外，通过监督人工神经网络的分类分析，能够以 100%的准确率鉴定鼠疫耶尔森氏菌菌株。对蛋白质组数据的深入分析表明，在 m/z 4350 和 6046 处存在耶尔森氏菌特异性生物标志物。此外，我们还可以在 m/z 7262、9238 和 9608 处鉴定出小肠结肠炎耶尔森氏菌的特异性生物标志物。对于假结核耶尔森氏菌，m/z 6474、7274 和 9268 的生物标志物组合是特异性的，而 m/z 3065、6637 和 9659 的峰组合则是鼠疫耶尔森氏菌菌株的特征。生物信息学方法和串联质谱用于揭示生物标志物离子的分子身份。通过这种方式，可以鉴定出 m/z 3065 处的鼠疫耶尔森氏菌特异性生物标志物为质粒编码的纤溶酶原激活物的一个片段，该片段是鼠疫感染中主要毒力因子之一。
