Goodacre R, Shann B, Gilbert R J, Timmins E M, McGovern A C, Alsberg B K, Kell D B, Logan N A
Institute of Biological Sciences, University of Wales, Aberystwyth, U.K.
Anal Chem. 2000 Jan 1;72(1):119-27. doi: 10.1021/ac990661i.
Thirty-six strains of aerobic endospore-forming bacteria confirmed by polyphasic taxonomic methods to belong to Bacillus amyloliquefaciens, Bacillus cereus, Bacillus licheniformis, Bacillus megaterium, Bacillus subtilis (including Bacillus niger and Bacillus globigii), Bacillus sphaericus, and Brevi laterosporus were grown axenically on nutrient agar, and vegetative and sporulated biomasses were analyzed by Curie-point pyrolysis mass spectrometry (PyMS) and diffuse reflectance-absorbance Fourier-transform infrared spectroscopy (FT-IR). Chemometric methods based on rule induction and genetic programming were used to determine the physiological state (vegetative cells or spores) correctly, and these methods produced mathematical rules which could be simply interpreted in biochemical terms. For PyMS it was found that m/z 105 was characteristic and is a pyridine ketonium ion (C6H3ON+) obtained from the pyrolysis of dipicolinic acid (pyridine-2,6-dicarboxylic acid; DPA), a substance found in spores but not in vegetative cells; this was confirmed using pyrolysis-gas chromatography/mass spectrometry. In addition, a pyridine ring vibration at 1447-1439 cm-1 from DPA was found to be highly characteristic of spores in FT-IR analysis. Thus, although the original data sets recorded hundreds of spectral variables from whole cells simultaneously, a simple biomarker can be used for the rapid and unequivocal detection of spores of these organisms.
通过多相分类方法确认属于解淀粉芽孢杆菌、蜡样芽孢杆菌、地衣芽孢杆菌、巨大芽孢杆菌、枯草芽孢杆菌（包括黑曲霉和球形芽孢杆菌）、球形芽孢杆菌和侧孢短芽孢杆菌的36株需氧产芽孢细菌在营养琼脂上无菌培养，通过居里点热解质谱法（PyMS）和漫反射吸收傅里叶变换红外光谱法（FT-IR）分析营养体和芽孢生物质。基于规则归纳和遗传编程的化学计量学方法被用于正确确定生理状态（营养细胞或芽孢），这些方法产生了可以简单地用生化术语解释的数学规则。对于PyMS，发现m/z 105具有特征性，它是一种吡啶鎓离子（C6H3ON+），由吡啶二羧酸（吡啶-2,6-二羧酸；DPA）热解产生，DPA是一种存在于芽孢中但不存在于营养细胞中的物质；这通过热解气相色谱/质谱法得到了证实。此外，在FT-IR分析中，发现来自DPA的1447 - 1439 cm-1处的吡啶环振动是芽孢的高度特征性信号。因此，尽管原始数据集同时记录了来自全细胞的数百个光谱变量，但一个简单的生物标志物可用于快速明确地检测这些生物体的芽孢。
