Sergeev Nikolay, Distler Margaret, Vargas Melany, Chizhikov Vladimir, Herold Keith E, Rasooly Avraham
FDA Center for Devices and Radiological Health, USA.
J Microbiol Methods. 2006 Jun;65(3):488-502. doi: 10.1016/j.mimet.2005.09.013. Epub 2005 Oct 20.
Bacillus cereus, B. thuringiensis and B. anthracis are closely related medically and economically important bacterial species that belong to the B. cereus group. Members of the B. cereus group carry genes encoding several important virulence factors, including enterotoxins, phospholipases and exotoxins. Since it is difficult to differentiate among B. cereus group members, and because Bacillus virulence factors are very important for pathogenesis, we explored the use of microarray-based detection of virulence factor genes as a tool for strain identification and for determining virulence. Our method requires an initial multiplex PCR amplification step, followed by identification of the PCR amplicons by hybridization to an oligonucleotide microarray containing genes for all three types of Bacillus virulence factors including B. anthracis virulence factors. The DNA chip described here contains 21 identical arrays used for analysis of seven samples in triplicates. Using the arrays, we found that virulence factors are present in several combinations in the strains analyzed. This work also demonstrates the potential of oligonucleotide microarrays for medical, food safety and biodefense analysis of microbial pathogens.
蜡样芽孢杆菌、苏云金芽孢杆菌和炭疽芽孢杆菌是医学和经济上重要的密切相关细菌物种，属于蜡样芽孢杆菌群。蜡样芽孢杆菌群的成员携带编码几种重要毒力因子的基因，包括肠毒素、磷脂酶和外毒素。由于蜡样芽孢杆菌群成员之间难以区分，且芽孢杆菌毒力因子对发病机制非常重要，我们探索了使用基于微阵列的毒力因子基因检测作为菌株鉴定和确定毒力的工具。我们的方法需要一个初始多重PCR扩增步骤，随后通过与包含所有三种芽孢杆菌毒力因子（包括炭疽芽孢杆菌毒力因子）基因的寡核苷酸微阵列杂交来鉴定PCR扩增子。这里描述的DNA芯片包含21个相同的阵列，用于对七个样品进行一式三份分析。使用这些阵列，我们发现毒力因子在分析的菌株中以几种组合形式存在。这项工作还证明了寡核苷酸微阵列在微生物病原体的医学、食品安全和生物防御分析中的潜力。
