Antwerpen Markus H, Zimmermann Pia, Bewley Kevin, Frangoulidis Dimitrios, Meyer Hermann
Bundeswehr Institute of Microbiology, Neuherbergstrasse 11, D-80937 Munich, Germany.
Mol Cell Probes. 2008 Oct-Dec;22(5-6):313-5. doi: 10.1016/j.mcp.2008.06.001. Epub 2008 Jun 17.
Specific identification of Bacillus anthracis and differentiation from closely related Bacillus cereus and Bacillus thuringiensis strains is still a major diagnostic problem. Commercially available diagnostic kits targeting plasmid-markers cannot differentiate between B. anthracis, non-anthracis Bacillus species harbouring anthrax-specific virulence plasmids, and plasmidless B. anthracis strains. A TaqMan PCR assay was designed targeting sequences of gene locus BA_5345 of the B. anthracis strain Ames. Specificity was determined by using a panel of 328 Bacillus strains; sensitivity was determined by probit analysis. All B. anthracis isolates (n=92) were specifically detected by using the genomic TaqMan PCR assay whereas 236 strains belonging to 19 Bacillus species other than B. anthracis were PCR negative. The detection limit was determined to be 12.7 copies per reaction (95% confidence interval 10.2-17.5 copies). Here we present an extensively evaluated and - to our current knowledge - specific TaqMan PCR assay for the detection of B. anthracis based on a chromosomal marker.
炭疽芽孢杆菌的特异性鉴定以及与密切相关的蜡样芽孢杆菌和苏云金芽孢杆菌菌株的区分仍然是一个主要的诊断难题。针对质粒标记物的市售诊断试剂盒无法区分炭疽芽孢杆菌、携带炭疽特异性毒力质粒的非炭疽芽孢杆菌物种以及无质粒的炭疽芽孢杆菌菌株。设计了一种针对炭疽芽孢杆菌菌株Ames基因座BA_5345序列的TaqMan PCR检测方法。通过使用一组328株芽孢杆菌菌株确定特异性；通过概率分析确定灵敏度。使用基因组TaqMan PCR检测方法特异性地检测到所有炭疽芽孢杆菌分离株（n = 92），而属于除炭疽芽孢杆菌外的19种芽孢杆菌物种的236株菌株PCR检测呈阴性。检测限确定为每个反应12.7个拷贝（95%置信区间10.2 - 17.5个拷贝）。在此，我们展示了一种经过广泛评估且据我们目前所知基于染色体标记物用于检测炭疽芽孢杆菌的特异性TaqMan PCR检测方法。
