Veterinary Public Health and Food Safety Department, Istituto Superiore di Sanità, National Reference Centre for Botulism, Viale Regina Elena 299, 00161 Rome, Italy.
Int J Food Microbiol. 2011 Mar 1;145 Suppl 1:S152-7. doi: 10.1016/j.ijfoodmicro.2011.02.001. Epub 2011 Feb 24.
A real-time PCR method for detection and typing of BoNT-producing Clostridia types A, B, E, and F was developed on the framework of the European Research Project "Biotracer". A primary evaluation was carried out using 104 strains and 17 clinical and food samples linked to botulism cases. Results showed 100% relative accuracy, 100% relative sensitivity, 100% relative specificity, and 100% selectivity (inclusivity on 73 strains and exclusivity on 31 strains) of the real-time PCR against the reference cultural method combined with the standard mouse bioassay. Furthermore, a ring trial study performed at four different European laboratories in Italy, France, the Netherlands, and Sweden was carried out using 47 strains, and 30 clinical and food samples linked to botulism cases. Results showed a concordance of 95.7% among the four laboratories. The reproducibility generated a relative standard deviation in the range of 2.18% to 13.61%. Considering the high level of agreement achieved between the laboratories, this real-time PCR is a suitable method for rapid detection and typing of BoNT-producing Clostridia in clinical, food and environmental samples and thus support the use of it as an international standard method.
一种实时 PCR 方法用于检测和分型 BoNT 产生的梭菌 A、B、E 和 F 型，该方法是在欧洲研究项目“生物追踪器”的框架上开发的。使用 104 株菌和 17 株与肉毒中毒病例相关的临床和食品样本进行了初步评估。结果表明，实时 PCR 相对于结合标准小鼠生物测定的参考文化方法具有 100%的相对准确性、100%的相对灵敏度、100%的相对特异性和 100%的选择性（73 株菌包容性和 31 株菌排他性）。此外，在意大利、法国、荷兰和瑞典的四个欧洲实验室进行了一项环试验研究，使用了 47 株菌和 30 株与肉毒中毒病例相关的临床和食品样本。结果表明，四个实验室之间的一致性为 95.7%。重现性产生的相对标准偏差在 2.18%至 13.61%范围内。考虑到实验室之间达成的高度一致性，这种实时 PCR 是一种用于快速检测和分型临床、食品和环境样本中 BoNT 产生的梭菌的合适方法，因此支持将其用作国际标准方法。
