Fach P, Hauser D, Guillou J P, Popoff M R
Centre National d'Etudes Vétèrinaires et Alimentaires/Laboratoire Central d'Hygiène Alimentaire, Paris, France.
J Appl Bacteriol. 1993 Sep;75(3):234-9. doi: 10.1111/j.1365-2672.1993.tb02771.x.
A polymerase chain reaction (PCR) was developed for the detection of Clostridium botulinum type A, a cause of human botulism. A two primer set and an oligonucleotide detection probe were used to specifically detect Cl. botulinum type A neurotoxin gene (BoNT/A). After 40 cycles of amplification, detection of a 798 bp amplified DNA fragment was carried out by agarose gel electrophoresis and Southern blot hybridization. This assay was able to detect 12.5 fg of purified target DNA or five bacteria per reaction. The sensitivity in artificially contaminated food samples after an 18 h enrichment step ranges from 10 to 10(3) bacteria per g according to the type of food samples. No cross-reactions were observed with the other Cl. botulinum toxinotypes and other bacteria found routinely in food. This PCR method may provide a suitable and rapid alternative to standard techniques for detection of Cl. botulinum type A in food samples.
已开发出一种聚合酶链反应（PCR）用于检测A型肉毒梭菌，它是人类肉毒中毒的一个病因。使用一组双引物和一个寡核苷酸检测探针来特异性检测A型肉毒梭菌神经毒素基因（BoNT/A）。经过40个循环的扩增后，通过琼脂糖凝胶电泳和Southern印迹杂交对798 bp的扩增DNA片段进行检测。该检测方法每个反应能够检测到12.5 fg的纯化目标DNA或五个细菌。在经过18小时富集步骤后，在人工污染的食品样本中的灵敏度根据食品样本类型在每克10至10³个细菌之间。未观察到与其他肉毒梭菌毒素型以及食品中常规发现的其他细菌发生交叉反应。这种PCR方法可能为检测食品样本中的A型肉毒梭菌提供一种合适且快速的标准技术替代方法。
