Detection Of Toxigenic Clostridium Difficile In Stool By

艰难梭菌相关性疾病(CDAD)是接受抗生素或抗肿瘤药物治疗的住院患者的主要问题。目前诊断CDAD的实验室方法包括培养和检测由生物体或细胞相关抗原产生的毒素。用于诊断CDAD的聚合酶链式反应方法在文献中已有报道，但这些报告的范围有限，所使用的许多引物对效率低下或扩增艰难梭菌基因组的不适当部分。我们最初针对艰难梭菌的两个毒素基因选择了5对特异的引物。我们选择了在扩增毒素A和毒素B方面最有效的两对引物。这两个基因的碱基序列都非常富含AT，我们发现这阻碍了灵敏的光周期分析的发展。我们已经开发了一种用于扩增子检测的ELISA格式，目前正在确定我们的检测方法的敏感性，然后再使用患者样本。

Clostridium difficile-associated disease (CDAD) is a major problem for hospitalized patients recieving antibiotics or antineoplastic agents. Current labotatory methods for diagnosing CDAD include culture and assays for detection of the toxins produced by the organism or a cell-associated antigen. PCR assays for the diagnosis of CDAD have been reported in the liturature, but these reports are limited in scope and many of the primer pairs used are inefficient or amplify inappropriate portions of the C. difficile genome. We originally selected 5 primer pairs specific for the two toxin genes of C. difficile. We have selected the two primer pairs that are the most efficient in amplifying toxin A and toxin B. The base sequences for these two genes are both very AT rich, and we discovered that this prohibits the development of a sensitive Light Cycler assay. We have developed an ELISA format for our amplicon detection and are now in the process of determining the sensitivity of our assay before using patient specimens.