Detection Of Toxigenic Clostridium Difficile In Stool By

艰难梭菌相关疾病（CDAD）是住院患者使用抗生素或抗肿瘤药物的主要问题。目前诊断CDAD的实验室方法包括培养和检测生物体产生的毒素或细胞相关抗原的测定。PCR诊断CDAD的方法在文献中已有报道，但这些报道的范围有限，而且许多使用的引物对效率低下或扩增艰难梭菌基因组的不适当部分。我们最初选择了艰难梭菌两个毒素基因特异性的5对引物。我们选择了对毒素A和毒素b扩增效率最高的两个引物对。这两个基因的碱基序列都是非常丰富的AT，我们发现这阻碍了敏感光循环试验的发展。我们已经开发了一种用于扩增子检测的ELISA格式，目前正在使用患者标本之前确定我们检测的灵敏度。

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.