Clostridium difficile ribotyping using automated fragment sizing technology

Abstract

This research project looked at an alternative methodology for ribotyping Clostridium difficile capillary gel electrophoresis. This method is based on the conventional ribotyping method as the intergenic spacer region between the 16S and 23S is copied by PCR. However the primers that were designed for this research project have a fluorescent tag attached. The PCR product can then be visualised using a fragment analyser which detects the fluorescent produces and assigns them a size. A small library had been previously constructed using this ribotyping method which was used in a blind study of 195 isolates supplied by the Anaerobic Reference Laboratory (ARL) Cardiff. After assigning ribotypes to all the isolates the results were then verified against those of ARL Cardiff. The results showed that an accuracy of 94.8% had been achieved. It also highlighted that the small number of ribotypes held in the library that had been constructed for this project had played a part in the number of unidentified ribotypes. This project showed that with a large library of ribotypes this method is reproducible quick and cost effective method for ribotyping Clostridium difficile.