Exploration Of A Multiple Real-time Polemerase Chain Reaction For Detecting Clostridium Difficile Infection

Objective: As the hypervirulent strains of Clostridium Difficile such as Ribotype 027 appears, the morbidity and mortality of Clostridium Difficile Infection increase. A fast and accurate laboratory diagnosis can improve the outcome of the patients and reduce the costs. This research intends to establish a quantitative PCR methods which can detect tcdA 、tcdB and TPI simultaneously.Method: Stools sent by clinicians that up to standard from October,2014 to March, 2015 were collected, isolated and cultured, the suspected colonies were identified by Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry and amplification of 16 sRNA. The strains isolated were then detected tcdA 、 tcdB and TPI by PCR and multiple qPCR based on Taqman probe.Results: 20 strains were isolated form 107 stools which match the demands. 4 strains are tcdA-tcd B-, and 16 strains are tcdA+tcdB+,tcdA-tcdB+ strain is not found. The positive rate is 18.7%. When detecting the standard strain using the primers and probe designed by us, the amplification rate of tcdA and TPI is 91% and 79%, and the limit of detection is 101 and 102, respectively.Conclusion: The proportion of strains that have toxin genes is high,we should pay attention to it. The primers and probes used in thisresearch should be improved.