Rapid Detection by Real Time Polymerase Chain Reaction (RT-PCR) of Environmental Reservoirs of Clostridioides Difficile: A Comparison with Culture Methods and the Relationship to the Treatment of Symptomatic Infection.

Clostridioides difficileinfection is an important healthcare-associated infection and inadequate decontamination of the healthcare environment results in transmission and outbreaks. Environmental surveillance for C. difficileis challenging and there are no internationally agreed recommendations on how to do it. In this thesis we compared two culture-based recovery methods in vitro(contact plates and flocked swabs). We then optimised a PCR assay for the detection of the Toxin B producing gene tcdBand the housekeeping gene cdd3. We undertook a six month period of sampling the environment of patients known to be colonised with C. difficilein a tertiary referral hospital in Dublin.In vitro, we found that contact plates were superior at spore recovery from hospital materials compared to flocked swabs. During the six month sampling period, 44 near patient areas of C. difficilecolonised patients were sampled resulting in a total of 352 samples. Although contact plates were superior at recovering C. difficilespores in vitro, in the clinical setting moistened flocked swabs recovered more C. difficileby both culture-based methods and by PCR. The sensitivity and specificity of the tcdBPCR assay was 76% and 91%, respectively, when compared to culture methods while the sensitivity and the specificity of the cdd3 PCR assay were 60% and 89%, respectively. PCR was not as sensitive or specific when compared to culture-based methods. However, with further work it could offer a more rapid and viable alternative to culture-based methods. This would have positive implications for infection prevention and control practices.