Diagnosis of Clostridium difficile infection: comparison of four methods on specimens collected in Cary-Blair transport medium and tcdB PCR on fresh versus frozen samples

Abstract

Clostridium difficile infection (CDI) caused by toxigenic strains of C. difficile is primarily a nosocomial infection with increasing prevalence. Stool specimens are typically collected in Cary-Blair transport medium to maximize culture-based detection of common stool pathogens. The goal of this study was to establish an analytically accurate and efficient algorithm for the detection of CDI in our patient population using samples collected in Cary-Blair transport medium. In addition, we wished to determine whether the sensitivity and specificity of PCR was affected by freezing samples before testing. Using 357 specimens, we compared four methods: enzyme immunoassay for the antigen glutamate dehydrogenase (Wampole™ C. DIFF CHEK-60 Assay, GDH), toxin A and B enzyme immunoassay (Remel ProSpecT™ C. difficile Toxin A/B Microplate Assay, Toxin EIA), cell culture cytotoxicity neutralization assay (Bartels™ Cytotoxicity Assay, CT), and real-time PCR targeting the toxin B gene (BD GeneOhm™ Cdiff Assay, PCR). The analytic sensitivity and specificity of each as determined using a combined gold standard were as follows: GDH, 100% and 93.2%; Toxin EIA, 82.9% and 82.9%; CT, 100% and 100%; PCR (performed on frozen specimens) 74.3% and 96.6%; respectively. However, the sensitivity and specificity of PCR improved to 100% when performed on 50 fresh stool samples collected in Cary-Blair. While CT remains a sensitive method for the detection of CDI, GDH offers an excellent initial screening method to rule out CDI. While the performance of each assay did not appear to be affected by collection in Cary-Blair medium, PCR performed better using fresh specimens.

Keywords: PCR; cary-Blair; clostridium difficile; frozen; glutamate dehydrogenase..