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.
Highlights
While glutamate dehydrogenase (GDH) antigen EIA and toxin Assay; Carlsbad, CA), and real-time PCR for for the detection of CDI, GDH offers an excellent EIA are well-studied on this medium, both toxin B gene
GDH antithe performance of each assay did not appear to are recommended for fresh samples without gen EIA, toxin EIA, and cell culture cytotoxicity be affected by collection in Cary-Blair medium, preservative
There were no samples that were positive for toxin-producing C. difficile but negative by GDH testing, resulting in a sensitivity and negative predictive value of 100%
Summary
In addi- mine the impact of freezing on PCR results, an tion, we sought to determine whether the additional 50 fresh specimens collected in Carysensitivity and specificity of PCR was affected Blair were tested upon arrival in the lab with. Clostridium difficile is an anaerobic, spore- by freezing samples before testing. GDH antigen EIA, toxin EIA, and PCR. All assays were performed according to the GDH antigen testing. There were no samples negative via CT but manufacturers’ instructions with the exception of specimens being collected in Cary-Blair transport medium (rather than raw), and freezing samples for PCR. The thawed transport tubes were vortex-mixed at high speed for 15 seconds and a sterile dry swab was dipped into the transport material. PCR for the tcdB gene was performed according to the manufacturer’s instructions
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