Clostridium difficile PCR Cycle Threshold Predicts Free Toxin.

There is no stand-alone diagnostic that can sensitively and rapidly detect fecal free toxins. We investigated the performance of the PCR cycle threshold ( ) for predicting free toxin status. Consecutive stool samples ( = 312) positive for toxigenic by the GeneXpert /Epi PCR assay were tested with the rapid membrane C. Diff Quik Chek Complete immunoassay (RMEIA). RMEIA toxin-negative samples were tested with the cell cytotoxicity neutralization assay (CCNA) and tgcBIOMICS enzyme-linked immunosorbent assay (ELISA). Using RMEIA alone or in combination with CCNA and/or ELISA as the reference method, the accuracy of was measured at different cutoffs. Using RMEIA as the reference method, a cutoff of 26.35 detected toxin-positive samples with a sensitivity, specificity, positive predictive value, and negative predictive value of 96.0% (95% confidence interval [CI], 90.2% to 98.9%), 65.9% (95% CI, 59.0% to 72.2%), 57.4% (95% CI, 52.7% to 62%), and 97.1% (95% CI, 92.8% to 98.9), respectively. Inclusion of CCNA in the reference method improved specificity to 78.0% (95% CI, 70.7% to 84.2%). Intercartridge lot variability measured as the average coefficient of variation was 2.8% (95% CI, 1.2% to 3.2%). Standardizing the input stool volume did not improve toxin specificity. The median values were not significantly different between stool samples with Bristol scores of 5, 6, and 7, between pediatric and adult samples, or between presumptive 027 and non-027 strains. In addition to sensitively detecting toxigenic in stool, on-demand PCR may also be used to accurately predict toxin-negative stool samples, thus providing additional results in PCR-positive stool samples to guide therapy.