Role of culture and toxin detection in laboratory testing for diagnosis ofClostridium difficile-associated diarrhea

Abstract

Two variations of an egg yolk agar base medium containing cycloserine, cefoxitin, and fructose (CCFA), one with 250 micrograms and other with 500 micrograms of cycloserine/ml of agar medium were compared to study the effect of the cycloserine concentration on recovery of Clostridium difficile from stool samples. In addition, the role of prior anaerobic reduction of these media in the detection of Clostridium difficile-associated diarrhea (CDAD) was tested. Each medium was studied over a two-month period, with outcome compared between the testing periods and to historical data from our institution. Clinical correlation of test results was performed. The use of the originally described formulation of CCFA with 500 microgram of cycloserine/ml of agar combined with 4 h of anaerobic reduction prior to specimen inoculation increased the rate of isolation of toxigenic Clostridium difficile from clinical specimens from 6 to 17% (p < 0.001). Combining direct detection of stool toxin and properly performed culture for toxigenic Clostridium difficile enhances the potential for diagnosis of CDAD. For optimal performance the culture medium should contain the originally proposed cycloserine concentration of 500 microgram/ml of agar and should be anaerobically reduced at least 4 h prior to specimen inoculation.

Yield of Stool Culture with Isolate Toxin Testing versus a Two-Step Algorithm Including Stool Toxin Testing for Detection of Toxigenic Clostridium difficile

“…However, the data are hard to interpret, since actual and described culture techniques (such as the use of enrichment procedures, heat or alcohol shock, and the composition of CCFA) vary (14), as do the tests against which they are compared, the number of tests performed, and the populations tested. Comparative North American and European studies have demonstrated that prereduced CCFA with a high cycloserine concentration, as proposed by George et al (15), may be optimal (26,30). To improve validity and comparability, we chose to use standard commercial media for culture, to assess toxigenicity by CCNA, to include only one observation per patient, and to do parallel testing (two-step algorithm and PCR) for all patients.…”

Six Rapid Tests for Direct Detection of Clostridium difficile and Its Toxins in Fecal Samples Compared with the Fibroblast Cytotoxicity Assay

“…While nucleic acid amplification of C. difficile toxin genes in stool remains an option, toxigenic culture is by far the most commonly recommended confirmatory method, and the Society for Healthcare Epidemiology of America in fact recommends culture in addition to direct fecal toxin testing (5,8,13,20,26). Toxigenic culture and one of the rapid methods evaluated in this study could be combined in a single algorithm as follows: (i) all fecal samples would be tested with an IA test; (ii) samples displaying a positive result would be signed out appropriately; (iii) negative samples would be sent on for confirmatory testing by toxigenic culture.…”

“…This sensitivity may be increased to Ͼ99% by combining CBA with toxigenic culture (i.e., C. difficile culture followed by CBA performed on the culture broth), but this increase comes at a cost of increased turnaround time (5,26). However, CBA is not standardized, requires tissue culture facilities, and has a turnaround time of at least 24 h to 4 days (26,31).…”

Diagnostic approach to Clostridium difficile infection

“…False negative results can occur in stored samples due to toxin degradation or by delay in transportation or by medication. In fact, a negative cytotoxicity assay does not completely rule out C. difficile as the cause of diarrhea as 30% of patients maybe missed [10].…”