Clostridium difficile Toxin, Molecular Detection, PCR, Feces
Sensitive, specific, and rapid diagnosis of Clostridium difficile-associated diarrhea and pseudomembranous colitis
Testing Algorithm Delineates situation(s) when tests are added to the initial order. This includes reflex and additional tests.
See Laboratory Testing for Infectious Causes of Diarrhea in Special Instructions.
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
Clostridium difficile is the cause of Clostridium difficile-associated diarrhea (CDAD), an antibiotic-associated diarrhea, and pseudomembranous colitis (PMC). In these disorders bacterial overgrowth of Clostridium difficile develops in the colon, typically as a consequence of antibiotic usage. Clindamycin and broad-spectrum cephalosporins have been most frequently associated with CDAD and PMC, but almost all antimicrobials may be responsible. Disease is related to production of toxin A and/or B. Treatment typically involves withdrawal of the associated antimicrobials and, if symptoms persist, orally administered and intraluminally active metronidazole, vancomycin, or fidaxomicin. Intravenous metronidazole may be used if an oral agent cannot be administered. In recent years, a more severe form of CDAD with increased morbidity and mortality has been recognized as being caused by an epidemic toxin-hyperproducing strain of Clostridium difficile (NAP1 strain). Many toxin-hyperproducing isolates also contain the binary toxin gene and are resistant quinolones. This test does not differentiate between toxin-hyperproducing and nontoxin-hyperproducing strains.
Traditionally, diagnosis relied upon 1) clinical and epidemiologic features, 2) culture (which is labor intensive and time consuming), 3) cytotoxicity assays, which are labor intensive and time consuming, and 4) toxin detection immunoassays (which are insensitive). The described PCR assay detects the regulatory gene (tcdC) responsible for production of toxins A and B. This test is used for rapid diagnosis of CDAD and PMC enabling prompt treatment that may reduce hospital stays for inpatients with CDAD.
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
A positive PCR result for the presence of the gene regulating toxin production (tcdC) indicates the presence of Clostridium difficile and toxin A and/or B.
A negative result indicates the absence of detectable Clostridium difficile tcdC DNA in the specimen, but does not rule-out Clostridium difficile infection. False-negative results may occur due to inhibition of PCR, sequence variability underlying the primers or probes, or the presence of Clostridium difficile in quantities less than the limit of detection of the assay.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
The assay must be performed on fresh stools, fresh-frozen stools, or stools in transport medium.
The assay has not been validated as a test of cure. Since nucleic acid may persist after effective treatment, follow-up testing of a positive result is not recommended.
Interfering substances in the stool may affect the accuracy of the assay; results should always be interpreted in conjunction with clinical and epidemiologic findings.
Submission of more than 1 specimen for testing is not recommended.
Testing of colostomy, ileostomy, or colonoscopically collected specimens has not been validated.
Patients may asymptomatically carry Clostridium difficile; clinical correlation is needed when deciding how to manage patients with a positive test result.
Results of the PCR assay were compared with those of Clostridium difficile toxin-detecting EIAs and culture of Clostridium difficile. Two hundred stool specimens were studied in a blinded manner. Clostridium difficile was isolated from 49 specimens by culture and 44 of these were confirmed as containing 1 of the genes associated with toxin production (toxigenic culture). Using toxigenic culture as the "gold standard," the sensitivities and specificities, respectively, of the assays were 48% and 98% for the Premier Toxin A/B EIA (Meridian diagnostics); 48% and 99% for the ImmunoCard toxin A and B test (Meridian); 48% and 84% for the Xpect Clostridium difficile toxin A/B test (Remel); 32% and 100% for the Triage Clostridium difficile panel (for toxin A, Biosite Diagnostics); and 86% and 97% for the PCR assay. No cross-reactivity was observed in the PCR assay with a panel of 51 pathogens and normal flora, including other Clostridium species. The analytical sensitivity of the PCR assay was 7 copies/mcL of extracted stool.
Clinical Reference Provides recommendations for further in-depth reading of a clinical nature
1. Aichinger E, Schleck CD, Harmsen WS, et al: Nonutility of repeat laboratory testing for detection of Clostridium difficile by use of PCR or enzyme immunoassay. J Clin Microbiol 2008;46:3795-3797
2. Sloan LM, Duresko BJ, Gustafson DR, et al: Comparison of real-time PCR for detection of the tcdC gene with four toxin immunoassays and culture in diagnosis of Clostridium difficile infection. J Clin Microbiol 2008;46:1996-2001
3. Verdoorn BP, Orenstein R, Rosenblatt JE, et al: High prevalence of tcdC deletion-carrying Clostridium difficile and lack of association with disease severity. Diagn Microbiol Infect Dis 2010;66:24-28
4. Karre T, Sloan L, Patel R, et al: Comparison of two commercial molecular assays to a laboratory-developed molecular assay for diagnosis of Clostridium difficile infection. J Clin Microbiol 2011;49:725-727