Bacteriology
Performance of TechLab C. DIFF QUIK CHEK™ and TechLab C. DIFFICILE TOX A/B II™ for the detection of Clostridium difficile in stool samples

https://doi.org/10.1016/j.diagmicrobio.2007.04.018Get rights and content

Abstract

Two membrane-bound enzyme immunoassays by TechLab, Blacksburg, VA, were evaluated and compared with the Triage® Micro C. difficile Panel (Biosite Diagnostics, San Diego, CA), with culture, and with cytotoxic assay. The TechLab panels were C. DIFF QUIK CHEK™ (QC-GDH) and C. DIFFICILE TOX A/B II™ (QC-toxinA/B), which detect glutamate dehydrogenase (GDH) and Clostridium difficile toxins A and B, respectively. The Triage Panel detects GDH (TR-GDH) and toxin A (TR-toxinA).

Methods

Stool samples were inoculated onto CCFA plates (Q-Labs, Quebec, Canada) after alcohol shock, and suspected colonies were identified by the MicroScreen C. difficile latex slide agglutination test (Microgen Bioproducts, Surrey, UK). TR-GDH, TR-toxinA, QC-GDH, and QC-toxinA/B tests were performed according to the manufacturers' instructions on all the samples. Samples positive for GDH or culture but negative for TR-toxinA and QC-toxinA/B were further tested by cytotoxin assay (CTA). CTA was also performed on samples that caused blackening of the Triage® Micro C. difficile Panel.

Results

A total of 313 of 401 stool samples were negative for GDH and toxins (78%). Eighty-eight samples were positive either for GDH or culture or both. Thirteen of these could not be evaluated for C. difficile-associated diarrhea (CDAD) because CTA test was not performed. Toxin/s was detected at least by one method in 46 (11.8%) of 388 samples that were positive for culture or GDH and were considered diagnostic of CDAD. The QC-GDH was more sensitive than culture and TR-GDH for the detection of C. difficile. However, in 18GDH-positive samples positive for either of the Triage or TechLab immunoassays, the culture remained negative. Ten (2%) results of the Triage immunoassays could not be evaluated because of discoloration of the panels. QC-GDH (93.5%) was more sensitive for detecting the presence of toxin-producing C. difficile than TR-GDH (79.5%). TR-toxinA was more specific for detecting the presence of toxin-producing C. difficile than QC-toxinA/B (100% and 96.9%, respectively).

Conclusions

The GDH tests had a faster turnaround time than the traditional culture methods. QC-GDH was most sensitive for the detection C. difficile-positive stools and was easy to use.

Introduction

Clostridium difficile was first identified as the primary cause of pseudomembranous colitis in 1978. Since then, the organism is recognized as the leading cause of hospital-acquired diarrhea called C. difficile-associated diarrhea (CDAD).

In the United States alone, the estimated number of cases of CDAD exceeds 250 000 per year, causing an economic burden of over $1 billion to the health care system annually (Wilkins and Lyerle, 2003, Kyne et al., 2002). Previously, reports of C. difficile outbreaks have been restricted to a single hospital. Recently, however, there appears to be a wider distribution with increased mortality. Consequently, in some parts, the disease has become a serious public health concern. In one Canadian hospital, the incidence of C. difficile cases increased from 35.6 per 100 000 population in 1991 to 156.3 per 100 000 in 2003. In the same period, the proportion of complicated cases increased from 7.1% to 18.2% and 30 days mortality rate from 4.7% to 13.8% (Pėpin et al., 2004).

The diagnosis of C. difficile is based upon the clinical history of recent antibiotic use, diarrhea, and laboratory tests for C. difficile markers including isolation of C. difficile, determination of the presence of glutamate dehydrogenase (GDH), and toxins. TechLab, Blacksburg, VA, has recently marketed 2 membrane-bound enzyme immunoassays (EIAs), C. DIFF Quick CHEK (QC-GDH) and TOX A/B Quick CHEK (QC-toxinA/B), that detect GDH and toxins A/B, respectively.

The aim of this study was to evaluate and compare the 2TechLab assays (QC-toxinA/B and QC-GDH) with the Triage® C. difficile Panel membrane-bound immunoassay (Biosite Diagnostics, San Diego, CA), the culture isolation ofC. difficile from stool samples, and an in-house cytotoxin assay (CTA). The Triage® C. difficile Panel assay containsdetection zones for GDH (TR-GDH) and toxin A (TR-toxinA).

Section snippets

Study protocol

All nonformed stools submitted from inpatients suspected of CDAD were included in the study. All tests (culture isolation, QC-toxinA/B, QC-GDH, TR-GDH, and TR-toxinA) were performed within 24 h of the arrival of specimens in the laboratory. A portion of the stool was frozen at −70 °C for further investigation if required. Samples positive for GDH or culture but negative for the presence of toxin/s by TR-toxinA and QC-toxinA/B were further tested by CTA. Also, samples on Triage panels that were

Statistical analysis

For the purpose of this study, the gold standard was considered a combination of several test results indicating a diagnostic positive for toxigenic C. difficile. A sample that was positive for either of the 2 screening methods (culture or GDH) and positive for toxin (toxin A, toxin A/B, or cytotoxic assay) was considered to be true positive for toxigenic C. difficile. This gold standard was used to calculate the sensitivities and specificities of the membrane-bound immunoassay tests. The

Assay results

In total, 401 specimens were processed during the study. All specimens were tested by the TechLab and Triage immunoassays and culture. Ten (2.5%) of 401 specimens could not be evaluated by the Triage® C. difficile panels because of discoloration of panels. Cytotoxic assay was performed on 54 specimens. Thirteen samples were excluded when analyzing performance of toxin assays because CTA was not performed in the face of positive culture or GDH but negative toxin immunoassay. For most of these

Discussion

At present, there is no sufficiently discriminatory or specific screening test for the diagnosis of CDAD; consequently, the Society for Healthcare Epidemiology of America has recommended that culture as well as a cytotoxic assay should be performed on stool specimens submitted for determination of CDAD (Gerding et al., 1995). The approach is problematic because both culture and CTA have slow turnaround times and require a minimum of 2 days to yield results.

Although CTA is the most sensitive

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