Evaluation of a rapid enzyme immunoassay for the detection ofClostridium difficile in stools

Barbut, Frédéric; Revel, R.; Ephraim, Richard K.D.; Leluan, P.; Lureau, P.; Petit, Jean‐Claude

doi:10.1007/bf01974552

Cited by 7 publications

(3 citation statements)

References 9 publications

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“…Current methods for diagnosing C. difficile infection are based on detection of the organism or the toxin genes and proteins or on the effect of the cytotoxin on tissue culture cells (2,(10)(11)(12). The only method that can provide information about the activities of the toxins is the cell cytotoxicity assay.…”

Section: Discussionmentioning

confidence: 99%

Harnessing the Glucosyltransferase Activities of Clostridium difficile for Functional Studies of Toxins A and B

Darkoh¹,

Kaplan²,

DuPont

2011

J Clin Microbiol

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The incidence of Clostridium difficile infection (CDI) has been increasing within the last decade. Pathogenic strains of C. difficile produce toxin A and/or toxin B, which are important virulence factors in the pathogenesis of this bacterium. Current methods for diagnosing CDI are mostly qualitative tests that detect either the bacterium or the toxins. We have developed an assay (Cdifftox activity assay) to detect C. difficile toxin A and B activities that is quantitative and cost-efficient and utilizes a substrate that is stereochemically similar to the native substrate of the toxins (UDP-glucose). To characterize toxin activity, toxins A and B were purified from culture supernatants by ammonium sulfate precipitation and chromatography through DEAE-Sepharose and gel filtration columns. The activities of the final fractions were quantitated using the Cdifftox activity assay and compared to the results of a toxin A-and B-specific enzyme-linked immunosorbent assay (ELISA). The affinity for the substrate was >4-fold higher for toxin B than for toxin A. Moreover, the rate of cleavage of the substrate was 4.3-fold higher for toxin B than for toxin A. The optimum temperature for both toxins ranged from 35 to 40°C at pH 8. Culture supernatants from clinical isolates obtained from the stools of patients suspected to be suffering from CDI were tested using the Cdifftox activity assay, and the results were compared to those of ELISA and PCR amplification of the toxin genes. Our results demonstrate that this new assay is comparable to the current commercial ELISA for detecting the toxins in the samples tested and has the added advantage of quantitating toxin activity.

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Section: Discussionmentioning

confidence: 99%

Harnessing the Glucosyltransferase Activities of Clostridium difficile for Functional Studies of Toxins A and B

Darkoh¹,

Kaplan²,

DuPont

2011

J Clin Microbiol

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“…However, this test has never become commercially available. More recently, we evaluated a very rapid enzyme immunoassay for the detection of C. difficile ‐specific antigen and we concluded that the good negative predictive value of this test made it particularly suitable for the rapid screening of specimens [22]. However, all positive results had to be confirmed by toxin detection before a definitive diagnosis of CDAD could be established.…”

Section: Discussionmentioning

confidence: 99%

Rapid detection of Clostridium difficile toxin A in stool specimens

Barbut

Macé

Lalande

et al. 1997

Clinical Microbiology and Infection

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OBJECTIVE: To evaluate a rapid (15-min) enzyme immunoassay in the format of an individual cassette (ImmunoCard toxin A, Meridian, BMD, Marne-la-Vallée, France) for the detection of Clostridium difficile toxin A in stool specimens. METHODS: We compared this new test with the cytotoxicity assay using MRC-5 cells, the ToxA test (TechLab, BioWhittaker, Fontenay-sous-bois, France) and toxigenic culture for the diagnosis of C. difficile-associated diseases (CDAD). A total of 236 stool specimens collected from 220 patients was simultaneously tested with the four methods. Discordant results were resolved by reviewing patients' clinical records. RESULTS: The prevalence of CDAD was 13.9%. Test sensitivities and specificities were 100% and 99% respectively for the cytotoxicity assay, 87.5% and 100% for ImmunoCard toxin A, 77.4% and 100% for the ToxA test and 100% and 98% for toxigenic culture. CONCLUSIONS: The ImmunoCard Toxin A is a very rapid, individual and easy-to-perform test for the diagnosis of CDAD. It provides same-day results and may be useful for both guiding appropriate treatment and controlling nosocomial spread of C. difficile.

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“…These tests include faecal culture on selective media, detection of a nonspecific antigen (glutamate dehydrogenase, GDH) and direct detection of toxins A and/or B from stools. This latter method can be performed either by the cytotoxicity assay or by a direct enzyme immunoassay (EIA) [11–22]. The cytotoxicity assay is considered by many authors as the standard because of its great sensitivity for the detection of toxin B [23].…”

Section: Introductionmentioning

confidence: 99%

A European survey of diagnostic methods and testing protocols for Clostridium difficile

Barbut

Delmée

Brazier

et al. 2003

Clinical Microbiology and Infection

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106

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The results of this study suggest marked discrepancies between laboratories and also between countries regarding the criteria by which C. difficile is investigated for, and the methods and the strategies that are used for the diagnosis of C. difficile. These discrepancies could be explained by the lack of clear guidelines for C. difficile diagnosis in each country, and by the importance that physicians attach to C. difficile. Precise guidelines for C. difficile diagnosis would be the first step to make possible accurate comparison of the incidence and the epidemiology of CDAD from one hospital to another or from one country to another.

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Evaluation of a rapid enzyme immunoassay for the detection ofClostridium difficile in stools

Cited by 7 publications

References 9 publications

Harnessing the Glucosyltransferase Activities of Clostridium difficile for Functional Studies of Toxins A and B

Harnessing the Glucosyltransferase Activities of Clostridium difficile for Functional Studies of Toxins A and B

Rapid detection of Clostridium difficile toxin A in stool specimens

A European survey of diagnostic methods and testing protocols for Clostridium difficile

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