Evaluation of real-time PCR and conventional diagnostic methods for the detection of Clostridium difficile-associated diarrhoea in a prospective multicentre study

Abstract

In this prospective multicentre study, an enzyme-linked fluorescent assay (VIDAS CDA2; bioMé rieux), an enzyme-linked assay [Premier Toxins A and B (PTAB); Meridian] and an in-house real-time PCR amplifying the tcdB gene were compared with the cell cytotoxicity assay used as the 'gold standard' for diagnosis of Clostridium difficile-associated diarrhoea (CDAD). Faecal samples from patients with a request for C. difficile diagnosis and samples from patients with diarrhoea hospitalized for at least 72 h were collected for 3 consecutive months from four university medical centres in The Netherlands. In total, 547 faecal samples were obtained from 450 patients. Of 540 samples available for all of the assays, 84 (15.6 %) showed a positive result in one or more assays. The cell cytotoxicity assay was positive in 31 samples (5.7 %) from 28 patients. A diagnosis of CDAD was not considered by the physician in 5 (23.8 %) of 21 patients with CDAD who were hospitalized for at least 72 h. Compared with the cell cytotoxicity assay, the sensitivity of VIDAS, PTAB and PCR was 83.9, 96.8 and 87.1 %, respectively. The specificity of VIDAS, PTAB and PCR was 97.1, 94.3 and 96.5 %, respectively. The positive and negative predictive values for VIDAS, PTAB and PCR were 63.4 and 99.0 %, 50.9 and 99.8 %, and 60.0 and 99.2 %, respectively. Of 61 samples that were positive in one, two or three of the assays, 56 were available for discordance analysis. Discordance analysis was performed by culture of toxinogenic strains. The concordance of VIDAS, PTAB and PCR with culture was 53.6 % (30/56), 55.4 % (31/56) and 71.4 % (40/56), respectively. It was concluded that real-time PCR had the highest concordance with toxinogenic culture and is therefore the preferred method for diagnosing CDAD in faecal samples. It was also concluded that diagnosis of patients with diarrhoea who have been hospitalized for more than 72 h should focus mainly on the detection of C. difficile, irrespective of the physician's request.

Clostridium difficile Infections in Children: Impact of the Diagnostic Method on Infection Rates

“…The increase in the number of cases cannot be explained solely by the difference in performance characteristics of the 2 tests: the sensitivity (68%-90% for EIA vs 88%-100% for PCR) and specificity (95.3%-99% for EIA vs 92.6%-98.4% for PCR) of the 2 tests are comparable. [12][13][14]26,27 The sudden increase in the incidence of CDI following the implementation of PCR could be partly explained by the fact that PCR detects toxin genes but cannot determine whether the organism is actively producing toxin. For that reason, some studies concluded that PCR is unreliable in differentiating CDI cases from asymptomatic carriers of a potentially toxigenic organism.…”

“…10,11 Several studies showed that PCR had an equivalent sensitivity and specificity (up to 100%) compared with toxigenic cultures. [12][13][14] However, PCR may not be useful when trying to differentiate carrier status from true CDI. 15 Moreover, given that clinical manifestations of CDI are milder in children compared with the adult population, 16 and given the current absence of testing strategies that accurately and optimally diagnose CDI, 17 we aimed to determine the proportion of pediatric patients diagnosed as having CDI by PCR who would also be diagnosed by EIA, and to compare the clinical characteristics of PCR + /enzyme-linked immunosorbent assay (ELISA) + vs PCR + /ELISA − patients.…”

High occurrence of variousClostridium difficilePCR ribotypes in pigs arriving at the slaughterhouse

“…All strains were further investigated by PCR ribotyping based upon Bidet et al (2000). Detection of toxin genes was done as described by van den Berg et al (2007).…”

Comparison of Two Commercial Molecular Assays to a Laboratory-Developed Molecular Assay for Diagnosis of Clostridium difficile Infection

“…Although some studies found that enzyme immunoassay (EIA) compared favorably to cytotoxicity assays (2, 9), several subsequent studies found EIA to have low sensitivity and specificity compared to those of both toxigenic culture and PCR (1,5,11). Also, some concluded that the performance of PCR is superior to that of other available methods (3,7,(10)(11)14). Although commercial FDA-approved assays have been evaluated compared to cytotoxicity assays and culture (4), our study is the first, to our knowledge, to compare two commercially available PCR methods to each other and to a laboratorydeveloped PCR assay with previously published performance characteristics.…”

“…Several laboratory-developed PCR assays for detection of C. difficile infection (CDI) have been described, and they have demonstrated favorable performances in comparative studies (3,7,10,14). We previously described a real-time PCR assay using a LightCycler (Roche Diagnostics, Indianapolis, IN), which we refer to herein as the LC-CDTX assay (11).…”