Comparison of Clostridioides difficile Stool Toxin Concentrations in Adults With Symptomatic Infection and Asymptomatic Carriage Using an Ultrasensitive Quantitative Immunoassay

Pollock, Nira R.; Banz, Alice; Chen, Xinhua; Williams, David N.; Xu, Hua; Cuddemi, Christine; Cui, Alice X; Perrotta, Matthew; Alhassan, Eaman; Riou, Brigitte; Lantz, Aude; Miller, Mark A.; Kelly, Ciarán P.

doi:10.1093/cid/ciy415

Cited by 62 publications

(72 citation statements)

References 25 publications

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“…Thus, ultrasensitive immunoassays can be used to further evaluate the potential of tcdB C T values to predict the presence of fecal toxin. C T values, at the proposed cutoffs, do not detect all samples with high toxin concentrations, not even those with very high concentrations (greater than the EIA and CCNA cutoffs) (10). Although there is a correlation between tcdB C T values and toxin concentration, the accuracy is suboptimal for use in clinical practice.…”

Section: Possible Ways Forwardmentioning

confidence: 84%

“…With the advent of quantitative ultrasensitive toxin immunoassays, which are capable of quantification at very low concentrations, from picogram per millimeter levels (11,15), an accurate assessment of toxin load can now be determined and the clinical value of using tcdB C T values to indirectly predict toxin can be further evaluated (9,11,15). In a recent study using PCR and an ultrasensitive toxin assay, multiple patients with C T values of Ͼ26.4 had detectable stool toxin, including values higher than analytical thresholds for EIA (ϳ1,000 pg/ml) and CCNA (TcdB of ϳ100 pg/ml) (10).…”

Section: Clinical Use Of C T Values Is Concerningmentioning

confidence: 99%

See 1 more Smart Citation

Ultrasensitive Detection of Clostridium difficile Toxins Reveals Suboptimal Accuracy of Toxin Gene Cycle Thresholds for Toxin Predictions

Sandlund¹,

Wilcox

2019

J Clin Microbiol

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The use of nucleic acid amplification tests (NAATs) for the diagnosis of Clostridium (Clostridioides) difficile infection (CDI) leads to overdiagnosis. To improve the clinical specificity of NAATs, there has been a recent interest in using toxin gene cycle thresholds (C T s) to predict the presence and absence of toxins. Although there is an association between C T values and fecal toxin concentrations, the predictive accuracy of the former is suboptimal for use in clinical practice. Ultrasensitive toxin immunoassays to quantify free toxins in stool offer a novel option for high-sensitivity fecal toxin detection rather than using surrogate markers for prediction.

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Section: Possible Ways Forwardmentioning

confidence: 84%

Section: Clinical Use Of C T Values Is Concerningmentioning

confidence: 99%

Ultrasensitive Detection of Clostridium difficile Toxins Reveals Suboptimal Accuracy of Toxin Gene Cycle Thresholds for Toxin Predictions

Sandlund¹,

Wilcox

2019

J Clin Microbiol

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“…An analytical cutoff for the Clarity assay was set at 12.0 pg/ml by comparing it statistically to CCCNA, the reference free-toxin test. Asymptomatic carriers can also have positive toxin assays (12), and CCCNA testing can lack sensitivity in clinically confirmed cases of CDI (21). Thus, future studies validating the clinical performance of the Clarity assay may provide additional insight into the role of ultrasensitive toxin detection.…”

Section: Discussionmentioning

confidence: 99%

“…The presence of toxins, not toxigenic C. difficile, in patients' stools correlates with disease severity (10)(11)(12), and current guidelines state that the detection of toxins is most relevant for CDI diagnosis (7,8). However, because of the poor sensitivity of currently available toxin EIAs, there is a need for a toxin test with high sensitivity that could be used as a stand-alone test (8).…”

mentioning

confidence: 99%

Ultrasensitive Detection of Clostridioides difficile Toxins in Stool by Use of Single-Molecule Counting Technology: Comparison with Detection of Free Toxin by Cell Culture Cytotoxicity Neutralization Assay

Hansen

Young

et al. 2019

J Clin Microbiol

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Laboratory tests for Clostridioides difficile infection (CDI) rely on the detection of free toxin or molecular detection of toxin genes. The Singulex Clarity C. diff toxins A/B assay is a rapid, automated, and ultrasensitive assay that detects C. difficile toxins A and B in stool. We compared CDI assays across two prospective multicenter studies to set a cutoff for the Clarity assay and to independently validate the performance compared with that of a cell culture cytotoxicity neutralization assay (CCCNA). The cutoff was set by two sites testing fresh samples from 897 subjects with suspected CDI and then validated at four sites testing fresh samples from 1,005 subjects with suspected CDI. CCCNA testing was performed at a centralized laboratory. Samples with discrepant results between the Clarity assay and CCCNA were retested with CCCNA when the Clarity result agreed with that of at least one comparator method; toxin enzyme immunoassays (EIA), glutamate dehydrogenase (GDH) detection, and PCR were performed on all samples. The cutoff for the Clarity assay was set at 12.0 pg/ml. Compared to results with CCCNA, the Clarity assay initially had 85.2% positive agreement and 92.4% negative agreement. However, when samples with discrepant results between the Clarity assay and CCCNA in the validation study were retested by CCCNA, 13/17 (76.5%) Clarity-negative but CCCNA-positive samples (Clarity+/CCCNA−) became CCCNA−, and 5/26 (19.2%) Clarity+/CCCNA− samples became CCCNA+, resulting in a 96.3% positive agreement and 93.0% negative agreement between Clarity and CCCNA results. The toxin EIA had 59.8% positive agreement with CCCNA. The Clarity assay was the most sensitive free-toxin immunoassay, capable of providing CDI diagnosis in a single-step solution. A different CCCNA result was reported for 42% of retested samples, increasing the positive agreement between Clarity and CCCNA from 85.2% to 96.3% and indicating the challenges of comparing free-toxin results to CCCNA results as a reference standard.

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“…However, the diagnosis of CDI in hospitals is confounded by multiple factors. First, five percent of the general population and more than twenty percent of the hospitalized population are asymptomatic carriers (8-11). Second, even among hospitalized patients, diarrhea is caused by CDI in only 10-20% of cases, despite concurrent leukocytosis or immune compromise (12, 13).…”

Section: Introductionmentioning

confidence: 99%

Novel Immunoassay for Diagnosis of OngoingClostridioides difficileInfections Using Serum and Medium Enriched for Newly Synthesized Antibodies (MENSA)

Haddad¹,

Nozick²,

Kim³

et al. 2020

Preprint

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47BACKGROUND. Clostridioides difficile infections (CDI) have been a challenging and 48 increasing serious concern in recent years. While early and accurate diagnosis is crucial, available assays 49 have frustrating limitations 50 51 OBJECTIVE. Develop a simple, blood-based immunoassay to accurately diagnose patients 52 suffering from active CDI. 53 54 MATERIALS AND METHODS. Uninfected controls (n=95) and CDI patients (n=167) were 55 recruited from Atlanta area hospitals. Blood samples were collected from patients within twelve days of a 56 positive CDI test and processed to yield serum and PBMCs cultured to yield medium enriched for newly 57 synthesized antibodies (MENSA). Multiplex immunoassays measured Ig responses to ten recombinant C. 58 difficile antigens. 59 60 RESULTS. Sixty-six percent of CDI patients produced measurable responses to C. difficile 61 antigens in their serum or MENSA within twelve days of a positive CDI test. Fifty-two of the 167 CDI 62 patients (31%) were detectable in both serum and MENSA, but 32/167 (19%) were detectable only in 63 MENSA, and 27/167 (16%) were detectable only in serum. 64 65 DISCUSSION. We describe the results of a multiplex immunoassay for the diagnosis of ongoing 66 CDI in hospitalized patients. Our assay resolved patients into four categories: MENSA-positive only, 67 serum-positive only, MENSA-and serum-positive, and MENSA-and serum-negative. The MENSA 68 positive-only patients accounted for 30% and may be attributed to nascent antibody secretion in MENSA 69 prior to seroconversion. Conversely, the serum positive-only subset may have been more advanced in 70 5 5 their disease course. Immunocompromise and misdiagnosis may have contributed to the 34% of CDI 71 patients who were not identified using MENSA or serum immunoassays. 72 73 IMPORTANCE. While there was considerable overlap between patients identified through 74 MENSA and serum, both methods detected additional, unique patients. The combined use of both 75 MENSA and serum to detect CDI patients resulted in the greatest identification of CDI patients. Together, 76 longitudinal analysis of MENSA and serum will provide a more accurate evaluation of successful host 77 humoral immune responses in CDI patients. 78In this observational investigation, we present the results of a pilot study validating the use of a 127 multi-antigen immunoassay and its ability to measure antibody secretion reflecting ongoing CDI in 128 MENSA samples from a complex hospital population and compare its performance with results from sera. 129 130 RESULTS 131

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Comparison of Clostridioides difficile Stool Toxin Concentrations in Adults With Symptomatic Infection and Asymptomatic Carriage Using an Ultrasensitive Quantitative Immunoassay

Abstract: Toxin concentration did not differentiate an individual with CDI from one with asymptomatic carriage. Median stool toxin concentrations in groups with CDI versus carriage differed, but only when groups were defined by detectable stool toxin (versus positive NAAT).

Cited by 62 publications

References 25 publications

Ultrasensitive Detection of Clostridium difficile Toxins Reveals Suboptimal Accuracy of Toxin Gene Cycle Thresholds for Toxin Predictions

Ultrasensitive Detection of Clostridium difficile Toxins Reveals Suboptimal Accuracy of Toxin Gene Cycle Thresholds for Toxin Predictions

Ultrasensitive Detection of Clostridioides difficile Toxins in Stool by Use of Single-Molecule Counting Technology: Comparison with Detection of Free Toxin by Cell Culture Cytotoxicity Neutralization Assay

Novel Immunoassay for Diagnosis of OngoingClostridioides difficileInfections Using Serum and Medium Enriched for Newly Synthesized Antibodies (MENSA)

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