Comparison of Strain Typing Results for Clostridium difficile Isolates from North America

Tenover, Fred C.; Åkerlund, Thomas; Gerding, Dale N.; Goering, Richard V.; Boström, Therése; Jönsson, Anna Maria; Wong, Edith Y.; Wortman, Alan T.; Persing, David H.

doi:10.1128/jcm.02446-10

Cited by 89 publications

(56 citation statements)

References 47 publications

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“…All 027/NAP1/BI strains identified were positive for all three markers ( Table 3). The positive results for 027NAP1/BI by the Xpert C. difficile/Epi were confirmed by PFGE (25) and/or sequencing of the tcdC gene (22). All nine strains identified were positive for all three markers ( Table 3).…”

Section: Discussionmentioning

confidence: 79%

“…The 027/ NAP1/BI strain is responsible for widespread outbreaks of C. difficile in North America (13,14,27). The hypervirulent strains have been reported to exhibit increased sporulation capacity, along with high levels of toxin production (1,25). The significance of 027/NAP1/BI as an epidemiological marker is known, and outbreaks caused by a toxin variant epidemic strain have renewed interest in detecting this strain.…”

Section: Discussionmentioning

confidence: 99%

“…The 027/NAP1/BI positive result by the Xpert C. difficile/Epi assay was confirmed by toxinotyping PFGE (25) and/or by sequencing (22), which identified the isolate as the epidemic strain 027/NAP1/BI (toxinotype III; binary toxin positive; 18-bp tcdC deletion).…”

Section: Specimensmentioning

confidence: 96%

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Detection of Toxigenic Clostridium difficile: Comparison of the Cell Culture Neutralization, Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile Assays

et al. 2012

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Clostridium difficile is the most important cause of nosocomial diarrhea. Several laboratory techniques are available to detect C. difficile toxins or the genes that encode them in fecal samples. We evaluated the Xpert C. difficile and Xpert C. difficile/Epi (Cepheid, CA) that detect the toxin B gene (tcdB) and tcdB, cdt, and a deletion in tcdC associated with the 027/NAP1/BI strain, respectively, by real-time PCR, and the Illumigene C. difficile (Meridian Bioscience, Inc.) that detects the toxin A gene (tcdA) by loop-mediated isothermal amplification in stool specimens. Toxigenic culture was used as the reference method for discrepant stool specimens. Two hundred prospective and fifty retrospective diarrheal stool specimens were tested simultaneously by the cell cytotoxin neutralization assay (CCNA) and the Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile assays. Of the 200 prospective stools tested, 10.5% (n ‫؍‬ 23) were determined to be positive by CCNA, 17.5% (n ‫؍‬ 35) were determined to be positive by Illumigene C. difficile, and 21.5% (n ‫؍‬ 43) were determined to be positive by Xpert C. difficile and Xpert C. difficile/ Epi. Of the 50 retrospective stools, previously determined to be positive by CCNA, 94% (n ‫؍‬ 47) were determined to be positive by Illumigene C. difficile and 100% (n ‫؍‬ 50) were determined to be positive by Xpert C. difficile and Xpert C. difficile/Epi. Of the 11 discrepant results (i.e., negative by Illumigene C. difficile but positive by Xpert C. difficile and Xpert C. difficile/Epi), all were determined to be positive by the toxigenic culture. A total of 21% of the isolates were presumptively identified by the Xpert C. difficile/Epi as the 027/NAP1/BI strain. The Xpert C. difficile and Xpert C. difficile/Epi assays were the most sensitive, rapid, and easy-to use assays for the detection of toxigenic C. difficile in stool specimens.

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Section: Discussionmentioning

confidence: 79%

Section: Discussionmentioning

confidence: 99%

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Detection of Toxigenic Clostridium difficile: Comparison of the Cell Culture Neutralization, Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile Assays

et al. 2012

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“…Toxigenic culture was performed using standard well-established methods (35) in D.N.G. 's laboratory.…”

Section: Methodsmentioning

confidence: 99%

Development and Validation of Digital Enzyme-Linked Immunosorbent Assays for Ultrasensitive Detection and Quantification of Clostridium difficile Toxins in Stool

et al. 2015

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The currently available diagnostics for Clostridium difficile infection (CDI) have major limitations. Despite mounting evidence that toxin detection is paramount for diagnosis, conventional toxin immunoassays are insufficiently sensitive and cytotoxicity assays too complex; assays that detect toxigenic organisms (toxigenic culture [TC] and nucleic acid amplification testing [NAAT]) are confounded by asymptomatic colonization by toxigenic C. difficile. We developed ultrasensitive digital enzymelinked immunosorbent assays (ELISAs) for toxins A and B using single-molecule array technology and validated the assays using (i) culture filtrates from a panel of clinical C. difficile isolates and (ii) 149 adult stool specimens already tested routinely by NAAT. The digital ELISAs detected toxins A and B in stool with limits of detection of 0.45 and 1.5 pg/ml, respectively, quantified toxins across a 4-log range, and detected toxins from all clinical strains studied. Using specimens that were negative by cytotoxicity assay/TC/NAAT, clinical cutoffs were set at 29.4 pg/ml (toxin A) and 23.3 pg/ml (toxin B); the resulting clinical specificities were 96% and 98%, respectively. The toxin B digital ELISA was 100% sensitive versus cytotoxicity assay. Twenty-five percent and 22% of the samples positive by NAAT and TC, respectively, were negative by the toxin B digital ELISA, consistent with the presence of organism but minimal or no toxin. The mean toxin levels by digital ELISA were 1.5-to 1.7-fold higher in five patients with CDI-attributable severe outcomes, versus 68 patients without, but this difference was not statistically significant. Ultrasensitive digital ELISAs for the detection and quantification of toxins A and B in stool can provide a rapid and simple tool for the diagnosis of CDI with both high analytical sensitivity and high clinical specificity.T he recent increases in the global incidence and severity of Clostridium difficile infection (CDI) (1-3) are of major concern, and CDI now consumes substantial resources for diagnosis, treatment, and infection control (2, 4). A recent U.S. prevalence survey of health care-associated infections (HAI) (5) found that C. difficile was the most commonly reported pathogen, causing 12.1% of HAI. Despite available therapies, treatment failure and recurrence are common (1, 2, 6). The emergence of epidemic strains capable of toxin hyperproduction and increased disease severity/mortality (2, 7, 8) has further increased the urgency to improve methods for diagnosis and treatment. Accurate diagnosis remains the cornerstone of effective management.Disease caused by C. difficile infection is due to the effects of two large protein exotoxins, toxins A and B (9, 10). While the presence of toxin is necessary to cause disease, the optimal method for diagnosing CDI remains controversial. Appropriate patient selection can improve diagnostic accuracy for all assays (2, 11). The classic gold standard stool assay, toxigenic culture (TC), is complex, lengthy, and unstandardized; furthermore, it is...

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“…These strains were chosen based on their impact in global outbreaks (51). Lastly, the effect of the S.b-conditioned medium in cytotoxicity caused by C. difficile filtered supernatants in mouse fibroblasts was also evaluated.…”

Section: By Utilizing a Well-established Hamster Model Of CDI We Showmentioning

confidence: 99%

ProbioticSaccharomyces boulardiiCNCM I-745 prevents outbreak-associatedClostridium difficile-associated cecal inflammation in hamsters

Koon

et al. 2016

American Journal of Physiology-Gastrointestinal and Liver Physi

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. difficile infection (CDI) is a common debilitating nosocomial infection associated with high mortality. Several CDI outbreaks have been attributed to ribotypes 027, 017, and 078. Clinical and experimental evidence indicates that the nonpathogenic yeast Saccharomyces boulardii CNCM I-745 (S.b) is effective for the prevention of CDI. However, there is no current evidence suggesting this probiotic can protect from CDI caused by outbreak-associated strains. We used established hamster models infected with outbreak-associated C. difficile strains to determine whether oral administration of live or heat-inactivated S.b can prevent cecal tissue damage and inflammation. Hamsters infected with C. difficile strain VPI10463 (ribotype 087) and outbreakassociated strains ribotype 017, 027, and 078 developed severe cecal inflammation with mucosal damage, neutrophil infiltration, edema, increased NF-B phosphorylation, and increased proinflammatory cytokine TNF␣ protein expression. Oral gavage of live, but not heated, S.b starting 5 days before C. difficile infection significantly reduced cecal tissue damage, NF-B phosphorylation, and TNF␣ protein expression caused by infection with all strains. Moreover, S.b-conditioned medium reduced cell rounding caused by filtered supernatants from all C. difficile strains. S.b-conditioned medium also inhibited toxin A-and B-mediated actin cytoskeleton disruption. S.b is effective in preventing C. difficile infection by outbreak-associated via inhibition of the cytotoxic effects of C. difficile toxins. (46). C. difficile is an anaerobic bacterium that produces two toxins -toxin A and toxin B -that mediate diarrhea, inflammation, and apoptosis of the mucosal epithelium in animals and humans (32). The effects of the toxin in target intestinal cells involve inactivation of the Rho family of GTPase, leading to cytoskeletal disorganization, epithelial cell apoptosis, and ultimately cell death (42, 54). C. difficile toxins also stimulate transcription of several proinflammatory genes, including tumor necrosis factor-␣ (TNF␣) (19) and activate transcription factors and mitogen-activated protein kinases involved in their proinflammatory effects (18,25). The mainstream CDI regimen includes the use of metronidazole, vancomycin, and fidaxomicin (55). However, many C. difficileinfected patients may also suffer from recurrent infections (13), while the recent epidemics that also involve new epidemic outbreak-associated strains (29) pose a major medical problem and epidemiological concern. These challenges have been met with a broad range of preventive approaches against CDI, including the use of probiotics, most notably Saccharomyces boulardii (S.b) alone or in combination with established antibiotic treatment (23,24).S.b is a nonpathogenic yeast that represents one of the well-studied probiotics against CDI in both experimental and clinical settings (24,40). Randomized double-blind placebocontrolled clinical trials have shown that S.b CNCM I-745 is an effective probiotic in the prophylaxis of antibi...

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Comparison of Strain Typing Results for Clostridium difficile Isolates from North America

Cited by 89 publications

References 47 publications

Detection of Toxigenic Clostridium difficile: Comparison of the Cell Culture Neutralization, Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile Assays

Detection of Toxigenic Clostridium difficile: Comparison of the Cell Culture Neutralization, Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile Assays

Development and Validation of Digital Enzyme-Linked Immunosorbent Assays for Ultrasensitive Detection and Quantification of Clostridium difficile Toxins in Stool

ProbioticSaccharomyces boulardiiCNCM I-745 prevents outbreak-associatedClostridium difficile-associated cecal inflammation in hamsters

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