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Rapid Simultaneous Molecular Stool-Based Detection of Toxigenic Clostridioides difficile by Quantitative TaqMan Real-Time PCR Assay

Kouhsari, E. and Douraghi, M. and Barati, M. and Yaseri, H.F. and Talebi, M. and Abbasian, S. and Moqarabzadeh, V. and Amirmozafari, N. (2019) Rapid Simultaneous Molecular Stool-Based Detection of Toxigenic Clostridioides difficile by Quantitative TaqMan Real-Time PCR Assay. Clinical Laboratory, 65 (4). pp. 461-469.

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Abstract

Background: Clostridioides difficile is a major cause of nosocomial infectious diarrhea in hospitalized patients throughout the world. Methods: A multiplex real-time PCR assay was developed and evaluated in comparison with toxigenic culture (TC) (as gold standard method) for direct detection of toxigenic C. difficile in fecal specimens. The multiplex real-time PCR assay simultaneously detected glutamate dehydrogenase (gluD), toxin A (tcdA), toxin B (tcdB), and binary toxin (cdtB) genes in stool samples. Results: The results of multiplex real-time PCR were compared to those of the TC method in 250 patients suspected of C. difficile infection. The prevalence of positive TC was 13.6. Forty-two stool samples (16.8) were determined to be gluD+ using multiplex real-time PCR. These included 35 (83.3) toxigenic (32 tcdA+, tcdB+ and three tcdB+) and 7 (20.0) were cdtB+. The multiplex real-time PCR assay had a sensitivity of 91.45, specificity of 99.54, and positive and negative predictive values of 97 and 98.6, respectively, compared to the TC method for diagnosis of C. difficile. The analytical sensitivity of the multiplex real-time PCR assay was estimated to be 102 CFU/g of stools and 0.0200 pg of genomic DNA from culture. The analytical specificity was determined to be 100 by using enteric and non-C. difficile standard bacterial strains. Conclusions: The molecular method developed in the study was rapid, sensitive, and specific for detection of toxigenic C. difficile. It is applicable to be performed in clinical laboratories and correlated well with the results obtained by TC. © 2019 Verlag Klinisches Labor GmbH. All rights reserved.

Item Type: Article
Additional Information: cited By 2
Uncontrolled Keywords: bacterial DNA; bacterial toxin; genomic DNA; glutamate dehydrogenase; bacterial protein; bacterial toxin; enterotoxin, adult; aged; amplicon; Article; bacterial gene; bacterial strain; bacterium culture; cdtA gene; cdtB gene; Clostridium difficile infection; controlled clinical trial; controlled study; diagnostic test accuracy study; feces analysis; female; genetic profile; genetic variability; human; limit of detection; major clinical study; male; molecular diagnosis; multiplex polymerase chain reaction; predictive value; quantitative assay; real time polymerase chain reaction; sensitivity and specificity; tcdA gene; tcdB gene; chemistry; Clostridioides difficile; diarrhea; feces; genetics; isolation and purification; laboratory technique; microbiology; middle aged; multiplex polymerase chain reaction; procedures; pseudomembranous colitis; real time polymerase chain reaction; receiver operating characteristic; very elderly, Aged; Aged, 80 and over; Bacterial Proteins; Bacterial Toxins; Clinical Laboratory Techniques; Clostridium difficile; Diarrhea; Enterocolitis, Pseudomembranous; Enterotoxins; Feces; Female; Humans; Male; Middle Aged; Multiplex Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction; ROC Curve; Sensitivity and Specificity
Subjects: QU بیوشیمی
فارماکولوژی QV
> دانشکده داروسازی > فارماکولوژی QV

آسیب شناسی بالینی QY
میکروب شناسی وایمنی شناسی QW
مقالات نمایه شده محققین دانشگاه در سایت ,Web of Science ,Scopus
Divisions: معاونت تحقیقات و فناوری
Depositing User: GOUMS
Date Deposited: 18 Dec 2019 11:36
Last Modified: 18 Dec 2019 11:36
URI: http://eprints.goums.ac.ir/id/eprint/10456

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