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Frontiers of Medicine >> 2018, Volume 12, Issue 2 doi: 10.1007/s11684-017-0560-5

Simultaneous detection and characterization of toxigenic:Clostridium difficile directly from clinical stool specimens

  1. The First People’s Hospital of Xiaoshan District, Hangzhou 311021, China
  2. Department of Microbiology
  3. Department of Disease Control and Prevention, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051, China
  4. Biotherapy Center for Medical Oncology, the First Affiliated Hospital, Zhejiang University, Hangzhou 310003, China
  5. Department of Laboratory Medicine, Hangzhou First People’s Hospital, Hangzhou 310006, China Department of Laboratory Medicine, Memorial Sloan Kettering Cancer Center
  6. Department of Pathology and Laboratory Medicine, Weill Medical College of Cornell University, New York, NY 10065, USA

Available online: 2018-04-02

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Abstract

We employed a multiplex polymerase chain reaction (PCR) coupled with capillary electrophoresis (mPCR-CE) targeting six Clostridium difficile genes, including tpitcdAtcdBcdtAcdtB, and a deletion in tcdC for simultaneous detection and characterization of toxigenic C. difficile directly from fecal specimens. The mPCR-CE had a limit of detection of 10 colony-forming units per reaction with no cross-reactions with other related bacterial genes. Clinical validation was performed on 354 consecutively collected stool specimens from patients with suspected C. difficile infection and 45 isolates. The results were compared with a reference standard combined with BD MAX Cdiff, real-time cell analysis assay (RTCA), and mPCR-CE. The toxigenic C. difficile species were detected in 36 isolates and 45 stool specimens by the mPCR-CE, which provided a positive rate of 20.3% (81/399). The mPCR-CE had a specificity of 97.2% and a sensitivity of 96.0%, which was higher than RTCA (x2 = 5.67, P = 0.017) but lower than BD MAX Cdiff (P = 0.245). Among the 45 strains, 44 (97.8%) were determined as non-ribotype 027 by the mPCR-CE, which was fully agreed with PCR ribotyping. Even though ribotypes 017 (n = 8, 17.8%), 001 (n = 6, 13.3%), and 012 (n = 7, 15.6%) were predominant in this region, ribotype 027 was an important genotype monitored routinely. The mPCR-CE provided an alternative diagnosis tool for the simultaneous detection of toxigenic C. difficile in stool and potentially differentiated between RT027 and non-RT027.

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