Reverse transcription polymerase chain reaction-based method for selectively detecting vegetative cells of toxigenic Clostridium difficile

Mitsutoshi Senoh, Haru Kato, Tomoko Murase, Hideharu Hagiya, Yasuaki Tagashira, Tadashi Fukuda, Masaaki Iwaki, Akihiko Yamamoto, Keigo Shibayama

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

The laboratory diagnostic methods for Clostridium difficile infection (CDI) include toxigenic culture, enzyme immunoassays (EIAs) to detect the toxins of C. difficile, and nucleic acid amplification tests (NAATs) to detect C. difficile toxin genes, but each of these methods has disadvantages; toxigenic cultures require a long time to produce results, EIAs have low sensitivity, and NAATs that target DNA cannot distinguish vegetative cells from spores and dead cells. Here we report a new detection method that uses reverse transcription polymerase chain reaction to target the toxin-gene transcripts. This method was able to specifically detect the vegetative cells of toxigenic C. difficile in fecal samples in spike tests, with a minimum detection limit of 5×102 colony-forming units per 100mg of stool specimen. The performance of this method was also demonstrated in a pilot scale evaluation using clinical fecal specimens, which showed that this method may be more sensitive than EIA and requires a shorter time than toxigenic culture. This method could potentially be applied in the clinical laboratory to detect C. difficile in fecal specimens. The ability of this method to discriminate the presence of vegetative cells from spores and dead cells could help to further the understanding of CDI.

Original languageEnglish
Pages (from-to)615-620
Number of pages6
JournalMICROBIOLOGY and IMMUNOLOGY
Volume58
Issue number11
DOIs
Publication statusPublished - Nov 1 2014

Fingerprint

Clostridium difficile
Reverse Transcription
Polymerase Chain Reaction
Immunoenzyme Techniques
Nucleic Acid Amplification Techniques
Clostridium Infections
Spores
Genes
Limit of Detection
Stem Cells
DNA

Keywords

  • Clostridium difficile
  • Detection method
  • RT-PCR
  • Vegetative cells

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Virology

Cite this

Reverse transcription polymerase chain reaction-based method for selectively detecting vegetative cells of toxigenic Clostridium difficile. / Senoh, Mitsutoshi; Kato, Haru; Murase, Tomoko; Hagiya, Hideharu; Tagashira, Yasuaki; Fukuda, Tadashi; Iwaki, Masaaki; Yamamoto, Akihiko; Shibayama, Keigo.

In: MICROBIOLOGY and IMMUNOLOGY, Vol. 58, No. 11, 01.11.2014, p. 615-620.

Research output: Contribution to journalArticle

Senoh, Mitsutoshi ; Kato, Haru ; Murase, Tomoko ; Hagiya, Hideharu ; Tagashira, Yasuaki ; Fukuda, Tadashi ; Iwaki, Masaaki ; Yamamoto, Akihiko ; Shibayama, Keigo. / Reverse transcription polymerase chain reaction-based method for selectively detecting vegetative cells of toxigenic Clostridium difficile. In: MICROBIOLOGY and IMMUNOLOGY. 2014 ; Vol. 58, No. 11. pp. 615-620.
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