Use of the Polymerase Chain Reaction for the Specific and Direct Detection of Clostridium dijficile in Human Feces

Gumerlock, Paul H.; Tang, Yajarayma J.; Meyers, Frederick J.; Silva, Joseph

doi:10.1093/clinids/13.6.1053

Cited by 93 publications

(63 citation statements)

References 21 publications

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“…Primers PG48 and B, derived from the C. difficile 16S rRNA gene, were used to confirm the identification of strains by PCR (24) and to control the quality of crude DNA extracts (positive PCR controls). 16S rRNA gene amplification confirmed that all 369 strains had been correctly identified as C. difficile.…”

Section: Methodsmentioning

confidence: 99%

Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

et al. 2004

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In addition to the two large clostridial cytotoxins (TcdA and TcdB), some strains of Clostridium difficile also produce an actin-specific ADP-ribosyltransferase, called binary toxin CDT. We used a PCR method and Southern blotting for the detection of genes encoding the enzymatic (CDTa) and binding (CDTb) components of the binary toxin in 369 strains isolated from patients with suspected C. difficile-associated diarrhea or colitis. Twenty-two strains (a prevalence of 6%) harbored both genes. When binary toxin production was assessed by Western blotting, 19 of the 22 strains reacted with antisera against the iota toxin of C. perfringens (anti-Ia and anti-Ib). Additionally, binary toxin activity, detected by the ADP-ribosyltransferase assay, was present in only 17 of the 22 strains. Subsequently, all 22 binary toxin-positive strains were tested for the production of toxins TcdA and TcdB, toxinotyped, and characterized by serogrouping, PCR ribotyping, arbitrarily primed PCR, and pulsed-field gel electrophoresis. All binary toxin-positive strains also produced TcdB and/or TcdA. However, they had significant changes in the tcdA and tcdB genes and belonged to variant toxinotypes III, IV, V, VII, IX, and XIII. We could differentiate 16 profiles by using typing methods, indicating that most of the binary toxin-positive strains were unrelated.

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Section: Methodsmentioning

confidence: 99%

Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

et al. 2004

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“…PCR with primers PG48 and PG49 derived from a C. difficile 16S rRNA gene was used to confirm identification, as previously described (18).…”

Section: Methodsmentioning

confidence: 99%

Prevalence and Genetic Characterization of Toxin A Variant Strains of Clostridium difficile among Adults and Children with Diarrhea in France

et al. 2002

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“…If inhibitory substances are present, they may be specific for certain proteins and may explain the discordant EIA results seen in this study where samples that yielded toxin-positive isolates were recorded by EIAs as GDH positive or TcdA or TcdB negative. Finally, Gumerlock et al proposed that fecal proteases degrade levels of toxins in the stool (38). It is possible that the length of time the samples spent in transit (mean transport time of 8 days) had a detrimental effect on toxin levels in the feces, reducing them below the level of detection of EIA-based assays.…”

Section: Discussionmentioning

confidence: 99%

Laboratory Detection of Clostridium difficile in Piglets in Australia

2014

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bClostridium difficile is a well-known enteric pathogen of humans and the causative agent of high-morbidity enteritis in piglets aged 1 to 7 days. C. difficile prevalence in Australian piglets is as high as 70%. The current diagnostic assays have been validated only for human infections, and there are no published studies assessing their performance in Australian piglets. We evaluated the suitability of five assays for detecting C. difficile in 157 specimens of piglet feces. The assays included a loop-mediated isothermal amplification (LMIA)-PCR for tcdA (illumigene C. difficile; Meridian), a real-time PCR for tcdB (GeneOhm Cdiff; Becton Dickinson), two-component enzyme immunoassays (EIA) for C. difficile glutamate dehydrogenase (GDH) (EIA-GDH) and TcdA/TcdB (EIA-TcdA/TcdB) (C. diff Quik Chek; Alere), and direct culture (DC) (C. difficile chromID agar; bioMérieux). The assays for detection of the organism were compared against enrichment culture (EC), and assays for detection of toxins/toxin genes were compared against EC followed by PCR for toxin genes (toxigenic EC [

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Use of the Polymerase Chain Reaction for the Specific and Direct Detection of Clostridium dijficile in Human Feces

Cited by 93 publications

References 21 publications

Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

Prevalence and Genetic Characterization of Toxin A Variant Strains of Clostridium difficile among Adults and Children with Diarrhea in France

Laboratory Detection of Clostridium difficile in Piglets in Australia

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