tcdA As a diagnostic target in a loop-mediated amplification assay for detecting toxigenic Clostridium difficile

Area of Science:

• Clinical microbiology
• Molecular diagnostics
• Infectious disease detection

Background:

• Two commercial assays, illumigene® and GeneOhm®, target different genes (tcdA and tcdB, respectively) for Clostridium difficile detection.
• Assessing the illumigene® assay's performance using the tcdA gene is crucial, especially for strains with tcdA deletions.

Purpose of the Study:

• To evaluate the efficacy of the illumigene® loop-mediated amplification assay targeting the tcdA gene for detecting toxigenic Clostridium difficile.
• To compare the illumigene® assay's performance against a reference method (GeneOhm® real-time PCR).

Main Methods:

• Testing of 38 C. difficile strains and 108 patient stool specimens using the illumigene® assay.
• Comparison with the GeneOhm® real-time PCR assay as the reference standard.
• Resolution of discordant results through repeat testing, anaerobic culture, and a custom real-time PCR assay.

Main Results:

• Both assays detected all 37 toxin B-positive C. difficile strains, including toxin A-negative, B-positive isolates.
• High agreement (96.3%) was achieved between the illumigene® assay and the reference method after resolving discordant results.
• No cross-reactivity was observed with other Clostridium species or toxin-negative C. difficile.

Conclusions:

• The illumigene® assay, targeting the tcdA 5' region, is a practical and effective molecular target for identifying toxigenic Clostridium difficile.
• The assay demonstrates reliable performance across various C. difficile strains, including those with tcdA deletions.