Comparative transcription analysis and toxin production of two fluoroquinolone-resistant mutants of Clostridium perfringens

BACKGROUND: Fluoroquinolone use has been listed as a risk factor for the emergence of virulent clinical strains of some bacteria. The aim of our study was to evaluate the effect of fluoroquinolone (gatifloxacin) resistance selection on differential gene expression, including the toxin genes involved in virulence, in two fluoroquinolone-resistant strains of Clostridium perfringens by comparison with their wild-type isogenic strains. RESULTS: DNA microarray analyses were used to compare the gene transcription of two wild types, NCTR and ATCC 13124, with their gatifloxacin-resistant mutants, NCTR(R) and 13124(R). Transcription of a variety of genes involved in bacterial metabolism was either higher or lower in the mutants than in the wild types. Some genes, including genes for toxins and regulatory genes, were upregulated in NCTR(R) and downregulated in 13124(R). Transcription analysis by quantitative real-time PCR (qRT-PCR) confirmed the altered expression of many of the genes that were affected differently in the fluoroquinolone-resistant mutants and wild types. The levels of gene expression and enzyme production for the toxins phospholipase C, perfringolysin O, collagenase and clostripain had decreased in 13124(R) and increased in NCTR(R) in comparison with the wild types. After centrifugation, the cytotoxicity of the supernatants of NCTR(R) and 13224(R) cultures for mouse peritoneal macrophages confirmed the increased cytotoxicity of NCTR(R) and the decreased cytotoxicity of 13124(R) in comparison with the respective wild types. Fluoroquinolone resistance selection also affected cell shape and colony morphology in both strains. CONCLUSION: Our results indicate that gatifloxacin resistance selection was associated with altered gene expression in two C. perfringens strains and that the effect was strain-specific. This study clearly demonstrates that bacterial exposure to fluoroquinolones may affect virulence (toxin production) in addition to drug resistance.