Comparison of the metabolic activities of four wild-type Clostridium perfringens strains with their gatifloxacin-selecte
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ORIGINAL PAPER
Comparison of the metabolic activities of four wild-type Clostridium perfringens strains with their gatifloxacin-selected resistant mutants Fatemeh Rafii • Miseon Park • Gonc¸alo Gamboa da Costa • Luisa Camacho
Received: 29 July 2009 / Revised: 26 September 2009 / Accepted: 30 September 2009 / Published online: 24 October 2009 Ó Springer-Verlag 2009
Abstract The production of short-chain fatty acids, reductive enzymes, and hydrolytic enzymes by four gatifloxacin-selected, fluoroquinolone-resistant, mutant strains of C. perfringens, with stable mutations either in DNA gyrase or in both DNA gyrase and topoisomerase IV, was compared with that produced by the wild-type parent strains to investigate the effect of mutations associated with the selection of gatifloxacin resistance on bacterial metabolic activities. The mutants differed from their respective wild-type parent strains in the enzymatic activities of azoreductase, nitroreductase, and b-glucosidase and in the ratio of butyric acid to acetic acid production. Microarray analysis of one wild type and the corresponding mutant revealed different levels of mRNA expression for the enzymes involved in short-chain fatty acid (SCFA) synthesis and for b-glucosidase and oxidoreductases. In addition to mutations in the target genes, selection of resistance to gatifloxacin resulted in strain-specific physiological changes in the resistant mutants of C. perfringens that affected their metabolic activities. Keywords Clostridium perfringens Fluoroquinolone Resistance Azoreductase Nitroreductase Gatifloxacin Communicated by Wolfgang Buckel. F. Rafii (&) M. Park Division of Microbiology, National Center for Toxicological Research, U.S. Food and Drug Administration, 3900 NCTR Road, Jefferson, AR 72079, USA e-mail: [email protected] G. Gamboa da Costa L. Camacho Division of Biochemical Toxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, 3900 NCTR Road, Jefferson, AR 72079, USA
Introduction Clostridium perfringens, a normal colonic bacterium of humans and other animals, is also a major opportunistic pathogen (Asha and Wilcox 2002; Collie et al. 1998; Meer et al. 1997; Sparks et al. 2001; Vaishnavi et al. 2005). In addition to producing various toxins that are involved in pathogenicity, it produces an assortment of reductive and hydrolytic enzymes and biologically active substances, including short-chain fatty acids (SCFA; Gorbach 1982; Ito et al. 1997; Kobayashi et al. 1995; McBain and Macfarlane 1997, 1998; Meer et al. 1997; Rafii et al. 1991). Some reductive and hydrolytic enzymes are considered carcinogen-metabolizing enzymes in human colonic contents because of their involvement in the activation or detoxification of genotoxic compounds (Gorbach 1982; McBain and Macfarlane 1997; Rafii et al. 1991; Rafii and Cerniglia 1993). C. perfringens ferments dietary carbohydrates in the colon and produces a large amount of SCFA, including butyric acid and acetic acid (Arimochi et al. 2006; Ito et al. 1997). Lumin
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