Differential gene expression in a Bacteroides fragilis metronidazole-resistant mutant

• Published in: Journal of antimicrobial chemotherapy Vol. 54; no. 1; pp. 100 - 108
• Main Authors: Diniz, Claudio Galuppo, Farias, Luiz M., Carvalho, Maria Auxiliadora R., Rocha, Edson R., Smith, C. Jeffrey
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01.07.2004; Oxford Publishing Limited (England)
• Subjects: anaerobes; Antibiotics. Antiinfectious agents. Antiparasitic agents; Bacteroides fragilis; Bacteroides fragilis - drug effects; Bacteroides fragilis - enzymology; Bacteroides fragilis - genetics; Biological and medical sciences; DNA Primers; DNA, Bacterial - genetics; drug resistance; Drug Resistance, Bacterial; Electrophoresis, Gel, Two-Dimensional; Fermentation; Gene Expression Regulation, Bacterial - drug effects; Medical sciences; Metronidazole - pharmacology; Microbial Sensitivity Tests; Mutagenesis, Insertional; Mutation - genetics; Pharmacology. Drug treatments; proteomics; RNA - genetics; RNA - isolation & purification; Bacteria; Gene expression; Bacteroides fragilis; Bacteroidaceae
• DOI: 10.1093/jac/dkh256
• ISSN: 0305-7453; 1460-2091

Objectives: The current work focused on molecular changes in a spontaneous Bacteroides fragilismutant selected by low concentrations of metronidazole as an adaptive response to the drug. Methods: A metronidazole-resistant strain derived from B. fragilis ATCC 25285 was selected by passage in the presence of drug using 0–4 mg/L gradient plates. Using a combination of proteomics for identification of differentially expressed proteins by two-dimensional electrophoresis and selected mutational analyses by single cross-over insertion and an allelic exchange, we have identified genes involved in the adaptive response to metronidazole. Results: There are significant changes in the protein profiles of resistant strains. These changes appeared to affect a wide range of metabolic proteins including lactate dehydrogenase (up-regulated) and flavodoxin (down-regulated), which may be involved in electron transfer reactions. Also, the enzymic activity of the pyruvate–ferredoxin oxidoreductase (PorA) complex was impaired. Mutant strains lacking the genes for flavodoxin and PorA were less susceptible to metronidazole than the sensitive parent, and a double flavodoxin/PorA mutant had even less susceptibility but none of the mutants were as resistant as the spontaneous metronidazole-resistant strain. Conclusions: Overall, the data indicated that there were global changes in the regulation of the physiology of the metronidazole-resistant strain. In addition, flavodoxin was identified as an important contributor to metronidazole sensitivity in B. fragilis.