Infections with the microaerophilic parasite Trichomonas vaginalis are treated with the
5-nitroimidazole drug metronidazole, which is also in use against Entamoeba histolytica, Giardia intestinalis and microaerophilic/anaerobic bacteria. Here we report that in T. vaginalis the
flavin enzyme thioredoxin reductase displays
nitroreductase activity with
nitroimidazoles, including
metronidazole, and with the nitrofuran
drug furazolidone. Reactive metabolites of
metronidazole and other
nitroimidazoles form covalent adducts with several
proteins that are known or assumed to be associated with
thioredoxin-mediated redox regulation, including
thioredoxin reductase itself,
ribonucleotide reductase,
thioredoxin peroxidase and cytosolic
malate dehydrogenase. Disulphide reducing activity of
thioredoxin reductase was greatly diminished in extracts of
metronidazole-treated cells and intracellular non-
protein thiol levels were sharply decreased. We generated a highly
metronidazole-resistant cell line that displayed only minimal
thioredoxin reductase activity, not due to diminished expression of the
enzyme but due to the lack of its
FAD cofactor. Reduction of free
flavins, readily observed in
metronidazole-susceptible cells, was also absent in the resistant cells. On the other hand,
iron-depleted T. vaginalis cells, expressing only minimal amounts of PFOR and hydrogenosomal
malate dehydrogenase, remained fully susceptible to
metronidazole. Thus, taken together, our data suggest a
flavin-based mechanism of
metronidazole activation and thereby challenge the current model of hydrogenosomal activation of
nitroimidazole drugs.