In this study, genotoxic potential of metronidazole. an antibiotic-antiparasitic agent widely used both clinical and veterinary areas. was evaluated using the piscine micronucleus test. Specimens of Oreochromis niloticus (Pisces: Cichlidae) were exposed to different concentrations; (5, 10, 15 mg/L) of metronidazole and 4 mg/L cyclophosphamide (as positive control) for 24, 48 and 72 h. A fluorescent staining technique with acridine orange (AO) was performed to improve the sensitivity of erythrocyte micronucleus assay with fish. AO-selectively stains newly formed immature erythrocytes (PCEs) therefore allows to obtain more sensitive results by detection of the micronucleated PCEs differed than mature erythrocytes (NCEs) by their RNA-containing cytoplasm. The ratio of PCE/NCE in peripheral blood was also evaluated to assess cytotoxicity. As a result. it was observed that the frequencies of micronucleated PCEs increased both dose and time dependently while PCE/NCE ratios decreased. Our results revealed that metronidazole has cytotoxic and genotoxic effects on fish. The use of acridine orange staining technique also seems useful in assessment of short-term genotoxic effects of chemicals. when fish are used as experimental animal. (C) 2004 Elsevier B.V. All rights reserved.