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Eighteen specimens of Pimelodus maculatus collected from Tibagi River (Sertaneja, PR, Brazil) were analyzed cytogenetically. The diploid number of 56 chromosomes was observed and karyotype was 20 M + 20 SM + 10 ST + 6 A with fundamental number (FN) of 106. Results of analyses from the nucleolus organizer regions (NORs), obtained by AgNO₃, CMA₃ and C-band staining showed marking in a terminal position on the long arm of a pair of subtelocentric chromosomes. The restriction enzyme AluI produced a linear differentiation similar to C-banding.