Sagittal otoliths are structures integrating the sensory system of teleost fish, assisting in balance and hearing and are normally formed by calcium carbonate as aragonite. Sagittal otoliths have intraspecific shape, the aim of this study was to check the frequency of anomalous otoliths and to characterize the crystallization of sagittal otoliths in Atherinella brasiliensis. For that, 45 specimens of A. brasiliensis were collected in the Paranaguá Estuarine Complex (25°32'59” S–48°23'20” W), from which the otoliths were removed from the palate region. Raman spectrometry was performed on 30 otoliths; aragonite deposition was observed in otoliths without morphological changes (n = 26) and aragonite (n = 2) and vaterite (n = 2) deposition in those with morphological changes. Therefore, the frequency of otoliths with morphological changes was 8.8%. The fact that this is the first report of otoliths with morphological changes and deposition of vaterite for A. brasiliensis, demonstrates the need for studies that correlate environmental changes with physiological stress in estuarine fish.