Background. In taxonomic studies of fish, the use of preserved samples has been a common practice. Under the framework of morphometrics, the effect of the preservation techniques on body shape is rarely taken into account. Changes during preservation can cause errors in the results, which may eventually lead to wrong conclusions. To explore the effect of a traditional preservation procedure, we quantified the changes in body shape and size of the golden mojarra, Diapterus aureolus (Jordan et Gilbert, 1882), using geometric morphometrics. Materials and Methods. Fish fresh samples were photographed and frozen for at least seven months. Then, they were set in 10% formaldehyde, and passed through a wash of tap water and submerged in 70% ethanol. After five months the samples were photographed again. The differences between the mean shapes of two data groups (fresh fish and fixed specimens) were calculated by using the Procrustes distance. Effects associated with preservation were visualized by comparing the mean shape from each group, using a thin plate spline. A canonical variate analysis was carried out to detect the degree of intra- and inter-specific variation with D. brevirostris as an outgroup. Also, matrices of correct classification, based on Mahalanobis distances, were obtained. Results. Procrustes distance between the two data sets was statistically significant (P < 0.001), suggesting that the fixation process produces changes in body shape. Canonical Variate Analysis verified that there were significant differences among three groups (fresh D. aureolus, fixed D. aureolus, and D. brevirostris; P < 0.001 for both CV1 and CV2), which were correctly classified (98.9%). In spite of the significant differences detected between the fresh and fixed data sets, these were less than those found in the outgroup. The pooled data of all specimens belonging to D. aureolus (fresh and fixed) produced a definitive correct classification (100%) between D. aureolus and D. brevirostris, indicating that the differences caused by the preservation method is not enough to confound species and, consequently, taxonomic integrity was totally acceptable. Conclusion. Our results show a change in size and shape that are consistent with changes obtained with this and other methods applied to other species: generally shapes change and sizes tend to shrink with preservation of specimens. This is probably caused by dehydration from freezing and thawing, exposure to ethanol, and dissolution of skeletal structures caused by formaldehyde. While this effect was clear, it was not enough to confuse species. Caution is advised when working with groups with a close relation (populations or not clearly-defined species). We recommend a similar exercise with a case study with these levels of divergence.
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