Gymnotus is the most studied genus of the order Gymnotiformes, but the morphological similarities of the different species make it difficult to identify taxa reliably. The present study is a continuation of the ongoing research into the taxonomic diversity of the stocks of Gymnotus sold as live bait in the Pantanal, Brazil. These studies have been based on cytogenetic analyses, DNA barcoding, and the analysis of coloration patterns. The results of the cytogenetic analysis confirmed the presence of three distinct strains, recognized as Gymnotus paraguensis, G. sylvius, and G. pantanal. However, the results revealed that the molecular operational taxonomic units identified as G. paraguensis actually include a relatively diverse set of fish, separated by considerable genetic distances. As the G. paraguensis specimens also presented considerable variation in coloration patterns, further genetic diversity analyses were conducted on these individuals, to test the hypothesis that more than one species is present in this cytotaxonomic unit. The haplotype network revealed a regional pattern in the distribution of this species. The results indicate that the observed variation in coloration patterns is associated with a high degree of phenotypic plasticity in G. paraguensis. These findings emphasize the importance of using an integrative approach for a more accurate diagnosis of Gymnotus, in particular, the species marketed as live bait for the fisheries of the upper Paraguay River basin in the Brazilian Pantanal.