A number of fish groups, such as Gobiidae, are highly diversified and taxonomically complex. Extensive efforts are necessary to elucidate their cryptic diversity, since questions often arise about the phylogenetic aspects of new species. Clarifications about the diversity and phylogeny of the Bathygobius species from the southwestern Atlantic are particularly needed. Evidence has been accumulating on the Brazilian coast regarding the possible presence of new species while doubts remain about the taxonomic status of others. The taxonomic identification of some species of Bathygobius has been problematic, given their generally conservative external morphology, and several species are recognized as cryptic. This situation hinders understanding the real diversity in this taxon. Taken together, genetic, cytogenetic and morphometric analyses have been effective in identifying new species of this genus. Here we describe the karyotypic features and morphological patterns of three Western South Atlantic species of Bathygobius. Furthermore, its cytochrome c oxidase I (COI) gene sequences were compared with those of species from Central America, North America and the Caribbean. The broad analyses performed demonstrated an unsuspected diversity, leading to the identification of an un-described new species (Bathygobius sp.2) and the geographic redefinition of another, Bathygobius sp.1, undoubtedly a branch of B. geminatus, hitherto inaccurately identified as B. mystacium on the coast of Brazil.
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