RESUMEN
Killifishes are a unique and diversified fish group composed of several annual species inhabiting ephemeral pools in tropical and subtropical regions. Moreover, many genera have probably undergone fast speciation with a high number of short-ranged species of controversial taxonomy. This is the case of the "Hypsolebias flavicaudatus" complex (Aplocheiloidei, Rivulidae) that includes nine recently described taxa of remarkable morphological similarity, endemic to Brazil. Considering the potential of cytotaxonomy of killifishes, as presently revised, we performed cytogenetic analyses in two sister species within this complex (H. flagellatus and H. janaubensis), including the first mapping of ribosomal genes in Hypsolebias. Despite sharing 48 chromosomes and similar distribution of heterochromatin and 5S rDNA, their karyotype formula probably differed as a result of pericentric inversions. In addition, H. flagellatus presented a single pair bearing 18S rDNA and GC-rich regions, while multiple GC-rich and 18S signals (up to 28 chromosomes) were detected in H. janaubensis. These results reinforce the dynamism of karyotype evolution in annual killifishes favored by population isolation and small effective size. Thus, cytogenetic variation seems to be closely associated with speciation in Aplocheiloidei, representing a useful tool for identifying similar species in Hypsolebias complex, which are vulnerable to extinction.