Revealing the phylogeography of Astyanax lacustris: insights from the São Francisco River clade

Abstract The genus Astyanax is highly diverse within the family Acestrorhamphidae, and species identification remains challenging due to overlapping morphological, morphometric, meristic, and coloration traits. As a result, several taxa have been considered as species complexes or treated as synonyms. Molecular and cytogenetic approaches are essential for clarifying taxonomic boundaries and assessing potential genetic divergence among Astyanax populations. This study aimed to investigate potential genetic and cytogenetic divergence within A. lacustris from the São Francisco (SF), Northeastern Caatinga and Coastal Drainages (CD), and Upper Paraná (PR) basins. Forty individuals from the Submiddle stretch of the São Francisco River (SMSF) were compared to 102 mitochondrial cytochrome c oxidase subunit I (COI) sequences identified as A. lacustris in the BOLD database from the three ecoregions. The mean K2P divergence between PR and the remaining basins was 3.8%. Both the Bayesian inference tree and the haplotype network revealed a distinct separation between PR samples and the other basins. Cytogenetic analyses also detected differences in karyotype formulas between these groups. Altogether, our results reveal deep genetic and cytogenetic divergence between the Paraná lineage and the São Francisco–Northeastern Caatinga and Coastal Drainages group, highlighting the need for refined taxonomic assessments within this complex.