Persistence despite isolation: Temporal genomic structure in the endemic and endangered killifish Aphanius baeticus (Actinopterygii, Cyprinodontiformes)

Many conservation efforts remain disproportionately focused on charismatic species, leaving much threatened biodiversity, particularly freshwater fishes, overlooked. The Iberian Peninsula is a hotspot of freshwater fish biodiversity, but many endemic species are under severe anthropogenic pressure. Among them, the Baetican toothcarp Aphanius baeticus (Doadrio, Carmona & Fernández-Delgado, 2002) is an endangered endemic species restricted to a small number of fragmented populations along the southwestern Atlantic coast of Spain. We investigated population structure, genetic diversity, divergence, and temporal genomic change in A. baeticus using complementary mitochondrial and nuclear genomic approaches. Analyses were based on mitochondrial cytochrome b, complete mitogenomes, and genome-wide SNPs derived from low-coverage whole-genome resequencing of 54 individuals sampled in 2008 and 2022, approximately 14 years apart. These datasets consistently recovered four major conservation genetic units corresponding to (i) Doñana coastal wetlands, (ii) the inland Guadalquivir basin, (iii) the Guadalete drainage, and (iv) the Iro–Conil system. Across both time periods (~10–14 generations), we detected increased genetic differentiation and a pronounced decline in nucleotide diversity across all populations, consistent with reduced connectivity and demographic decline. While historical gene exchange was supported, likely driven by episodic dispersal during extreme hydrological events, contemporary patterns indicate substantial erosion of that connectivity. Together, these results demonstrate that a formerly metapopulation-like network has transitioned toward increasingly isolated and erosion-prone units within just a few generations. This temporal genomic framework provides an early-warning baseline for conservation genetics and is transferable to other fishes facing comparable conservation pressures.