Salinity-associated differential gene expression in natural populations of the euryhaline killifish Aphanius iberus

Salinity is a major ecological driver in aquatic environments and strongly influences the physiology, distribution, and survival of fish species. While many fishes are restricted to narrow salinity ranges, euryhaline species can tolerate large osmotic fluctuations despite the substantial physiological adjustments required. The Spanish toothcarp, Aphanius iberus, an endemic Mediterranean killifish, is one of such exceptional species capable of inhabiting environments ranging from freshwater to hypersaline systems. Despite this remarkable resilience, the molecular mechanisms underlying salinity tolerance and osmoregulatory plasticity in this species remain poorly understood.

We used RNA sequencing to analyze gill and gastrointestinal tract transcriptomes from individuals belonging to four wild populations distributed along a natural salinity gradient: freshwater-like low-salinity habitats (0.76 PSU); brackish habitats (4.8 and 7.4 PSU at sampling); and hypersaline lagoon-associated habitats (52 PSU). Differential gene expression analyses revealed strong tissue-specific patterns and salinity-associated differences among population contrasts. A substantially higher number of differentially expressed genes was detected in the gills, where genes associated with ion transport, cytoskeletal remodeling, and energy metabolism varied across contrasting osmotic environments, consistent with the central role of this tissue in osmoregulation. In the gastrointestinal tract, pathways related to lipid and carbohydrate metabolism were particularly represented, especially in brackish population contrasts. In addition, genes previously associated with osmotic stress, cellular remodeling and genome regulation, including ATPases, histones and transposable element-related sequences, showed significant expression differences across populations.

These findings offer novel insights into the molecular architecture of salinity tolerance in euryhaline fishes, highlighting coordinated transcriptional responses across tissues involved in ion regulation and metabolic adjustment and improving our understanding of how euryhaline fishes cope with extreme and fluctuating osmotic environments. From a conservation perspective, identifying the molecular basis of this physiological plasticity contributes to the management of this highly threatened endemic species and the dynamic coastal habitats it inhabits.