The Mediterranean habitat of the Nile soft-shelled turtle (Trionyx triunguis)

The Mediterranean soft-shell turtle (Trionyx triunguis) is classified as critically endangered by the IUCN. Effective conservation requires a clear understanding of its reproductive strategies and population structure. By combining mitochondrial DNA tandem repeat-region profiling with genome-wide SNP data obtained through 3RADseq, we gained high-resolution insights into the genetic composition and breeding behavior of Mediterranean populations. Our results revealed complex reproductive dynamics, including multiple paternity, sperm storage, and repeated nesting within a single season —strategies that enhance genetic diversity in small, fragmented populations. Using SNP-based kinship inference, we estimated the number of breeding females and identified full and half-sibling groups, offering a robust genomic framework for assessing population size and structure. Genetic similarity patterns highlighted moderate differentiation among Israeli river populations, suggesting some connectivity, while samples from Türkiye were clearly distinct, reflecting long-term geographic and genetic separation. This integrative approach provides a scalable, repeatable tool for long-term monitoring. The combined use of maternal and biparental markers enables detailed tracking of genetic diversity, breeding contributions, and demographic trends—key elements for designing informed, adaptive conservation strategies.