Chemical Composition and Nutritional Indices of Autochthonous Trifolium repens Populations from Different Origins

White clover (Trifolium repens L.) is a major legume in Mediterranean agroecosystems. This study systematically evaluates 15 autochthonous white clover populations from the Trikala region of Greece, focusing on chemical composition and derived nutritional indices relevant for germplasm characterization and breeding. Fifteen local populations were evaluated under controlled pot cultivation over two consecutive years. Clonal plants were harvested at the early flowering stage. Key traits—crude protein (CP), Ash, Fat, crude fibre (FIBRE), acid detergent fibre (ADF), neutral detergent fibre (NDF), digestible dry matter (DDM), dry matter intake (DMI), and relative feed value (RFV)—were measured. Combined ANOVA revealed significant differences among populations for all traits (p ≤ 0.001), while genotype × year interactions were present but generally minor compared to genotypic effects. Broad-sense heritability was high across most traits (H2 = 90.8–99.4%), demonstrating strong genetic control. CP showed positive correlations with DDM, DMI, and RFV, whereas ADF and NDF were negatively correlated with intake and digestibility. Canonical and discriminant analyses showed that a reduced set of traits (CP, Ash, FIBRE, RFV) contributed strongly to differentiation among populations. Hierarchical clustering (heatmap) confirmed these groupings based on fibre and digestibility-related traits. Populations such as Dendrochori and Gorgogyri consistently showed favorable chemical and nutritional profiles, while Fiki and Dendrochori showed the highest stability across years. The present study highlights substantial genetic variability among local white clover populations and identifies trait structures of relevance for germplasm characterization. These findings enhance the characterization of genetic diversity in Trifolium repens and support its potential use in future breeding research under Mediterranean environments.