Phylogenetic Resolution and Conflict in the Species-Rich Flowering Plant Family Leguminosae

The Tree of Life is central to evolutionary biology, yet resolving deep, recalcitrant phylogenetic relationships remains challenging due to complex processes such as incomplete lineage sorting (ILS), hybridization, and polyploidization. Although previous phylogenetic studies have advanced our understanding of Leguminosae (Fabaceae), a species-rich and ecologically diverse family, many deep relationships at the tribal and higher levels remain unresolved. Incorporating newly generated genome skimming data for 231 species with previously issued plastid genomic, mitochondrial genomic and transcriptomic data, we reconstructed a phylogeny of the family using whole plastomes, 39 mitochondrial genes, and 1559 low-copy nuclear genes, achieving dense taxonomic sampling across almost all recognized tribes and major unplaced lineages. Our results supported the monophyly of the six subfamilies and 49 recognized tribes, identified ten clades worthy of recognition as new tribes in subfamily Papilionoideae, and clarified many contentious relationships. However, nuclear-nuclear and cytonuclear conflicts persist at multiple nodes among trees inferred from different datasets and analytical methods. We proposed the most probable resolution for 22 contentious nodes by applying nuclear gene-tree quartet analysis with corroboration from support of nuclear Maximum Likelihood (ML) and ASTRAL trees. Our results indicate ILS significantly contributes to observed phylogenetic conflicts, while gene flow represents an additional and previously underappreciated factor that mainly contributes to cytonuclear conflicts, particularly along the branches of the Angylocalyceae + Dipterygeae + Amburaneae (ADA) clade and Wisterieae. These processes likely underlie recalcitrant phylogenetic relationships, such as those within the 50-kb inversion clade of Papilionoideae. Our study uses multiple data partitions and analytical methods to resolve contentious phylogenetic relationships in Leguminosae, resulting in a robust phylogenomic framework to guide further investigations in this economically important and exceptionally diverse family.