A haplotype-complete chromosome-level assembly of octoploid <i>Urochloa humidicola</i> cv. Tully reveals multiple genomic compositions and evolutionary histories in the species

We developed a haplotype-resolved, chromosome-scale genome assembly of the Urochloa humidicola (Rendle) Morrone & Zuloaga cultivar Tully, an apomictic C4 forage grass cultivated in the tropics worldwide. We assembled a 4.1 Gb genome into 48 chromosomes (2n = 8x = 48), capturing 99.5% of the BUSCO markers, and annotating 259,254 protein-coding genes. Subgenome assignment revealed an octoploid AABBBBCC structure with three ancestral lineages (A, B, C), and an aneuploid composition of 14A, 22B, and 12C chromosomes. Comparative analyses with related Urochloa species identified U. dictyoneura and U. arrecta as potential progenitors of the B and C subgenomes, respectively. The likely progenitor of the A subgenome remains an unknown wild species from the Humidicola clade. Analysis of LTR-retrotransposons and gene collinearity further indicated a close relationship between A and B ancestries, and a distinct evolutionary path for C. Competitive read mapping across additional U. humidicola accessions supported multiple evolutionary histories in the species, with AABBBB (lacking C ancestry), being the most common. We found that previously described subpopulation structures can be explained by the presence or absence of C ancestry, and that sexual U. humidicola are likely to be autopolyploid from the B ancestry. The genome is available as assembly GCA_965614515.2. This assembly provides the first complete reference for U. humidicola and reveals a multi-ancestral origin and reticulated evolution in U. humidicola. It provides a foundation for studying complex polyploid evolution, regulation of apomixis and biological nitrification inhibition, and molecular breeding strategies.