Refined chloroplast annotations, repeat profiles, and phylogenomic evidence reveal maternal lineage shifts and independent evolution in the Triticum-Aegilops complex

The Triticum-Aegilops complex has a complex reticulate evolutionary history shaped by multiple hybridization and polyploidization events. While nuclear genome relationships have been extensively explored, the maternal inheritance patterns embedded in chloroplast genomes remain unresolved. This study aims to elucidate the maternal lineage dynamics using comparative chloroplast genomics across key Triticum and Aegilops species. We assembled complete chloroplast genomes of three Triticum aestivum cultivars (Keumkang, Saekeumkang, and Olgeuru) and curated a dataset of 20 chloroplast genomes from 15 Triticum and Aegilops species. Annotation refinement revealed conserved gene content and structural stability, with lineage-specific SSR motifs AAAAT/ATTTT and AATAG/ATTCT supporting a maternal contribution of Aegilops speltoides to polyploid wheats. Phylogenetic and SNP-PCA analyses consistently clustered B and G genome-bearing taxa, reinforcing A. speltoides as the maternal donor in these lineages. In contrast, Aegilops tauschii formed a distinct clade and clustered near other Aegilops, suggesting an independent chloroplast origin of the D genome, potentially involving A. mutica as a maternal ancestor. Repeat profiles, codon usage patterns, and selective pressure analyses further underscored the evolutionary conservation and lineage specificity of chloroplast genomes within the complex. Comparative genomics and mVISTA alignments identified several hypervariable regions, such as trnK-rps16 and rpl32-trnL, as potential markers. Codon usage analysis showed strong bias toward A/U-ending codons, and neutrality and ENc-GC3 plots indicated natural selection as the dominant force shaping codon usage. Our findings provide strong evidence for maternal lineage continuity in B/G genome lineages via A. speltoides, while supporting a distinct chloroplast origin for the D genome lineage. This highlights the role of chloroplast genomics in resolving wheat evolutionary history and supports alternative models involving A. mutica in D genome evolution.