Comparative Genomic Analysis of Four Macadamia Species Reveals Lineage-Specific Gene Family Dynamics and Shared Whole-Genome Duplication

Macadamia is an emerging woody nut with high economic value. Although genome assemblies are now available for all four species, a comprehensive evolutionary analysis integrating divergence time estimation, lineage-specific gene family dynamics, and whole-genome duplication (WGD) history has not yet been undertaken. Here, we performed a comparative genomic analysis of four macadamia species (M. integrifolia, M. tetraphylla, M. jansenii, and M. ternifolia) alongside 11 other plant species. A total of 72,992 gene families were identified, with species-specific gene families numbering 277 in M. integrifolia, 234 in M. tetraphylla, 236 in M. ternifolia, and 183 in M. jansenii. KEGG enrichment analysis revealed that M. integrifolia-unique genes were enriched in terpenoid biosynthesis and ABC transporter pathways, whereas the unique genes of the other three species were predominantly associated with phenylpropanoid biosynthesis and amino acid/carbohydrate metabolism. Phylogenetic analysis revealed a well-supported monophyletic macadamia clade, with M. integrifolia and M. tetraphylla forming one subclade, and M. ternifolia and M. jansenii forming another. Divergence time estimation indicated that the genus diverged approximately 10.3 million years ago (Mya), with species-level divergences occurring at 7.8–8.4 Mya. Genomic synteny analysis revealed a conserved genomic architecture, with 57.36%–59.68% of genes being shared in syntenic blocks among species. Gene family dynamics revealed lineage-enriched patterns in the terpenoid biosynthesis pathways: expansion in M. integrifolia but contraction in the other three species. WGD analysis detected two WGD events. On a relative Ks scale, the more recent WGD event overlapped with the species-level divergence, whereas the older WGD event was shared by the common ancestor of the genus. These findings provide a comprehensive genomic foundation for understanding macadamia evolution and may facilitate breeding and conservation efforts.