Integrative chromosome‐scale genome analysis of cupuassu provides insights into witches' broom disease resistance and expands genomic resources for Theobroma

Cupuassu (Theobroma grandiflorum) is a fruit tree native to the Brazilian Amazon and increasingly relevant to regional bioeconomies. Its cultivation is severely affected by witches' broom disease (WBD), caused by Moniliophthora perniciosa. While a chromosome-scale genome of the susceptible genotype C1074 is available, the lack of a resistant reference has limited investigation into the genomic basis of resistance. Here, we present the first chromosome-scale assembly of the WBD-resistant genotype C174 (415.8 Mb) and a comparative analysis with C1074 integrating structural variant detection, gene-duplication profiling, transposable-element (TE) annotation, and time-resolved host-pathogen transcriptomics. C174 exhibits distinctive tandem and dispersed duplications, genotype-specific TE insertions, and coordinated defense-gene expression, together with higher heterozygosity indicative of broader allelic diversity. Genome-wide, TEs show differential expression and spatial proximity to immune loci, suggesting potential regulatory importance. Although C1074 encodes more nucleotide-binding domain leucine-rich repeat receptors (541 vs. 434), most remain transcriptionally inactive, whereas C174 shows sequential activation of pattern-recognition receptors, mitogen-activated protein kinase components, transcription factors, and pathogenesis-related proteins. Within the Chromosome 6 previously identified resistance quantitative trait locus, two duplicated DUF4220/DUF594 genes (where DUF is domain of unknown function) unique to C174-orthologous to a maize gene implicated in fungal response before-are infection-induced and display signatures of episodic positive selection. Together, these results establish a high-quality genomic framework for exploring the molecular architecture of WBD response in T. grandiflorum. The datasets generated here-including the C174 and C1074 reference genomes, immune-related variant catalog, and prioritized defense-gene lists-constitute a comprehensive open resource for evolutionary, functional, and breeding research of Theobroma species.