Genome-Wide Characterization and Evolutionary Conservation of the Mildew Locus O Gene Family in Coffea Species Reveal Candidate Powdery Mildew Susceptibility Genes

Coffee (Coffea spp.) represents a high-value perennial crop globally, yet its productivity is increasingly threatened by pathogens such as powdery mildew (PM).While PM resistance is often achieved through loss-of-function mutations in Mildew Locus O (MLO) susceptibility genes, these have remained poorly explored in coffee despite the recent availability of high-quality chromosomelevel genome assemblies.A total of 90 MLO genes were identified across the four Coffea genomes and classified into seven major phylogenetic clades.Notably, six members of the Clade V-ii subgroup, orthologous to known PM susceptibility factors, exhibited exceptional sequence and three-dimensional structural conservation supported by AlphaFold3 modeling.These candidates were characterized by conserved C-terminal motifs, including a DFSF consensus sequence as well as specific enrichment of defense-related cis-regulatory elements, in this case TC-rich repeats, within their promoter regions.These findings highlight their potential utility as molecular targets for resistance breeding and genome-editing strategies in coffee improvement programs.This study provides comprehensive insights into the evolutionary conservation of the MLO gene family in Coffea species and identifies candidate susceptibility-associated genes for PM resistance.