Intraspecies sequence-graph analysis of the Phytophthora theobromicola genome reveals a dynamic structure and variable effector repertoires

Phytophthora theobromicola is an emerging cacao pathogen recently identified in Brazil as an aggressive agent of black pod rot. We generated genome assemblies for two P. theobromicola isolates using long-read sequencing and five additional isolates using short reads. Comparative analysis revealed a genome size and predicted gene content comparable to P. citrophthora, a closely related species with a broad host range that includes both citrus and cacao. An intraspecies sequence-graph analysis revealed a highly dynamic genome structure with high proportion of variable effectors. Syntenic orthology analysis across 13 Phytophthora species identified orthologous gene groups conserved only in cacao pathogens and others specific to P. theobromicola. RxLR effectors and CAZymes were particularly enriched among lineage-specific syntenic groups, with RxLRs preferentially located near transposable elements and within gene-sparse, repeat-rich regions. Transcriptome analysis of infected cacao tissues showed that 88% of predicted effectors were expressed, with pods exhibiting the highest number of upregulated genes. Notably, several RxLRs classified as P. theobromicola-specific syntenic orthologs were highly expressed in infected tissues, suggesting that these lineage-specific effectors may play key roles in host-pathogen interactions unique to cacao. Together, our findings highlight the dynamic architecture and functional plasticity of the P. theobromicola genome, providing foundational insights into its virulence strategies and supporting future studies on host adaptation and effector evolution in emerging cacao pathogens.可可生疫霉（Phytophthora theobromicola）是一种新发可可病原菌，近期在巴西被鉴定为可可黑荚病的强致病原。本研究利用长读长测序技术对两株可可生疫霉分离株进行了基因组组装，并通过短读长测序完成了另外五株分离株的基因组组装。比较基因组分析显示，其基因组大小与预测基因含量均与亲缘关系较近的广宿主范围病原菌柑橘疫霉（Phytophthora citrophthora）相近，后者可侵染柑橘与可可两种宿主。种内序列图谱分析揭示了该菌高度动态的基因组结构，其中可变效应蛋白占比极高。对13种疫霉属物种开展的共线性直系同源分析，鉴定出仅在可可病原菌中保守的直系同源基因簇，以及可可生疫霉特异性的直系同源基因簇。RxLR效应子（RxLR effectors）与碳水化合物活性酶（Carbohydrate-Active Enzymes，CAZymes）在谱系特异性共线性基因簇中显著富集，其中RxLR效应子优先定位于转座因子附近，以及基因稀疏、重复序列富集的基因组区域。对受侵染可可组织的转录组分析显示，88%的预测效应蛋白均有表达，其中可可荚果组织的上调基因数量最多。值得注意的是，若干被归类为可可生疫霉特异性直系同源基因簇的RxLR效应子在受侵染组织中呈高表达，提示这些谱系特异性效应子可能在可可专属的宿主-病原体互作中发挥关键作用。综上，本研究结果揭示了可可生疫霉基因组的动态结构与功能可塑性，为解析其致病策略提供了基础见解，同时可为后续针对新发可可病原菌的宿主适应性与效应子进化研究提供支撑。