Plasmopara viticola, the causal agent of grapevine downy mildew, exhibits substantial intraspecific variation in pathogenicity and genetic diversity, yet the genomic features underlying this variation remain incompletely characterized. Here, we sequenced and assembled two P. viticola isolates, PvH (from Vitis vinifera) and PvS (from V. amurensis), using PacBio HiFi sequencing, and performed comparative genomic analysis. Two complete genome assemblies (17 chromosomes) of P. viticola (PvH: 115.3 Mb; PvS: 113.0 Mb) were generated and revealed that nearly 90% of the putative effectors exist as local duplicated gene clusters. Comparative genomics uncovered distinct intraspecific expansion, deletion, and diversification of putative effectors driven by local segmental, tandem, and proximal duplication events in P. viticola. Specifically, PvH exhibited a ~1.4-fold increase in CRNs (PvH: 237; PvS: 183; PV221: 169) and harbored 35 strain-specific CRNs. These differential effectors were predominantly clustered in complex structural variation hotspots (SVs, duplication and inversion). Notably, 104 putative effectors—including 21 RxLRs, 59 CRNs, and 24 CAZymes—were located within inversion regions. Together, our results highlight a highly dynamic genome architecture in P. viticola, in which SV and local gene duplication are closely associated with effector diversification. This study provides a genome-resolved comparative framework for understanding intraspecific genomic diversity in P. viticola and establishes a foundation for future population-level and functional investigations.

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