Identification and biological characteristics of Colletotrichum species combined with cultivar response analysis unravel grape ripe rot resistance mechanisms

• Two phylogenetically distinct Colletotrichum strains CvYL2a and YLRG13 were identified via multi-locus (ITS, ACT, GAPDH) analysis, differing in morphology, reproduction and infection ability, with CvYL2a showing stronger pathogenicity. • CvYL2a exhibits variety-specific infection patterns on grape leaves by completing the biotrophic to necrotrophic infection transition on ‘Thompson Seedless’, remaining confined solely to the biotrophic phase on ‘Beaumont’, and completely failing to infect the highly resistant ‘Liuba-8’ altogether. • H₂O₂ acts as a key defensive signal in grape- Colletotrichum interactions, with elevated levels in incompatible systems mediating resistance by regulating pathogen infection structure development and trophic transition. Grapes ( Vitaceae Vitis L.) are of significant economic importance worldwide, but are severely impacted by grape ripe rot, a fungal disease caused by Colletotrichum species. This study aimed to investigate the economic importance of grapes with varying resistance levels and their interactions with the grape ripe rot fungus to understand pathogenic mechanisms and host defense strategies. Two Colletotrichum strains, CvYL2a ( Colletotrichum viniferum ) and YLRG13 ( Colletotrichum gloeosporioides ), were chosen for a comparative biological characterization. Further inoculation studies were conducted using CvYL2a, which exhibited increased virulence and sporulation capacity. Susceptible Vitis vinifera cv. ‘Thompson Seedless’ leaves and berries were inoculated with CvYL2a for compatible interaction analysis. In contrast, leaves and berries of the resistant Vitis piasezkii accession ‘Liuba-8’ and Vitis riparia cv. ‘Beaumont’ were utilized for incompatible interaction analysis. Analyzed were inoculated ‘Thompson Seedless’ and ‘Beaumont’ berries to compare the development of infection structures (appressoria, penetration hyphae), and the accumulation of H₂O₂. CvYL2a completed the biotrophic-to-necrotrophic colonization cycle in ‘Thompson Seedless’ leaves, but it only achieved biotrophic colonization in resistant ‘Beaumont’ leaves. The highly resistant ‘Liuba-8’ suppressed appressorium formation and exhibited no hyphal growth in leaves. Appressorium formation was delayed, and hyphal differentiation was restricted in ‘Beaumont’ berries. Moreover, there was a significant increase in H 2 O 2 production during incompatible interactions, underscoring its importance as a crucial defense signaling molecule. These findings illustrate that resistance levels of grape cultivars vary based on their impacts on pathogenetic structure development and trophic lifestyle transitions.