The agriculturally important genus Colletotrichum is an emerging model pathogen for studying defense in Arabidopsis. During the process of screening for novel pathogenic Colletotrichum isolates on Arabidopsis, we found significant differences in defense responses between detached and attached leaf assays. A near-adapted isolate Colletotrichum linicola A1 could launch a typical infection only on detached, but not attached, Arabidopsis leaves. Remarkably, resistance gene-like locus RCH1-mediated resistance in intact plants also was compromised in detached leaves during the attacks with the virulent reference isolate C. higginsianum. The differences in symptom development between the detached leaf and intact plant assays were further confirmed on defense-defective mutants following inoculation with C. higginsianum, where the greatest inconsistency occurred on ethylene-insensitive mutants. In intact Arabidopsis plants, both the salicylic acid- and ethylene-dependent pathways were required for resistance to C. higginsianum and were associated with induced expression of pathogenesis-related genes PR1 and PDF1.2. In contrast, disease symptom development in detached leaves appeared to be uncoupled from these defense pathways and more closely associated with senescence: an observation substantiated by coordinated gene expression analysis and disease symptom development, and chemically and genetically mimicking senescence.
Understanding the infection biology of fungi is the key step in devising suitable control strategies for plant diseases. Recently, the Arabidopsis-Colletotrichum higginsianum (causal agent of anthracnose) system has emerged as a seminal paradigm for deciphering the infection biology underlying fungus-plant interactions. We describe here three staining methods coupled with confocal microscopy: trypan blue, aniline blue and dual trypan blue-aniline blue fluorescence staining. Trypan blue and aniline blue staining were employed to scan the infection structures of the hemibiotrophic fungus C. higginsianum and host response in A. thaliana leaf tissues. The two techniques then were combined to observe the contrast between in planta fungal infection structures, i.e., infection vesicles, primary hyphae and secondary hyphae, and the host plant defense responses, i.e., papilla formation and hypersensitive response. These staining techniques also were applied to the lentil-C. truncatum pathosystem to demonstrate their applicability for multiple pathosystems.
Understanding the infection biology of fungi is the key step in devising suitable control strategies for plant diseases. Recently, the Arabidopsis-Colletotrichum higginsianum (causal agent of anthracnose) system has emerged as a seminal paradigm for deciphering the infection biology underlying fungus-plant interactions. We describe here three staining methods coupled with confocal microscopy: trypan blue, aniline blue and dual trypan blue-aniline blue fluorescence staining. Trypan blue and aniline blue staining were employed to scan the infection structures of the hemibiotrophic fungus C. higginsianum and host response in A. thaliana leaf tissues. The two techniques then were combined to observe the contrast between in planta fungal infection structures, i.e., infection vesicles, primary hyphae and secondary hyphae, and the host plant defense responses, i.e., papilla formation and hypersensitive response. These staining techniques also were applied to the lentil-C. truncatum pathosystem to demonstrate their applicability for multiple pathosystems.
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