Microscopic, physiological and molecular studies of pathogenesis in Monilinia fructicola, the brown rot pathogen of stone fruits

Monilinia fructicola (G. Wint.) Honey is one of the most destructive pathogens of stone fruit species worldwide. Mechanisms of pathogenesis and the nature of host factors that influence infection and expression of cell wall degrading enzymes are poorly understood for this plant-microbe interaction. A fungal transformation is established that will be very helpful in future studies of gene function and expression during pathogenesis by M. fructicola. Agrobacterium tumefaciens-mediated transformation of M. fructicola generates on average 13 transformants per 105 spores. More than half of the transformants have integrations in the targeted genetic location. T-DNA integration analysis indicates that recombination during non-specific T-DNA integration in this fungus is unlike that reported in Agrobacterium-mediated transformation in plants. M. fructicola appressoria are required for full symptom development on Prunus petals and may serve as resting structures during the quiescent infection period on stage II fruit. M. fructicola appressorium formation depends to a large degree on the stage of fruit development on nectarine. Chemical factors including nutrients, self-inhibitors, fruit volatiles and physical factors such as hydrophobicity and topographical features regulate M. fructicola appressorium development. The results of reporter gene assays, semi-quantitative -RT-PCR analyses, and pathogenicity tests with Mfcut1 mutants are consistent with a role for Mfcut1, a M. fructicola cutinase gene, in brown rot lesion development. Fruit peel phenols influence intracellular redox status to regulate expression of cell wall degrading enzymes that, in turn, may be related to quiescence of nascent infections in stage II fruit. The plant phenols, chlorogenic acid and caffeic acid, have little or no effect on fungal growth, but markedly inhibit the production of fungal cell-wall degrading enzymes, including polygalacturonase and cutinase. Such inhibition is related to changes in redox homeostasis as measured by extracellular redox potentials and intracellular glutathione concentrations. Appressorium formation is inhibited by caffeic acid, chlorogenic acid and reduced glutathione. The three redox-active compounds inhibit brown rot symptom development on flower petals and fruit. Stage II fruit peels have higher total phenol and chlorogenic acid contents than stage III fruit. The data are consistent with peach fruit phenols having a role in regulating quiescence in latent infections of stone fruits by M. fructicola.