UC Davis

Fruit infections by fungal pathogens significantly impact the quality, marketability, and safety of produce in preharvest and postharvest scenarios. Fruit ripening, which transforms the fruit into an attractive, nutritious, and delicious food for consumers, also dramatically increases susceptibility to fungal disease. Ripening is an exceptionally complex collection of physiological and biochemical changes produced by large-scale transcriptional reprogramming in a relatively short period time, and identifying individual components of ripening that impact susceptibility is a major challenge. Furthermore, the infection strategies of necrotrophic pathogens are diverse and adapted to the ripening stage of the host.

The major objective of this Ph.D. dissertation has been to isolate specific elements of fruit ripening and fruit-pathogen interactions that have a disproportionate impact on disease outcome and which may serve as targets for breeding or postharvest control. I hypothesized that ripening-associated susceptibility is driven by accumulation of susceptibility factors (such as cell wall-degrading enzymes), and that pathogens can respond to and even manipulate these features during infection. Given the complexity of ripening and fruit-pathogen interactions, I have employed a systems-level approach, leveraging multiple multiple techniques including disease development assays, physiological assessments of ripening, measurement of phytohormones, mutant studies, cell wall glycomics, and various functional transcriptomic methods. The bulk of this research has been conducted using infections of tomato (Solanum lycopersicum) by the model necrotroph Botrytis cinerea, or gray mold (Chapters 1, 2, and 3). However, I have also used the principles and techniques developed in the tomato-B. cinerea pathosystem to expand our knowledge of fruit-pathogen interactions to other systems, specifically infections of tomato by two other necrotrophic pathogens, Rhizopus stolonifer and Fusarium acuminatum (Chapters 1 and 2), and the agriculturally important pathosystem of nectarine and Monilinia laxa, or brown rot (Chapter 4). Through this research, I have detailed infection strategies and virulence factors (Chapters 1, 3, and 4), characterized fruit responses at different developmental stages (Chapters 2, 3, and 4), and identified susceptibility factors that increase during ripening in tomato (Chapter 3).

A major emerging theme of this research has been the importance of pectin integrity and degradation in the fruit cell wall to the infection success. Both host and pathogen catabolism of pectin influence susceptibility to disease. Expression of fungal pectin-degrading enzymes is emphasized on unripe fruit, where the cell wall is more intact than it is in ripe fruit (Chapter 1). The massive upregulation of the tomato gene pectate lyase during ripening, which facilitates fruit softening, substantially increases susceptibility to B. cinerea, as demonstrated by CRISPR mutants (Chapter 2). Moreover, B. cinerea can induce the expression of multiple host pectin-degrading enzymes during infections of unripe fruit, effectively recruiting these enzymes to accelerate softening and increase susceptibility (Chapter 3). This induction appears to be dependent on the B. cinerea pectin-degrading enzymes BcPG1 and BcPG2, which may be required for the initial establishment of infection on unripe fruit (Chapter 3).

The role of the phytohormone ethylene in fruit-pathogen interactions has also obtained additional clarity through this research. Ethylene has contradictory functions in fruit-pathogen interactions, as it both promotes ripening and participates in plant defense signaling. High levels of ethylene in ripe tomato fruit and the inoculated unripe fruit of a hypersusceptible tomato mutant likely result in a net increase susceptibility, regardless of any defense signaling gained, though low, well-regulated levels of ethylene may still be useful during defense of unripe fruit up to 3 days post inoculation (Chapter 2). The eventual accumulation of increasing amounts of ethylene in response to B. cinerea after 4 days post-inoculation may be the trigger for induction of host cell wall degradation and other susceptibility-promoting ripening processes (Chapter 3). Comparable findings from gene expression analyses and ethylene measurements in the nectarine-M. laxa pathosystem indicate that this dual functionality of ethylene is similar elsewhere (Chapter 4).

Untangling increased susceptibility from fruit ripening is of tremendous importance in the ongoing battle against postharvest food losses. My research has progressed our understanding of how specific aspects of pathogen strategy, host response, and transcriptional and physiological changes during fruit ripening impact fungal disease. Additionally, the systems-level approaches I have deployed in the tomato-B. cinerea may serve as guidance for research in non-model pathosystems, as demonstrated in the study of the nectarine-M. laxa pathosystem. Ultimately, this research will assist in the management of postharvest diseases in fruit crops and facilitate breeding efforts for increased resistance to fungal pathogens in ripe fruit.