Internal Seed Colonization, Flower-to-Seed Transmission, and Seedling Association of Didymella tanaceti in Pyrethrum

Seedborne inoculum has been proposed as a potential source of primary infection for Didymella tanaceti in pyrethrum ( Tanacetum cinerariifolium ). However, the location of the pathogen within seed associated tissues - that include the pericarp, seed coat and embryo - and the role of seedborne inoculum in disease establishment remain unclear. This study investigated seed colonization, internal survival, and seed-to-seedling transmission of D. tanaceti using light microscopy, histopathology, scanning electron microscopy (SEM), and a fluorescently labeled isolate. In naturally infected seeds, following surface sterilization and culture-based identification, putative D. tanaceti was recovered from the outer seed layer of 27.5% of seeds, from internal tissues (inner seed layer and embryo) of 7.5% of seeds, and from both outer and internal tissues of 3.8% of seeds. Histological and SEM analyses revealed the presence of fungal propagules associated with inner tissues, particularly within inter-tegumentary spaces. In a separate flower inoculation experiment, fluorescent D. tanaceti was detected in 37.3% and 36.0% of seeds from pre- and post-anthesis inoculated treatments, respectively. Among D. tanaceti -positive seeds recovered following post-anthesis inoculation, 40.7% appeared externally asymptomatic despite lacking embryos. Finally, in a separate seed-to-seedling assay, fluorescent D. tanaceti was detected on cotyledons of 64% of seedlings emerging from inoculated seeds. These findings highlight the epidemiological importance of seedborne inoculum in pyrethrum production systems.