Weakening and Delayed Mortality of Fusarium oxysporum by Heat Treatment: Flow Cytometry and Growth Studies

Abstract

Survival of Fusarium oxysporum f. sp. niveum following heat treatments was studied using flow cytometric, physiological, and microscopic assays. We exposed germinating conidia to sublethal temperatures from 36 to 42°C for 60 min, followed by rhodamine 123 staining and flow cytometry, and found increasing levels of fluorescence that reflect a change in mitochondrial membrane potential, indicating a weakening induced by stress. Viability of conidia or germinating conidia of the fungus exposed to heat decreased with increasing temperature, as assessed by fluorescent staining. However, viability was higher than that assessed with the 5-day-long plate count method and was further reduced 13 and 24 h after treatment, suggesting delayed mortality of the heat-treated germinating conidia. Delayed mortality was substantiated by observing these conidia with light and fluorescent scanning electron microscopy and by subculturing single germinating conidia that had been previously heated. Programmed cell death was not observed in heat-treated conidia or germinating conidia of F. oxysporum based on the detection of plasma membrane phosphatidylserine translocation, cell-cycle measurements, detection of DNA fragmentation, or microscopic observation of apoptotic bodies. We hypothesize that propagules, which survived the heating and apparently are alive, may undergo further irreversible detrimental processes, eventually leading to their death by yet unidentified mechanisms. These findings suggest that pathogen propagules also might be affected under lower temperatures, possibly facilitating pathogen control by heating.