Physiological changes in host plants in response to the broad spectrum of fungal modes of infection are still not well understood. The current study was conducted to better understand the infection of in vitro cultures of Pinus sylvestris L. seedlings by three trophically diverse fungal species, Fusarium oxysporum E. F. Sm. & Swingle, Trichoderma harzianum Rifai and Hebeloma crustuliniforme (Bull.) Quél. Biochemical methods and microscopy were utilized to determine (i) which factors (apoplastic and cellular pH, reactive oxygen species, glutathione and cell death) play a role in the establishment of pathogenic, saprotrophic and mycorrhizal fungi, and (ii) whether cell death is a common response of conifer seedling tissues when they are exposed to trophically diverse fungi. Establishment of the pathogen, F. oxysporum, was observed more frequently in the meristematic region of root tips than in the elongation zone, which was in contrast to T. harzianum and H. crustuliniforme. Ectomycorrhizal (ECM) hyphae, however, were occasionally observed in the studied root zone and caused small changes in the studied factors. Colonization of the meristematic zone occurred due to host cell death. Independently of the zone, changes in cellular pH resulting in an acidic cytoplasm conditioned the establishment of F. oxysporum. Additionally, cell death was negatively correlated with hydrogen peroxide (H2O2) in roots challenged by a pathogenic fungus. Cell death was the only factor uniquely associated with the colonization of host roots by a saprotrophic fungus. The mechanism may differ, however, between the zones since apoplastic pH was negatively correlated with cell death in the elongation zone, whereas in the meristematic zone, none of the studied factors explained cell death. Colonization by the ECM fungus, H. crustuliniforme, was associated with a decreasing number of cells with acidic apoplast and by production of H2O2 in the elongation zone resulting in cell death. Saprotrophic and ECM fungi had a greater effect on cell acidification in the meristematic zone than the pathogenic fungus.
Keywords: colonization; conifer; fungal trophic strategy; host cell death; pH; reactive oxygen species.