Closed Systems to Study Plant-Filamentous Fungi Associations: Emphasis on Microscopic Analyses

In nature, filamentous fungi interact with plants. These fungi are characterized by rapid growth in numerous substrates and under minimal nutrient requirements. Investigating the interaction of these fungi with their plant hosts under controlled conditions is of importance for many researchers aiming to proceed with molecular or microscopical investigations of their favorite plant-fungus interaction system. The speed of growth of these fungi complicates transferring plant-fungal interaction systems in laboratory conditions. The issue is more complicated when monoxenic conditions are desired, to ensure that only two members (a fungus and a plant) are present in the system under study. Here, two simple closed systems for investigating plant-filamentous fungi associations under laboratory, monoxenic conditions are described, along with their limitations. The plant and fungal growth conditions, methods for sampling, staining, sectioning, and subsequent microscopical imaging of colonized plant tissues with affordable, common laboratory tools are described. Key features • Setting up closed systems for microscopical observations of plant-filamentous fungi (emphasis on model legumes-) associations and temporal in vivo observations of the association(s). • Preparing root samples for microscopical observations: staining, sectioning, and mounting on microscopical slides. • Using low-cost equipment for performing microscopical observations and imaging. • Using fluorescence microscopy: provision of common fluorophores to highlight specific plant and fungal tissues, compartments, and structures.