Visualising endophytic fungi within leaves by detection of (1→3)-ß-d-glucans in fungal cell walls

The presence of endophytic fungi within symptomless leaves of vascular plants is usually recognised indirectly through culturing methods. In order to understand the biology of fungi isolated as endophytes, there is a need to directly observe their hyphae within the leaves of their hosts. Such observations provide information about the mode of infection, the extent of colonisation within the leaf, and the reaction of the plant to infection by the fungus. Many endophytic fungi develop highly localised infections with small amounts of mycelium, making such direct observations difficult. We describe a method adapted from an electron microscopy protocol that labels one of the constituent components of fungal cell walls with a fluorescent dye and enables them to be observed in thin sections under a compound microscope.