This article reports on the use of high pressure freezing followed by
freeze substitution (HPF/FS) to study ultrastructural details of
host–pathogen interactions in fungal diseases of plants. The
specific host–pathogen systems discussed here include a powdery
mildew infection of poinsettia and rust infections of daylily and
Indian strawberry. The three pathogens considered here all attack the
leaves of their hosts and produce specialized hyphal branches known as
haustoria that invade individual host cells without killing them. We
found that HPF/FS provided excellent preservation of both haustoria
and host cells for all three host–pathogen systems. Preservation
of fungal and host cell membranes was particularly good and greatly
facilitated the detailed study of host–pathogen interfaces. In
some instances, HPF/FS provided information that was not available
in samples prepared for study using conventional chemical fixation. On
the other hand, we did encounter various problems associated with the
use of HPF/FS. Examples included freeze damage of samples,
inconsistency of fixation in different samples, separation of plant
cell cytoplasm from cell walls, breakage of cell walls and membranes,
and splitting of thin sections. However, we believe that the
outstanding preservation of ultrastructural details afforded by
HPF/FS significantly outweighs these problems and we highly
recommend the use of this fixation protocol for future studies of
fungal host-plant interactions.