Confocal imaging of living fungal hyphae challenged with the fungal antagonist viscosinamide

The combination of confocal laser scan- ning microscopy (CLSM) and staining with vital fluo- rescence dyes has promising potential for studying subcellular structures in living filamentous fungi and oomycetes. CLSM offers high resolution images and the possibility of optical sectioning of the specimen. Vital dyes make it possible to do this while the cells are still alive and with minimum disturbance of cel- lular processes. In this study confocal microscopy and the vital dyes Nile red, SYTO 13, DIOC7(3), and car- boxy SNARF-1 were used on several species. Subse- quently the technique was used to detect effects of the fungal antagonist viscosinamide. These fluores- cent dyes were expected to stain hydrophobic ele- ments, nuclei, mitochondria, and cytoplasm respec- tively. Nile red and DIOC7(3) were found to be es- pecially well suited for confocal imaging. Nile red stained the vacuolar membranes in Rhizoctonia solani