The glyoxysomal nature of microbodies was determined in Botryosphaeria dothidea hyphae based on morphology and in situ enzyme characteristics by transmission electron microscopy and cytochemistry. Bound by a single membrane, microbodies had a homogeneous matrix and varied in size ranging from 200 to 400 nm in diameter. Microbodies often had crystalline inclusions that consisted of parallel arrays of fine tubules in their matrices. Microbodies and lipid globules were placed in close association with each other, forming microbody-lipid globule complexes in hyphae. The cytochemical activities of catalase and malate synthase were localized in microbodies, showing intense electron density of the organelle. In addition, immunogold labeling detected the presence of catalase in a multivesicular body-like organelle and the cell wall as well as in the matrix and crystalline inclusion of microbodies, supporting the enzyme secretion outward. Meanwhile, isocitrate lyase was localized only in matrices of microbodies. These results suggest that the microbodies complexed with lipid globules in B. dothidea hyphae are functionally defined as glyoxysomes which may enable the fungus to survive latent periods using lipids via the glyoxylate cycle and catalase secretion.