Distribution of Diaminobenzidine Reaction Products in Zoospores of Phytophthora palmivora

Abstract

Because microbodies in many filamentous fungi fail to stain with the 3,3''-diaminobenzidine (DAB) technique, the subcellular localization of DAB reaction products at two pHs was explored in an electron microscopic study of zoospores of Phytophthora palmivora. At pH 9.2, but not at pH 7.2, DAB reaction product was in the matrix of U-bodies and microbodies, indicating that both organelles contained catalase. In contrast to a homogeneous distribution in microbodies, reaction product was limited to the core of U-bodies and absent from their halo and shell regions. At both pH 7.2 and 9.2, reaction product filled mitochondrial cristae, the site of cytochrome c oxidase. In addition to the organellar compartmentalization of reaction product, DAB reactions at pH 9.2 and 7.2 enhanced the prominence of an asymmetry in the trilamellar structure of the zoospore plasma membrane and the appearance of a cell coat covering the plasma membrane and flagellar sheath. This coat formed during zoosporogenesis, and DAB reactions intensified cell coat material that lined the inner surfaces of membranes involved in cytoplasmic cleavage. Since no reaction product formed in tissues incubated in reaction media to which sodium azide was added or DAB eliminated, reaction products were enzymatically produced. Because the DAB reaction highlights plasma membrane asymmetry and the presence of a cell coat, this technique should be useful in morphologically marking plasma membranes, enabling their recognition during zoosporogenesis or in isolated subcellular fractions of this fungus.