Morphology, Plasmolysis, and Tetrazolium Bromide Stain as Criteria for Determining Viability of Phytophthora Oospores

ABSTRACT

Viability of Phytophthora oospores was determined by morphology, ability to plasmolyze in osmotica and by rose-colored staining with tetrazolium bromide. Morphologically, normal oospores contained one or more nuclei as well as a well-defined central ooplast detectable by light microscopy. Abnormal oospore walls were thicker than normal and did not contain nuclei or an ooplast. Storage of 14-day-old oospores in distilled water for 14 days at 2 C resulted in loss of viability and germinability; however, 30-day-old oospores tolerated storage at 2 C and germinated well. In testing for viability normal oospores plasmolyzed in osmotica (4 M NaCl or 3 M sucrose solutions) but abnormal oospores did not. Cytoplasm of normal oospores contracted to form a central ball-like structure, deplasmolyzed in distilled water and germinated in soil extract. Abnormal oospores did not plasmolyze and did not germinate. Normal oospores also stained rose by the tetrazolium bromide assay. Positive correlations were obtained between percentage of normal-appearing oospores and percentage of oospores which stained rose in color with tetrazolium bromide (r = 0.997), between percentage of normal-appearing oospores and percentage of germination (r = 0.998), between percentage of normal oospores and percentage of oospores that plasmolyzed in osmotica (r = 1.0), and between percentage of oospores that plasmolyzed in osmotica and percentage of germination (r = 0.985).

Key Words:

• pellucid body
• ooplast
• plasmolysis
• tetrazolium bromide
• germination