SINGLE GENOTYPE AXENIC CULTURES OF CRONARTIUM-QUERCUUM F-SP FUSIFORME

Single genotype axenic cultures of Cronartium quercuum f. sp. fusiforme were isolated from individually cultured haploid basidiospores. Successful single spore initiations were obtained only on Harvey and Grasham (4) basal medium amended with 0.1 g/L CaCO3, 1 g/L each of yeast extract and peptone, and 10 g/L of bovine serum albumin (HGYP + BSA). Nurse culture techniques were necessary for significant numbers of successful colony initiations. Nurse cultures were produced by casting basidiospores from a group of five clustered telial columns onto cellulose nitrate-acetate membranes that covered four test media, including HGYP + BSA. Basidiospores germinated and hyphae grew on these membranes for 2 wk but did not penetrate through to the underlying media. After 2 wk, the membrane was removed from the dishes to yield the conditioned nurse media. Four to 6 days after membrane removal, single basidiospores were inoculated onto the nurse media where overlying nurse spores had previously been cast. Single basidiospore colonies routinely developed in approximately 2 mo on HGYP + BSA nurse medium. This study is believed to be the first report of a rust fungus axenically cultured repeatedly from single spores.