Mycelia of Saprolegnia ferax were used to study the functional significance of metabolism of the Δ24,25-bond in the sterol side chain of oomycetous fungi. The fungus produced a mixture of Δ5-24-desalkyl (cholesterol and cholesta-5,24-dienol) and 24-alkylidene sterols (24-methylenecholesterol and 24Z-ethylidenecholesterol). Through feeding and trapping studies using 24-tritiolanosterol, the mechanism of C-24 alkylation was confirmed to involve a 1,2-hydride shift from C-24 to C-25. The mechanism of C-24 alkylation and Δ24-reduction was interfered with by 25-azacholesterol. When 10 ppm of the sterol biosynthesis inhibitor was fed to S. ferax, growth remained essentially unchanged, although Δ24-reduction and S-adenosylmethionine C-24-transferase activity were blocked producing an altered sterol composition. In contrast to the growth response, oosporogenesis was completely abolished. The results are interpreted to imply that S. ferax produces multiple sterol end products each of which plays different physiological roles in sterol-controlled developmental regulation of fungal ontogeny.