Pathogenesis of apple by the facultative parasite Penicillium expansum was critically dependent upon certain physiological events during the primary stages of spore germination. A comparison of two prototrophic isogenic strains of the fungus, one of which was virulent, the other avirulent, revealed that both produced an apple-macerating factor when the spores germinated in vitro. However, germination and growth of the avirulent strain in apple extract and in apple fruit was significantly lower than with the virulent strain. This contrasting behaviour was attributed to the differential sensitivity of the two strains to an inhibitory substance of host origin. Radioactive isotope studies indicated that, with the avirulent organism, the inhibitor interfered with the incorporation of 14C into RNA from labelled [6-14C]orotic monohydrate during germination; under similar conditions RNA synthesis by the virulent strain was stimulated. Incorporation of 14C into protein from labelled [U-14C]L-arginine monohydrochloride was not influenced by the inhibitor, nor was the differential sensitivity of the two strains attributable to differences in respiration, although apple extract greatly reduced the evolution of 14CO2 from [1-14C]D-glucose in both strains. The inhibitor, which was water-soluble and dialyzable, appeared to have a fundamental effect upon the metabolism of the spores in the initial stages of germination, and is postulated to be a primary determinant in the virulence of P. expansum. The data support the hypothesis that the mechanism controlling the pathogenicity of a prototroph may be distinct from that which governs the basic nutrition of the organism.