Physiology of sexual reproduction in Pythium sylvaticum

An investigation has been made of the physiology of sexual reproduction in the heterothallic fungus, Pythium sylvaticum. Strains 431a, 431b, 431c, 431d and 431e were obtained from leading University for use in this project. It was confirmed that this species has a requirement for sterol for sexual reproduction, as is well known in all Phytophthora spp. Pythium sylvaticum has a sexual system similar to that reported in heterothallic Achlya spp. The strains of Pythium sylvaticum ranged in sexual activity from strong male, through strains of intermediate potency to strong female. Strains c and d. were strong females while strains e and b were strong males. Strain a varies between male and female depending on the strain with which it was paired. Female strains produce oogonia and male strains, antheridia. When strains c and e were paired on opposite sides of an agar plate oospores formed in the centre of the plate when the strains came into contact. When the paired strains were separated by polycarbonate membrane, the induction of oogonia and antheridia occurred but no oospores were formed. Oospores only formed when the hyphae came into contact. The pores in the membrane (0. 2um) pore size, are too small to allow hyphae to pass through, thus fertilisation can not occur and no oospores form. However, there must be some agent or agents produced by each strain which diffuse through the membrane and initiate and control reproduction in the other strain. When 72h old cultures of male and female were paired, antheridial branches were produced by the male after 4.5h and oogonial initials by the female after 6h. Pairing induced with uninduced cultures confirmed the female first induces the male to produce antheridial branches. The induced male then induces production of oogonial initials by the female. Attempts to isolate the inducing hormones were made, but proved unsuccessful. The induction of sexual reproduction v/ithout hyphal contact and the sequential production of antheridia and oogonia is very similar to that described in heterothallic Achlya spp. Sexual reproduction is controlled by steroid hormones in Achlya. To test the hypothesis that the sterols required for reproduction by Pythium sylvaticum are metabolised to steroid hormones, sterol metabolism in this organism was studied. Cholesteryl palmitate was added to the medium in which male and female were grown and paired, or in which strains were grown alone. Cholesterol and metabolites more polar than cholesterol were produced, but only minor differences were noted in the metabolites produced by induced and uninduced strains which could not be related to reproductive phenomena.