INTERSPECIFIC HYBRIDIZATION BETWEEN PSEUDOCERCOSPORELLA-HERPOTRICHOIDES AND P-ANGUIOIDES ACHIEVED THROUGH PROTOPLAST FUSION

The relatedness of the cereal eyespot pathogen, Pseudocercosporella he rpotrichoides, to the weakly pathogenic species P. anguioides was inve stigated using protoplast fusion to produce somatic hybrids. Crosses b etween P. anguioides and strains representing the two main pathotypes of P. herpotrichoides (W and R) were conducted. Procedures for protopl ast isolation and regeneration developed for P. herpotrichoides were s uccessfully used, without modification, with P. anguioides. PEG-induce d fusion of protoplasts, isolated from auxotrophic mutants of P. herpo trichoides and P. anguioides, generated prototrophic fusion products o n minimal regeneration medium at low frequency (0.00016%-0.0016%). Sub culture of fusion products on non-selective media generated spontaneou s segregants with a range of morphological phenotypes. Characterizatio n of parental nutritional and isozyme markers revealed novel marker co mbinations, confirming that genetic recombination between the species had occurred, presumably by parasexual processes following karyogamy. Both pathotypes of P. herpotrichoides appeared able to hybridize with P. anguioides under these conditions. Marker segregation was not rando m, suggesting structural differences in the genomes of the two species of the expression of genomic incompatibility. Determination of spore sizes of species and hybrids showed that the recombinants formed conid ia intermediate in length between the parental species. Mean cell leng th of conidia remained relatively constant at 9.7 +/- 0.2 mum (standar d error) per cell, with longer spores containing more cell compartment s. No evidence to indicate that the hybrid recombinants were diploid w as obtained from the spore analysis and these strains remained stable after exposure to the haplodizing agent carbendazim. Clear evidence fo r the segregation of pathogenicity to wheat cv. Beaver was obtained fr om a 9 wk pathogenicity test with a range of hybrids and parental isol ates. P. anguioides was only weakly pathogenic to wheat, whereas both pathotypes of P. herpotrichoides caused severe lesion development. Int erspecific hybrids varied in pathogenicity. Some were non-pathogenic, like the P. anguioides parent, while others gave infection levels comp arable to the p. herpotrichoides R-type parent. Pathogenicity was not impaired by the presence of the cysteine auxotrophic marker of the R-t ype. The successful recovery of interspecific hybrids indicates that t hese two species are relatively closely related.