Evidence of Genetic Exchange by Parasexual Recombination and Genetic Analysis of Pathogenicity and Mating Type of Parasexual Recombinants in Rice Blast Fungus, Magnaporthe oryzae

Abstract

A selectable marker gene conferring resistance to bialaphos (BI) was introduced into rice blast isolate Y90-71BI and another conferring resistance to blasticidin S (BS) into isolate 3514-R-2BS of Magnaporthe oryzae to demonstrate exchange of DNA. Colonies obtained from co-cultures of these two isolates were resistant to both BI and BS and had both resistance genes as shown by Southern blot analysis of their genomic DNA. Conidia from these BI-BS-resistant isolates had only one nucleus per cell after staining with 4',6-diamidino-2-phenylindole (DAPI). Using flow cytometry, however, these BI-BS-resistant isolates were found to be haploid. Segregation of BI-BS-resistant isolates for pathogenicity (avirulence to virulence) on rice line K59-1 was consistent with a 1:1 ratio, as was segregation for mating type. These BI-BS-resistant isolates were thus apparently derived from parasexual exchange of DNA and the segregation of pathogenicity and of mating type of the parasexual recombinants might correspond to that of the progeny of the offspring of the sexual cross.

Parasexual Cycle Provides Genetic Segregants Equivalent to Sexual Progeny in the Rice Blast Fungus Magnaporthe oryzae

“…We revealed that the segregation of avirulence to virulence (avr/vir) in the progeny derived from crossing Y90-71 and 3514-R-2 on Hattan 3 24 and line K59-1 (unpublished data) was in a 1:1 ratio. The segregation of avr/vir in the parasexual recombinants (BI-BSresistant parasexual recombinants) was consistent with a 1:1 ratio on line K59-1, but not on Hattan 3 15 . We postulated that the segregation of avr/vir of the BI-BS-resistant parasexual recombinants on Hattan 3 was consistent with that of sexual progeny and that the antibiotic-resistance genes and genes for virulence on Hattan 3 might be linked as a result of transformation.…”

“…Mating type was determined by crossing M. oryzae isolates with tester isolates, Y93-164a-1 (MAT1-1, race 132) and Y93-245c-2 (MAT1-2, race 137), as described previously 15 .…”

“…The pathogenicity of the BI-BS-resistant sexual progeny was determined using a previously described method 15 .…”

“…All grew on PDA containing both antibiotics. Pathogenicity was assayed 6 to 7 days after spraying conidial suspension on Hattan 3 and K59-1 seedlings 15,24 (A: avirulence, V: virulence). a): Noguchi et al 15 .…”

“…While numerous studies on the parasexual cycle have been conducted in the rice blast fungus 3,5,6,8,13,14,22,25 , no genetic analyses have been performed on parasexual recombinants that included avirulence genes. Therefore, we carried out a genetic study after obtaining parasexual recombinants from co-cultures of two different antibiotic-resistant isolates of the fungus: isolate Y90-71BI derived from introducing plasmid pBARKS1, a bialaphos (BI)-resistance vector that contains the Ignite/Basta-resistance (bar) gene 1,16 , into isolate Y90-71, and isolate 3514-R-2BS derived from introducing plasmid pBF101, a blasticidin S (BS)-resistance vector that contains the blasticidin S deaminase gene (BSD) 9 , into isolate 3514-R-2 15 . We revealed that the segregation of avirulence to virulence (avr/vir) in the progeny derived from crossing Y90-71 and 3514-R-2 on Hattan 3 24 and line K59-1 (unpublished data) was in a 1:1 ratio.…”