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The conidia of Pyricularia oryzaewere cultured on barley grain medium for 17 days in this study. Two hundred milliliters of barley grains of each tested strain were cultured in a 500 mL Erlenmeyer flask. The conidia were eluted with 300 mL of sterile water from the cultivated barley grains in each flask. The barley grains were placed in aluminum plates for continued cultivation. The conidia were eluted with 48 h of interval culture, and the elution method used each time was the same as that used for the first elution. All the tested strains could be eluted 6 times; thus, we obtained a large number of conidia. The numbers of conidia produced in the elutions of all the strains were significantly different, with the smallest number produced in the first elution, followed by a gradual increase and then a gradual decrease after reaching the peak; the number of conidia produced in the sixth elution, that is, the final elution, was also low. The total number of conidia and the total number of effective conidia differed among the strains. A large number of effective conidia can be obtained by using the method described in this study, and much time can be saved because the conidia can be eluted six times in one culture cycle.

The objective of this study was to preliminarily analyze the differences in pathogenicity and physiologic races between Pyricularia oryzaeisolates from indicaand japonicarice varieties. Five to 7 isolates of P. oryzaefrom indicaand japonica,having good sporulation capacity, were screened out in each experimental year. To determine the pathogenicity of P. oryzae, different kinds of rice varieties were inoculated with mixtures of indicaand japonicaisolates. A total of 46 isolates, including 23 indicaisolates and 23 japonicaisolates, were screened out in four years. In total, 1252 rice varieties, including 490 indica, 296 japonicaand 466 indica-japonicahybrid rice varieties, were inoculated. Our results suggested that the degree of disease was more severe in most indicavarieties infected with indicaisolates than in those infected with japonicaisolates. That is, indicaisolates were more inclined to infect indicavarieties. A majority of japonicaand indica-japonicahybrid rice varieties infected with japonicaisolates showed more severe infection than those infected with indicaisolates. In other words, japonicaisolates more easily infected japonicaand indica-japonicahybrid rice varieties. The differences in quantity among the varieties indicated high resistance to rice blast in the corresponding kinds of rice inoculated with indicaand japonicaisolates, fully explaining the above viewpoint. Most of the 23 indicaisolates belonged to indicaraces. Among the 23 japonicaisolates, half were indicaraces and half were japonicaraces. In addition, there were significant differences between the isolates selected in the first two years and those selected in the second two years. The isolates in the first two years were all of indicaraces, and those of japonicaraces all appeared in the second two years. The results showed that the race of isolates shared little association with the kind of rice variety collected but was associated with the year of neck blast sample collection.

ABSTRACTPeroxisomes are ubiquitous organelles in eukaryotic cells that fulfill multiple important metabolisms. Pex13 and Pex14 are key components of the peroxisomal docking complex in yeasts and mammals. In the present work, we functionally characterized the homologues of Pex13 and Pex14 (Mopex13 and Mopex14) in the rice blast fungus Magnaporthe oryzae. Mopex13 and Mopex14 were peroxisomal membrane distributed and were both essential for the maintenance of Mopex14/17 on the peroxisomal membrane. Mopex13 and Mopex14 interacted with each other, and with Mopex14/17 and peroxisomal matrix protein receptors. Disruption of Mopex13 and Mopex14 resulted in a cytoplasmic distribution of peroxisomal matrix proteins and the Woronin body protein Hex1. In the ultrastructure of Δmopex13 and Δmopex14 cells, peroxisomes were detected on fewer occasions, and the Woronin bodies and related structures were dramatically affected. The Δmopex13 and Δmopex14 mutants were reduced in vegetative growth, conidial generation and mycelial melanization, in addition, Δmopex13 showed reduced conidial germination and appressorial formation and abnomal appressorial morphology. Both Δmopex13 and Δmopex14 were deficient in appressorial turgor and nonpathogenic to their hosts. The infection failures in Δmopex13 and Δmopex14 were also due to their reduced ability to degrade fatty acids and to endure reactive oxygen species and cell wall-disrupting compounds. Additionally, Mopex13 and Mopex14 were required for the sexual reproduction of the fungus. These data indicate that Mopex13 and Mopex14, as key components of the peroxisomal docking complex, are indispensable for peroxisomal biogenesis, fungal development and pathogenicity in the rice blast fungus.

Two cDNA libraries enriched for transcripts differentially expressed in plants of two rice lines with similar genetic backgrounds and same leaf blast resistance but different responses to neck blast using suppression subtractive hybridization (SSH). After differential screening and sequence analysis of the selected clones, 90 unique cDNA clones were found, of which 74 clones were with known functions according to the putative functions of their homologous genes in the database. They may be involved in pathogen response, signal transduction, transcription, etc. Expression differences of 17 out of the 26 selected cDNA clones in resistant and susceptible lines were confirmed by RT-PCR. Expression profilings of the 26 cDNA clones at the early stages after inoculation were also revealed by RT-PCR. This is the first report on the rice neck blast resistance at mRNA level and will facilitate the further study of genetic mechanism of neck blast resistance.

An F8recombinant inbred population was constructed using a commercial indicarice variety Zhong 156 as the female parent and a semidwarf indicavariety Gumei 2 with durable resistance to rice blast as the male parent. Zhong 156 is resistant to the fungus race ZC15at the seedling stage but susceptible to the same race at the flowering stage. Gumei 2 is resistant to ZC15at both stages. The blast resistance of 148 recombinant inbred lines was evaluated using the blast race ZC15. Genetic analysis indicated that the resistance to leaf blast was controlled by three genes and the presence of resistant alleles at any loci would result in resistance. One of the three genes did not have effects at the flowering stage. Two genes, tentatively assigned as Pi24(t)and Pi25(t), were mapped onto chromosome 12 and 6,respectively, based on RGA (resistance gene analog), RFLP and RAPD markers. Pi24(t)conferred resistance to leaf blast only, and its resistance allele was from Zhong 156. Pi25(t)conferred resistance to both leaf and neck blast, and its resistance allele was from Gumei 2. In a natural infection test in a blast hot-spot, Pi25(t)exhibited high resistance to neck blast, while Pi24(t)showed little effect.