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The vertical distribution of leaf blast lesions caused by the fungus Pyricularia grisea was studied to estimate the degree of leaf blast suppression in rice multilines in experimental paddy fields for 4 years. Leaf blast in 1 : 1 and 1 : 3 mixtures of susceptible rice cultivar Sasanishiki and its resistant near-isogenic line, Sasanishiki BL7, developed slower than that in pure stands of Sasanishiki. The average distance of lesions on leaves from the ground in the 1 : 3 mixtures was significantly lower than that in the pure stands at the end of leaf blast epidemics (at booting stage). This result shows that the distribution of leaf blast lesions in the upper layer differs between the susceptible pure stands and the 1 : 3 mixtures at the end of leaf blast epidemics.

To investigate the frequency distribution of purple seed stain of soybean caused by Cercospora kikuchii in two experimental fields in 2004, we set up rows 75 cm apart and sowed two asymptomatic seeds at each of positions 20 cm apart in each row. We sowed purple-stained seeds infected with the pathogen as inocula at four points instead of asymptomatic seeds in each field. We assessed disease incidence in harvested seeds by counting the numbers of purple-stained and asymptomatic seeds. To determine the spatial distribution of the disease, we grouped the field points into analytical units of several sizes. Beta-binomial and binomial distributions described the distribution patterns of purple-stained seeds. The smallest value of α, a beta-binomial parameter, occurred with analytical units that contained three or nine points next to each other within a single row, suggesting that these units showed the most aggregated distribution of the disease, each of the patches of seeds infected with C. kikuchii could be defined approximately by the area covered by three or nine points (75 × 60 or 75 × 180 cm), and the disease tended to infect plants next to each other within rows.

Leaf blast suppression in multilines was evaluated based on the number of susceptible lesions observed in a pure stand of susceptible rice cultivar Sasanishiki, and in 1 : 1 and 1 : 3 mixtures of Sasanishiki and a resistant near-isogenic line, Sasanishiki BL4 or BL7, from 1998 to 2001. The number of lesions first observed in fields in the 1 : 1 and 1 : 3 mixtures were close to theoretical numbers calculated using the number of lesions observed in the pure stands and the ratios of the susceptible Sasanishiki in the mixtures. The ratio of the number of lesions in the 1 : 1 and 1 : 3 mixtures to the number in the pure stand was 0.29 and 0.09, respectively. The relationship between these ratios and the ratios of susceptible Sasanishiki in mixtures was defined in an equation to estimate the degree of leaf blast suppression. Validation studies for the ratios of the number of lesions in the 1 : 1 and 1 : 3 mixtures to the number in the pure stand were conducted in two different locations and showed that the ratios are almost acceptable. The calculated autoinfection to alloinfection ratio was 1.3 and 1.4 in the 1 : 1 and 1 : 3 mixtures, respectively, suggesting that the calculated ratio will affect the degree of leaf blast suppression. Thus, predictors were obtained to estimate leaf blast suppression for effective blast control in multilines.

Azoxystrobin was recently registered in Japan for the control of purple seed stain of soybean caused by Cercospora kikuchii, because the pathogen has developed resistance to thiophanate-methyl. To investigate the effects of these fungicides on the frequency of C. kikuchii strains resistant to thiophanate-methyl and on the genotype structure of the population, we sowed purple-stained seeds, approximately 40% of which were infected with resistant strains, as inocula with asymptomatic seeds and applied thiophanate-methyl and azoxystrobin during the reproductive growth of soybeans. The isolation frequency of resistant strains increased more than 99% by thiophanate-methyl but was not significantly increased by azoxystrobin. In amplified fragment length polymorphism (AFLP) DNA fingerprinting, genotypic diversity was significantly decreased by thiophanate-methyl but was not affected by azoxystrobin. In addition, the similarity of the AFLP genotype structure was increased by thiophanate-methyl but not by azoxystrobin. These results suggest that thiophanate-methyl selectively inhibited the proliferation of sensitive strains, which resulted in a small number of genotypes, most of which were resistant strains. Azoxystrobin was found to nonselectively inhibit proliferation of the pathogen, which retained a large number of genotypes including thiophanate-methyl-sensitive or thiophanate-methyl-resistant strains or both.

We collected 247 isolates of Cercospora kikuchii from soybean seeds with typical purple stain symptoms from 15 prefectures in Japan. Of the 247 isolates, 93 were sensitive to thiophanate-methyl, a benzimidazole used to control this soybean disease; the remaining 154 were highly resistant to the fungicide. To examine genetic variability among the population of 247 isolates, we developed amplified fragment length polymorphism (AFLP) markers. An AFLP primer pair generated DNA fingerprint polymorphisms among the sample isolates, and with the unweighted pair-grouping method to cluster arithmetic means of the similarity coefficients among all pairs of the fingerprint patterns, the isolates were divided into four lineages (I to IV). Of the 247 isolates, 225 belonged to lineage I, including all isolates that were resistant to thiophanate-methyl. To determine whether the resistance of these isolates was related to mutations in the β-tubulin gene, we amplified partial nucleotide sequences of the gene from 29 representative isolates, including 12 that were resistant to thiophanate-methyl, by means of the polymerase chain reaction. The resistant isolates had identical nucleotide sequence with a one-step change at codon 198, in which the amino acid glutamic acid had been replaced by alanine. The evidence thus suggests that thiophanate-methyl resistance might have arisen in lineage I, the largest of the four lineages.

The japonica rice (Oryza sativa) cultivar Chubu 32 has a high level of partial resistance to blast, which is mainly controlled by a dominant resistance gene located on chromosome 11. The partial resistance to the rice blast fungus (Magnaporthe grisea) in Chubu 32 has isolate specificity; isolate IBOS8-1-1 is more aggressive on Chubu 32 than are other isolates. We hypothesized that the gene-for-gene relationship fits this case of a partial resistance gene in Chubu 32 against the avirulence gene in the pathogen. The partial resistance gene in Chubu 32 was mapped between DNA markers C1172 (and three other co-segregated markers) and E2021 and was designated Pi34. In the 32 F3 lines from the cross between a chromosome segment substitution line (Pi34−) from Koshihikari/Kasalath and Chubu 32, the lines with high levels of partial resistance to the M. grisea isolate Y93-245c-2 corresponded to the presence of Pi34 estimated by graphic genotyping. This indicated that Pi34 has partial resistance to isolate Y93-245c-2 in compatible interactions. The 69 blast isolates from the F1 progeny produced by the cross between Y93-245c-2 and IBOS8-1-1 were tested for aggressiveness on Chubu 32 and rice cultivar Koshihikari (Pi34−). The progeny segregated at a 1 : 1 ratio for strong to weak aggressiveness on Chubu 32. The results suggested that Y93-245c-2 has one gene encoding avirulence to Pi34 (AVRPi34), and IBOS8-1-1 is extremely aggressive on Chubu 32 because of the absence of AVRPi34. This is the first report of a gene-for-gene relationship between a fungal disease resistance gene associated with severity of disease and pathogen aggressiveness.