<i>MC69</i> is involved in appressorial penetration and pathogenicity of <i>M. oryzae</i>.

<p>(A) <i>MC69</i> is required for pathogenicity of <i>M. oryzae</i> strain Ina72. Conidial suspension of the wild-type strain Ina72 (Ina72 WT) and the <i>mc69</i> mutants (<i>mc69-9</i>, <i>-12</i>, <i>-87</i>) were inoculated on barley (cv. Nigrate) and rice (cv. Shin No. 2) leaves, and incubated for 4 and 7 days, respectively. (B) Germination, appressorium formation and appressorial penetration of Ina72 WT and the <i>mc69</i> mutants. The ratio of germination was calculated as the mean percentage of conidia germinated after 24 h on rice (cv. Shin No. 2) leaf sheath cells. The mean percentage of appressorium formation on rice leaf sheath cells among the germinated conidia is presented. Three replicates of ∼50 conidia were counted for each observation. The mean percentage of appressorial penetration by the <i>mc69</i> mutants 32 h after inoculation is presented. Standard errors are indicated by the vertical bars. (C) Appressorial penetration assays on rice leaf sheath cells. Conidia from Ina72 WT and <i>mc69-87</i> germinated and formed melanized appressoria. All appressoria formed by Ina72 WT penetrated and produced infectious hyphae but no appressoria formed by <i>mc69-87</i> produced infectious hyphae in most of area. Photographs were taken 32 h after inoculation. Scale bar = 40 µm. (D) Invasive growth rating of rice leaf sheath cells 32 h after inoculating with Ina72 WT and <i>mc69-87</i>. The levels for invasive growth rating are given above. For details of the invasive growth levels and rating see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002711#s4" target="_blank">Materials and Methods</a>. Scale bar = 20 µm. (E), (F) Complementation of <i>mc69</i> mutant with the wild type allele of <i>MC69</i>. (E) Appressorial penetration by Ina72 WT, <i>mc69-87</i> (<i>mc69</i>) and the <i>MC69</i> re-introduced strain (<i>mc69</i>+<i>MC69</i>). Mean percentage of appressorial penetration is recorded 32 h after inoculation in rice leaf sheath cells. Four replicates of ∼50 appressoria were counted for each observation. (F) Blast symptoms caused by Ina72 WT, <i>mc69</i> and <i>mc69</i>+<i>MC69</i> on barley (cv. Nigrate) and on rice (cv. Shin No 2) 3 and 4 days after inoculation, respectively.