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Inhibitory Effects of Antioxidant Reagent in Reactive Oxygen Species Generation and Penetration of Appressoria of Alternaria alternata Japanese Pear Pathotype

First, fourth, and fifth authors: Laboratory of Stress Cytology, Graduate School of Agricultural Science, and second author: Organization of Advanced Science and Technology, Kobe University, 1-1 Rokkodai, Nada, Kobe 657-8501, Japan; and third author: Laboratory of Nano-Structure Physiology, Okazaki Institute for Integrative Bioscience, 5-1 Higashiyama, Myodaiji, Okazaki, 444-8787, Japan.

In the Japanese pear pathotype of Alternaria alternata, H₂O₂ is generated solely from penetration pegs and not from other portions of subcuticular hyphae within the pectin layers of host leaves. A close association between H₂O₂ generation and fungal aggressiveness is expected because the pegs are important for fungal penetration into the host epidermis. To determine the potential role of reactive oxygen species in microbial pathogenicity, we studied the inhibitory effects of the antioxidant reagent ascorbic acid and the NADPH oxidase inhibitor diphenylene iodonium on infection of the pathogen. In our study, we showed H₂O₂ generation to be inhibited by inoculation with the mixture of ascorbic acid or diphenylene iodonium and spores at the pegs in the spore-inoculated host leaves. The decrease of generation in the pegs resulted in penetration failure, indicating that H₂O₂ generation probably contributed to strengthening of the penetration and probably was recruited by NADPH oxidase.