Identification of Mushroom Species by Automated rRNA Intergenic Spacer Analysis (ARISA) and Its Application to a Suspected Case of Food Poisoning with <i>Tricholoma ustale</i>

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  • SUGAWARA Ryota
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity
  • YAMADA Sayumi
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity
  • TU Zhihao
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity
  • SUGAWARA Akiko
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity
  • HOSHIBA Toshihiro
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity
  • EISAKA Sadao
    Sapporo City, Kiyota Ward Office, Health & Welfare Department
  • YAMAGUCHI Akihiro
    Department of Food Science and Human Wellness, Rakuno Gakuen Uninversity

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Other Title
  • 自動rRNA遺伝子間多型解析（ARISA）によるキノコ種鑑別の検討とカキシメジによる食中毒疑い事例への適用の試み
  • ジドウ rRNA イデンシ カン タケイ カイセキ(ARISA)ニ ヨル キノコシュ カンベツ ノ ケントウ ト カキシメジ ニ ヨル ショクチュウドク ウタガイ ジレイ エ ノ テキヨウ ノ ココロミ

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Automated rRNA intergenic spacer analysis (ARISA), a method of microbiome analysis, was evaluated for species identification of mushrooms based on the specific fragment sizes. We used 51 wild mushroom-fruiting bodies collected in the centre of Hokkaido and two cultivated mushrooms. Samples were hot-air-dried and DNA were extracted by a beads beating procedure. Sequencing analysis of portions of the rRNA gene (rDNA) provided 33 identifications of mushrooms by genus or species. The results of ARISA identification based on the combination of the fragment sizes corresponding to two inter spacer regions (ITS2 and ITS1) of rDNA within±0.1% accuracy showed that 27 out of the 33 species had specific fragment sizes differentiated from other species. The remaining 6 species formed 3 pairs that showed overlapping fragment sizes. In addition, within-species polymorphisms were observed as 1 bp differences among 32 samples of 13 species. ARISA was applied to investigate a case of suspected food poisoning in which the mushroom was thought to be a toxic Kakishimeji. The morphological identification of the mushroom was ambiguous since the remaining sample lacked a part of the fruiting body. Further, yeast colonies had grown on the surface of the fruiting body during storage. The ARISA fragment size of the mushroom showed 7 bp difference from that of the candidate toxic mushroom. Although ARISA could be a useful tools for estimation of mushroom species, especially in case where the fruiting bodies have deteriorated or been processed, further studies are necessary for reliable identification. For example, it may be necessary to adopt more informative genes which could provide clearer species-specific polymorphisms than the ITS regions.

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