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Species identification of white false hellebore (Veratrum album subsp. oxysepalum) by loop-mediated isothermal amplification (LAMP)

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Abstract

Purpose

Food poisoning is frequently caused by the accidental ingestion of toxic plants with strong morphological similarities to edible plants. The plant, false helleborine (Veratrum album), is commonly involved in such incidents. Cases of poisoning by toxic plants require rapid and accurate identification for appropriate medical treatment or forensic investigation in the event of death. However, morphological examinations require experience in systematic botany, and chemical analysis of natural compounds can be challenging. This study aimed to develop a sensitive, rapid, and specific method for identifying V. album and V. stamineum.

Methods

We developed and optimized a loop-mediated isothermal amplification (LAMP) detection method. Total genomic DNA was extracted from approximately 10 mg (dry weight) of leaf from 26 specimens comprising the target plants, allied plants, and other causative plants of poisoning. These samples were analyzed the LAMP assay, and mixed samples, cooked samples, and simulated gastric contents were also tested.

Results

The limit of detection of our LAMP assay was 0.08 ng of target DNA, and the assay was specific to V. album and V. stamineum. The LAMP assay could be performed in approximately 30 min. V. album was also successfully identified in mixed samples, cooked samples, and simulated gastric contents. In addition, the LAMP assay signal could be effectively assessed with the naked eye.

Conclusions

The present LAMP assay is a highly sensitive, rapid, and specific method for identifying V. album and V. stamineum.

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Acknowledgements

The authors thank Prof. Kiyotaka Koyama (Meiji Pharmaceutical University), Dr. Tatsuhiko Suzuki (Tokyo University of Science), and Nikko Botanical Garden for providing the plant samples used in this study. The authors thank Mr. Kenichi Sato and Mr. Jun Kimura of the Nippon Gene Corporation for technical advice on Genie II and primer design. We also thank Mr. Shigehiko Miyamoto of Kaneka for assisting with LAMP assay visualization. The authors are grateful to Dr. Takuya Aikawa of the Tohoku Research Center, Forestry and Forest Products Research Institute, and Mr. Masashi Kitamura of the Forensic Science Laboratory, Ishikawa Prefectural Police H.Q. for technical advice on the LAMP assay. The authors are also grateful to Mr. Satoshi Kubota for technical advice on preparing the simulated gastric samples and Dr. Ritsuko Sugita for her invaluable advice regarding this study. This work was supported by JSPS KAKENHI grant nos. 20392269 and 17K15886.

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Authors and Affiliations

  1. National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa, Chiba, 277-0882, Japan

    Hitomi S. Kikkawa & Kouichiro Tsuge

  2. Medical Plant Garden, Tokyo Metropolitan Institute of Public Health, 21-1 Nakajimacho, Kodaira, Tokyo, 187-0033, Japan

    Masako Aragane

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  1. Hitomi S. Kikkawa
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Correspondence to Hitomi S. Kikkawa.

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Kikkawa, H.S., Aragane, M. & Tsuge, K. Species identification of white false hellebore (Veratrum album subsp. oxysepalum) by loop-mediated isothermal amplification (LAMP). Forensic Toxicol 37, 308–315 (2019). https://doi.org/10.1007/s11419-018-00461-y

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