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International Journal of Legal Medicine

, Volume 133, Issue 4, pp 1065–1073 | Cite as

Rapid identification of Gloriosa superba and Colchicum autumnale by melting curve analysis: application to a suicide case involving massive ingestion of G. superba

  • Makoto SakuradaEmail author
  • Naoki Yoshioka
  • Azumi Kuse
  • Kanako Nakagawa
  • Mai Morichika
  • Motonori Takahashi
  • Takeshi Kondo
  • Migiwa Asano
  • Yasuhiro Ueno
Original Article
  • 23 Downloads

Abstract

The plant species Gloriosa superba and Colchicum autumnale produce extremely poisonous colchicine as a major toxic metabolite. Almost all previous studies on colchicine poisoning have focused on drug analysis and clinical and pathological aspects. In this study, we developed a rapid, highly sensitive method to identify G. superba and C. autumnale. This method, which can distinguish between G. superba and C. autumnale using even minute amounts of plant material, is based on duplex real-time PCR in combination with melting curve analysis. To discriminate between the two genera of colchicine-containing plants, we designed new primer pairs targeting the region of the ycf15 gene, which is present in C. autumnale but not G. superba. By producing PCR amplicons with easily distinguishable melting temperatures, we were able to rapidly and accurately distinguish G. superba from C. autumnale. The new primer pairs generated no PCR amplicons from commercially available human DNA or various plant DNAs except for G. superba and C. autumnale. Sensitivity testing indicated that this assay can accurately detect less than 0.031 ng of DNA. Using our method in conjunction with colchicine drug analysis, we successfully identified G. superba in the stomach contents of a suicide victim who ingested massive quantities of a colchicine-containing plant. According to these results, duplex real-time PCR analysis is very appropriate for testing forensic samples, such as stomach contents harboring a variety of vegetables, and enables discrimination between G. superba and C. autumnale in forensic and emergency medical fields.

Keywords

Real-time PCR Melting curve analysis Colchicine Gloriosa superba Colchicum autumnale Forensic analysis of stomach contents 

Notes

Acknowledgments

We thank Edanz Group (www.edanzediting.com/ac) for editing the English text of a draft of this manuscript.

Compliance with ethical standards

Informed consent was obtained, and autopsy sample collection was conducted with the permission of the Ethics Committee of the Kobe University Graduate School of Medicine (#1799).

Conflict of interest

The authors declare that they have no conflicts of interest.

Supplementary material

414_2019_2060_MOESM1_ESM.docx (243 kb)
ESM 1 (DOCX 242 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Division of Legal MedicineKobe University Graduate School of MedicineKobeJapan
  2. 2.Forensic Science LaboratoryHyogo Prefectural Police HeadquartersKobeJapan
  3. 3.Hyogo Prefectural Institute of Public Health ScienceKakogawaJapan
  4. 4.Division of Legal MedicineEhime University Graduate School of MedicineToonJapan

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