Journal of the Korea Institute of Military Science and Technology (한국군사과학기술학회지)

The Korea Institute of Military Science and Technology (한국군사과학기술학회)
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Microbial Forensics: Comparison of MLVA Results According to NGS Methods, and Forensic DNA Analysis Using MLVA

미생물법의학: 차세대염기서열분석 방법에 따른 MLVA 결과 비교 및 이를 활용한 DNA 감식

  • Hyeongseok Yun (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Seungho Lee (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Seunghyun Lim (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Daesang Lee (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Sehun Gu (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Jungeun Kim (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Juhwan Jeong (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Seongjoo Kim (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Gyeunghaeng Hur (Defense Advanced Science and Technology Research Institute, Agency for Defense Development) ;
  • Donghyun Song (Defense Advanced Science and Technology Research Institute, Agency for Defense Development)
  • 윤형석 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 이승호 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 임승현 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 이대상 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 구세훈 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 김정은 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 정주환 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 김성주 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 허경행 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터) ;
  • 송동현 (국방과학연구소 국방첨단과학기술연구원 Chem-Bio센터)
  • Received : 2024.02.01
  • Accepted : 2024.05.08
  • Published : 2024.08.05
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Abstract

Microbial forensics is a scientific discipline for analyzing evidence related to biological crimes by identifying the origin of microorganisms. Multiple locus variable number tandem repeat analysis(MLVA) is one of the microbiological analysis methods used to specify subtypes within a species based on the number of tandem repeat in the genome, and advances in next generation sequencing(NGS) technology have enabled in silico anlysis of full-length whole genome sequences. In this paper, we analyzed unknown samples provided by Robert Koch Institute(RKI) through The United Nations Secretary-General's Mechanism(UNSGM)'s external quality assessment exercise(EQAE) project, which we officially participated in 2023. We confirmed that the 3 unknown samples were B. anthracis through nucleic acid isolation and genetic sequence analysis studies. MLVA results on 32 loci of B. anthracis were analysed by using genome sequences obtained from NGS(NextSeq and MinION) and Sanger sequencing. The MLVA typing using short-reads based NGS platform(NextSeq) showed a high probability of causing assembly error when a size of the tandem repeats was grater than 200 bp, while long-reads based NGS platform(MinION) showed higher accuracy than NextSeq, although insertion and deletion was observed. We also showed hybrid assembly can correct most indel error caused by MinION. Based on the MLVA results, genetic identification was performed compared to the 2,975 published MLVA databases of B. anthracis, and MLVA results of 10 strains were identical with 3 unkonwn samples. As a result of whole genome alignment of the 10 strains and 3 unknown samples, all samples were identified as B. anthracis strain A4564 which is associated with injectional anthrax isolates in heroin users.

Keywords

Acknowledgement

본 논문은 2024년 정부의 재원으로 국방과학연구소에서 수행한 연구결과임(912A01201).

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