Comparative analysis of Sanger and next generation sequencing methods for 16S rDNA analysis of post-mortem specimens
Comparative analysis of Sanger and next generation sequencing methods for 16S rDNA analysis of post-mortem specimens
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DOI：
10.1080/00450618.2017.1402957
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发表时间：
2019-07
影响因子：
1
通讯作者：
Yoonjung Cho;M. H. Lee;H. Kim;Moonhee Park;Min-Hee Kim;Hansol Kwon;Jong-Bae Kim;Yang Han Lee;Dong Sub Lee
中科院分区：
文献类型：
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作者：
Yoonjung Cho;M. H. Lee;H. Kim;Moonhee Park;Min-Hee Kim;Hansol Kwon;Jong-Bae Kim;Yang Han Lee;Dong Sub Lee
摘要法医微生物学被广泛应用于在犯罪现场鉴定人的身份，并通过检验微量证据来确定死因和死亡时间。特别是，从死后标本中鉴定细菌非常重要，因为死亡原因可能是临床显著的细菌感染，导致致命的结果，如败血症。应用生物系统公司的MicroSEQ 500 16S rDNA细菌鉴定系统（MicroSEQ ID）是一种基于细菌16S rDNA基因Sanger测序的分子技术，用于通过培养分离细菌的分析。MiSeq Illumina平台使用下一代测序（NGS）进行16S rDNA的宏基因组分析和细菌的培养独立鉴定。在这项研究中，我们用这两种方法分析了65个死后标本，并比较了方法、时间效率、成本效率和性能方面的工作流程，以确定这些技术在法医应用中的局限性和优势。我们发现MiSeq更具时间效率，并且在分析超过30个样品时表现出优越的成本效益。此外，MiSeq与MicroSeq ID相比还有许多其他优势，包括更简单地鉴定难以培养的细菌，以及使用更大的16S rDNA文库，从而更精确地鉴定细菌种类。
Abstract Forensic microbiology is widely applied to identify people at crime scenes and determine the cause and time of deaths by examination of trace evidence. In particular, identification of bacteria from post-mortem specimens is important because the cause of death could be an infection with clinically significant bacteria that results in a fatal outcome, such as sepsis. The MicroSEQ 500 16S rDNA Bacteria Identification system (MicroSeq ID) from Applied Biosystems is a molecular technique for profiling of bacteria isolated by cultures, based on Sanger sequencing of the bacterial 16S rDNA gene. The MiSeq Illumina platform uses next-generation sequencing (NGS) for metagenomic analysis of 16S rDNA and culture-independent identification of bacteria. In this study, we analysed 65 post-mortem specimens by both methods and compared the workflows involved with regard to methodology, time-efficiency, cost-efficiency, and performance to determine the limitations and benefits of these techniques for forensic application. We found that MiSeq is more time-efficient, and exhibits superior cost-efficiency when more than 30 samples are analysed. In addition, MiSeq has a number of other advantages over MicroSeq ID, including a simpler identification of bacteria that are difficult to culture and use of a larger library of 16S rDNA, allowing more precise identification of bacterial species.