Dou Shujie, Ma Guanju, Liang Yu, Shen Jie, Zhao Guangzhong, Fu Guangping, Fu Lihong, Cong Bin, Li Shujin
College of Forensic Medicine, Hebei Key Laboratory of Forensic Medicine, Research Unit of Digestive Tract Microecosystem Pharmacology and Toxicology, Hebei Medical University, Chinese Academy of Medical Sciences, No. 361 Zhongshan Road, Shijiazhuang, 050017, Hebei, China.
Hainan Tropical Forensic Medicine Academician Workstation, Haikou, China.
Int J Legal Med. 2025 May;139(3):1019-1030. doi: 10.1007/s00414-024-03383-0. Epub 2024 Dec 16.
Determining the time since deposition (TsD) and sex of saliva stains is crucial for revealing the time of the crime's occurrence and clarifying the nature of the crime. This process not only shortens the time required to solve the case but also helps narrow down the scope of investigation, thereby enhancing the efficiency of case resolution. Currently, the forensic study of the microbial composition in long-term saliva stains remains a relatively underexplored field. The purpose of this study was to explore the succession pattern of long-placed human saliva stains microbial communities and identify relevant microbial markers for estimating TsD and identifying the sex of the donor, in order to be an effective alternative tool for solving practical forensic cases. Therefore, in this study, saliva stains exposed to indoor environmental conditions for up to 140 days were collected and 16S rRNA full-length sequencing was performed using single-molecule real-time sequencing technology based on the PacBio sequencing platform. The study reveals that after 140 days of placement, the relative abundance of Firmicutes significantly decreased (p = 0.00304). At the genus level, the relative abundances of Streptococcus (p = 0.0008), Rothia (p = 0.0448), Gemella (p = 0.016), and Veillonella (p = 0.0208) also significantly decreased. Additionally, significant differences were found in the microbial communities between saliva stains from males and females (p = 0.00013). Then, we constructed a TsD estimating model for microbial community markers based on random forest, and the results showed that the mean absolute error was 9.59 days, and the accuracy of sex classification model based on stepwise logistic regression model and 4 bacterial markers was 84.21%. This indicates that saliva stains that have been in place for a long time still retain significant forensic value, and microbial markers can be used to determine the time since deposition (TsD) of dried saliva stains as well as to identify the sex of the donor.
确定唾液斑痕的沉积时间（TsD）和性别对于揭示犯罪发生时间和厘清犯罪性质至关重要。这一过程不仅缩短了破案所需时间，还有助于缩小调查范围，从而提高案件侦破效率。目前，长期唾液斑痕中微生物组成的法医学研究仍是一个相对未被充分探索的领域。本研究的目的是探索长期放置的人类唾液斑痕微生物群落的演替模式，并识别用于估计沉积时间和鉴定捐赠者性别的相关微生物标志物，以便成为解决实际法医学案件的有效替代工具。因此，在本研究中，收集了暴露于室内环境条件下长达140天的唾液斑痕，并基于PacBio测序平台使用单分子实时测序技术进行了16S rRNA全长测序。研究表明，放置140天后，厚壁菌门的相对丰度显著下降（p = 0.00304）。在属水平上，链球菌属（p = 0.0008）、罗氏菌属（p = 0.0448）、孪生球菌属（p = 0.016）和韦荣球菌属（p = 0.0208）的相对丰度也显著下降。此外，男性和女性唾液斑痕的微生物群落存在显著差异（p = 0.00013）。然后，我们基于随机森林构建了微生物群落标志物的沉积时间估计模型，结果显示平均绝对误差为9.59天，基于逐步逻辑回归模型和4种细菌标志物的性别分类模型的准确率为84.21%。这表明长期存在的唾液斑痕仍保留着显著的法医学价值，微生物标志物可用于确定干燥唾液斑痕的沉积时间以及鉴定捐赠者的性别。
