Evaluating Bioinformatic Pipeline Performance for Forensic Microbiome Analysis*,†,‡

Microbial communities have potential evidential utility for forensic applications. However, bioinformatic analysis of high-throughput sequencing data varies widely among laboratories. These differences can potentially affect microbial community composition and downstream analyses. To illustrate the importance of standardizing methodology, we compared analyses of postmortem microbiome samples using several bioinformatic pipelines, varying minimum library size or minimum number of sequences per sample, and sample size. Using the same input sequence data, we found that three open-source bioinformatic pipelines, MG-RAST, mothur, and QIIME2, had significant differences in relative abundance, alpha-diversity, and beta-diversity, despite the same input data. Increasing minimum library size and sample size increased the number of low-abundant and infrequent taxa detected. Our results show that bioinformatic pipeline and parameter choice affect results in important ways. Given the growing potential application of forensic microbiology to the criminal justice system, continued research on standardizing computational methodology will be important for downstream applications.     

Volatile Organic Compound Profiling from Postmortem Microbes using Gas Chromatography–Mass Spectrometry

Volatile organic compounds (VOCs) are by-products of cadaveric decomposition and are responsible for the odor associated with decomposing remains. The direct link between VOC production and individual postmortem microbes has not been well characterized experimentally. The purpose of this study was to profile VOCs released from three postmortem bacterial isolates (Bacillus subtilis, Ignatzschineria indica, I. ureiclastica) using solid-phase microextraction arrow (SPME Arrow) and gas chromatography–mass spectrometry (GC-MS). Species were inoculated in headspace vials on Standard Nutrient Agar and monitored over 5 days at 24°C. Each species exhibited a different VOC profile that included common decomposition VOCs. VOCs exhibited upward or downward temporal trends over time. Ignatzschineria indica produced a large amount of dimethyldisulfide. Other compounds of interest included alcohols, aldehydes, aromatics, and ketones. This provides foundational data to link decomposition odor with specific postmortem microbes to improve understanding of underlying mechanisms for decomposition VOC production.     

大鼠肝细胞DNA含量与早期死亡时间关系的图像分析研究

实验选择 15只大鼠 ,处死后 ,在 2 4h内 ,每隔 1h取肝组织块进行细胞学涂片、福尔马林液固定、Feul gen染色、自动图像分析仪测量、统计学处理数据。结果表明 ,肝细胞DNA含量在死后 2 4h内 ,随死亡时间的延长而较有规律的下降。其中积分光密度、平均光密度、异形指数是用来研究死亡 2 4h内肝细胞DNA降解规律 ,并准确推断死亡时间的较好指标。     

家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化

目的探讨急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化。方法用1%牛胆酸钠诱导家兔急性胰腺炎动物模型建立,于死后不同时间和正常对照组,观察其淀粉酶变化。结果家兔急性胰腺炎死后72h眼玻璃体液淀粉酶含量(x)与死亡时间(y)存在相关关系,并导出其二项式回归方程为y=8.7420 0.7699x-0.0083x2(R2=92.62792,F=14.89734,P=0.001)。可作为推定早期损伤时间的参考指标。结论家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶改变为法医早期死亡时间推断提供了灵敏客观的实验依据。     

家兔死后眼玻璃体液锌、镍含量与 PMI关系的研究

目的寻找一种精确推定PMI的方法。方法应用电感耦合等离子体质谱（ICP－MS）,检测了家兔死后96小时内眼玻璃体液锌、镍元素含量,探讨其与PMI的相关性。结果家兔死后24h内眼玻璃体液锌、镍元素含量与PMI显著相关,可作为推定早期PMI的参考指标,其二项式回归方程分别为y=0.1404x2－1.3351x＋3.8298（R2=0.9202）、y=0.0043x2－0.0596x＋0.2665（R2=0.9103）。结论家兔眼玻璃体液中Zn、Ni元素含量变化是推定早期PMI的参考指标之一。     

利用DNA和RNA进行死亡时间推断的研究进展

死亡时间是法医和刑事侦查工作者在调查犯罪行为和其他死亡事件中所需要的重要信息之一。但是目前死亡时间推断所使用的传统方法常受到经验和环境的影响，从而降低推断的准确性。由于DNA和某些RNA在细胞内的含量十分稳定，而且个体死亡后这些遗传物质随着死亡时间的延长呈有规律地降解，所以通过测定死后细胞内的DNA和RNA含量推断死亡时间成为法医学的研究热点。     

Dental hygiene students' matching accuracy when comparing antemortem dental radiographs and oral photographs to simulated postmortem WinID3® odontograms

Matching dental antemortem (AM) and postmortem (PM) data for human identification is especially challenging when the workforce is limited. Dental hygienists have served mass fatality incidents (MFIs) due to dental-related expertise. However, forensics within dental hygiene education and research on transferable skills is limited. This qualitative balance design study assessed senior dental hygiene students' match accuracy of simulated cases varying in dental identifiers based on AM full mouth series (FMS) radiographs and oral photographs to PM WinID3^® odontograms to demonstrate possible disaster victim identification (DVI) transferable skills gained during formal education. A convenience sample of senior dental hygiene students (n = 31) was presented information on WinID3^® interpretation, then presented with 5 mismatched cases and asked to visually interpret each to make 10 total matches; five based on AM FMS with simulated PM WinID3^® odontograms and five based on AM photographs with PM WinID3^® odontograms. Match accuracy scores ranged from 41.9% to 58.1% for cases with 1–10 identifiers, and 77.4% to 93.5% for cases with 11–40 identifiers. Accuracy when matching AM radiographs to PM odontograms versus AM photographs to PM odontograms was compared and revealed no statistical differences in match accuracy depending on image type (p = 0.388 to 1.000). Results of this pilot study suggests transferable match accuracy skills resulted from the participants' dental hygiene formal education. These baseline skills with additional specialized training support the rationale for dental hygienists serving on DVI teams. More research is needed in education and practice when preparing dental hygienists for forensic-based service.     

死后眼组织变化与死亡时间推断

眼组织推断死亡时间一直是国内外法医学界的研究热点之一,从肉眼观到镜下观,从细胞水平到分子水平,不断有新的研究成果和报告出现。大量研究资料表明,随着死亡时间的延长,眼角膜、视网膜、玻璃体液及房水等均出现规律性的变化,并与死亡时间高度相关,可用于死亡时间推断。本文对其在法医学领域的研究进展作以综述。     

甲维盐中毒死亡小鼠体内的分布及死后再分布

目的研究甲氨基阿维菌素苯甲酸盐(简称甲维盐)中毒死亡小鼠的致死血浓度、靶器官组织、毒物蓄积库和死后毒物再分布的特征。方法采用灌胃法建立中毒小鼠模型,动态观察急性中毒组、亚急性中毒组小鼠的主要中毒症状和临床死亡时间。观察中毒死后小鼠各器官组织病理形态学改变,应用酶联免疫吸附试验测定死后0、24、48、72h甲维盐体内分布及死后再分布,采用高效液相色谱法测定中毒小鼠的致死血浓度和死后各时间点的血中甲维盐浓度。结果中毒小鼠均在灌胃后15~30 min内依次出现神经、呼吸系统症状。急性中毒组小鼠的临床死亡时间为(45.8±7.9)min,亚急性中毒组为(8.0±1.4)d。甲维盐的急性致死血浓度范围为447.164 0~524.463 5 mg/L。光镜及荧光显微镜下各器官组织均见明显的病理改变;小鼠中毒死后72 h内,血、心、肝、脾、肺、肾及脑甲维盐浓度变化具有规律性(P0.05)。结论甲维盐中毒作用的主要靶器官为心、肝、肾、肺、脑和接触部位(胃),其主要蓄积库为肾、肝,甲维盐在小鼠体内存在死后再分布现象。     

Fatal Fentanyl Patch Misuse in a Hospitalized Patient with a Postmortem Increase in Fentanyl Blood Concentration

Opioid‐related mortality happens, even in healthcare settings. We describe serial postmortem fentanyl blood concentrations in a hospital inpatient who fatally abused transdermal fentanyl. This is a single‐patient case report. A 42‐year‐old man with lymphoma was started on transdermal fentanyl therapy while hospitalized for chronic abdominal pain. The patient was last seen awake 1.3 h prior to being found apneic and cyanotic. During the resuscitation attempt, a small square‐shaped film was removed from the patient's oropharynx. Femoral blood was collected 0.5 and 2 h postmortem, and the measured fentanyl concentration increased from 1.6 to 14 ng/mL. Study limitations include potential laboratory or collection errors and missing data. (i) Providers must be vigilant for signs of fentanyl patch abuse. (ii) Postmortem blood concentrations are not static postmortem, likely secondary to decreasing pH, increased aqueous solubility, and tissue redistribution, and are therefore unlikely to accurately represent antemortem blood concentrations.