Doctoral Dissertation Research: Impact of Various Storage Temperatures andTtime on Microbiome of Forensically Relevant Biological Samples

博士论文研究：不同储存温度和时间对法医相关生物样本微生物组的影响

• 批准号: 2017519
• 负责人: Baneshwar Singh
• 金额: $ 3.05万
• 依托单位: Virginia Commonwealth University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2017519&HistoricalAwards=false
• 关键词: Doctoral; Dissertation; Research; Impact; Various

The confirmatory identification of forensically relevant body fluids is a fundamental part to forensic investigations as this information can aid investigators in crime scene reconstruction and resolving mixtures such as those often encountered in sexual assault cases. The majority of current body fluid identification methods are presumptive, and the sample consumption is not ideal for forensic casework. For new methods to be accepted into the community, methods should be robust, reliable, sensitive, and confirmatory to each body fluid while producing stable results over time. This study aims to determine how short- and long-term storage conditions of human biological samples impact the microbial community structures associated with these samples, and to determine whether these conditions impact the accuracy of a newly developed microbial signature-based method for the body fluid identification. Biological sample storage regulations were created without microbial evidence in mind, and it is therefore important to determine whether these samples are stable under various storage times and temperatures. This project will 1) look at the microbial signatures of each body fluid to determine whether there are significant differences in their compositions once stored at various temperatures at extended periods of time and 2) determine whether these storage conditions impact the accuracy of the previously developed body fluid identification method. Samples from ten individuals will be stored at four different temperatures (-20°C, 4°C, 25°C, 37°C) for 3 storage periods (30 days, 60 days, 90 days). To evaluate these changes, 16S rDNA dual-index high throughput sequencing will be performed using the MiSeq FGx sequencing platform. Data analysis will follow the mothur MiSeq SOP. The variation in bacterial signatures will be assessed between all temperature/time groups. Ensemble subspace classification and L1-norm regularized classification methods previously developed will be tested using the bacterial structures obtained to determine how storage conditions may impact the classification accuracy. The findings from this study will be useful in streamlining the previously developed body fluid identification method and will provide additional suggestions for standard operating procedures. This is not only useful for the forensic science community but can contribute to stability, standardization, and mobility of microbial research as a whole.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

确认法律相关的体液是法医投资的基本部分，因为这些信息可以帮助调查人员重建和解决诸如性侵犯案件中经常遇到的混合物。当前的大多数体液识别方法都是假定的，样品消耗并不是法医案例的理想选择。为了使新方法被接受到社区中，随着时间的流逝，方法应对每种体液进行稳健，可靠，敏感和确认，同时产生稳定的结果。这项研究旨在确定人类生物学样品的短期和长期存储条件如何影响与这些样品相关的微生物群落结构，并确定这些条件是否会影响新开发的基于微生物的基于微生物签名方法的准确性。生物样品储存法规是在没有微生物证据的情况下创建的，因此，确定这些样品在各种存储时间和温度下是否稳定很重要。该项目将1）查看每种体液的微生物特征，以确定一旦在长时间的各种温度存储在各种温度下，其组成是否存在显着差异，并且2）确定这些存储条件是否影响了先前开发的体液识别方法的准确性。来自十个个体的样本将在四个不同的温度（-20°C，4°C，25°C，37°C）中存储3个存储期（30天，60天，90天）。为了评估这些变化，将使用Miseq FGX测序平台进行16S rDNA双索引高吞吐测序。数据分析将遵循Mothur Miseq SOP。细菌特征的变化将在所有温度/时间组之间评估。集合子空间分类和先前开发的L1-norm正则分类方法将使用获得的细菌结构进行测试，以确定存储条件如何影响分类精度。这项研究的发现将有助于简化先前开发的体液识别方法，并为标准操作程序提供其他建议。这不仅对法医科学界有用，而且可以为整个微生物研究的稳定性，标准化和流动性做出贡献。该奖项反映了NSF的法定任务，并通过使用该基金会的智力优点和更广泛的影响来评估NSF的法定任务。