Harnessing the potential of the environmental microbiome in forensic science

2021 ◽  
pp. 61-67
Author(s):  
Agata Jagiełło ◽  
◽  
Anna Woźniak ◽  
Błażej Szczerba ◽  
Rafał Płoski ◽  
...  
Keyword(s):  
Forensic Science ◽  
Dna Analysis ◽  
Genetic Test ◽  
Massively Parallel Sequencing ◽  
Geographical Location ◽  
Soil Samples ◽  
Massively Parallel ◽  
Soil Microbiome ◽  
Microbiome Analysis ◽  
Dna Profiles

A scientific consortium led by the Central Forensic Laboratory of the Police has undertaken to develop a method for DNA analysis of the soil microbiome to be used in forensic investigations. The aim of the project entitled Soil Microbiome Analysis Forensic Tool – SMAFT (http://smaft.eu/), financed by the National Center for Research and Development (DOB-BIO10/03/01/2019), is to develop a new tool that enables the association of a trace in the form of a soil sample with a specific geographical location. The first part of the paper introduces the concept of the microbiome and presents the possibilities of using microbiome DNA analysis in forensic science. In the second part, the stages of the SMAFT project are described in detail, beginning from the collection of soil samples from different sites in Poland across all seasons and isolation of microbiome DNA through massively parallel sequencing (MPS) technology-based analysis of isolates and the development of a genetic test containing a set of metagenomic markers allowing for effective individualization of soil samples, up to the creation of an IT system enabling analysis and interpretation of the obtained results, which includes a database of soil microbiome DNA profiles from various locations in Poland.

scholarly journals Impact of the Human Microbiome in Forensic Sciences: a Systematic Review

2020 ◽  
Vol 86 (22) ◽  
Author(s):  
Manuel G. García ◽  
María D. Pérez-Cárceles ◽  
Eduardo Osuna ◽  
Isabel Legaz
Keyword(s):  
Systematic Review ◽  
Forensic Science ◽  
Human Microbiome ◽  
Geographical Location ◽  
Postmortem Interval ◽  
Forensic Sciences ◽  
Content Type ◽  
Microbiome Analysis ◽  
Location Of Death ◽  
Microbiome Research

ABSTRACT Numerous studies relate differences in microbial communities to human health and disease; however, little is known about microbial changes that occur postmortem or the possible applications of microbiome analysis in the field of forensic science. The aim of this review was to study the microbiome and its applications in forensic sciences and to determine the main lines of investigation that are emerging, as well as its possible contributions to the forensic field. A systematic review of the human microbiome in relation to forensic science was carried out by following PRISMA guidelines. This study sheds light on the role of microbiome research in the postmortem interval during the process of decomposition, identifying death caused by drowning or sudden death, locating the geographical location of death, establishing a connection between the human microbiome and personal items, sexual contact, and the identification of individuals. Actinomycetaceae, Bacteroidaceae, Alcaligenaceae, and Bacilli play an important role in determining the postmortem interval. Aeromonas can be used to determine the cause of death, and Corynebacterium or Helicobacter pylori can be used to ascertain personal identity or geographical location. Several studies point to a promising future for microbiome analysis in the different fields of forensic science, opening up an important new area of research.

Identifying cancer mutations in neuroendocrine prostate cancer (NEPC) through massively parallel DNA sequencing of formalin-fixed paraffin-embedded (FFPE) tissue.

2012 ◽  
Vol 30 (5_suppl) ◽  
pp. 110-110
Author(s):  
Himisha Beltran ◽  
Kyung Park ◽  
Scott T. Tagawa ◽  
Roman Yelensky ◽  
Doron Lipson ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Dna Sequencing ◽  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Ffpe Tissue ◽  
Massively Parallel ◽  
Sequence Coverage ◽  
High Confidence ◽  
Biopsy Material ◽  
Synonymous Mutations

110 Background: NEPC is an aggressive and lethal variant of prostate cancer that most commonly arises from existing prostate adenocarcinoma (PCA). Little is known about the underlying biology of NEPC, as metastases are rarely biopsied. The purpose of this study is to determine the spectrum of somatic mutations in NEPC by using a novel DNA sequencing platform and to evaluate for targetable molecular alterations. Methods: 44 NEPC and mixed NEPC/PCA were evaluated and 77 high density foci of NEPC, PCA, and benign areas were selected for DNA extraction. Massively parallel sequencing via HiSeq2000 was performed using indexed seq libraries constructed by adapter ligation followed by hybridization with optimized capture probes. Data was processed using publicly available and newly validated software tools to make mutation assignments for base alterations, indels, and CNAs in 182 cancer associated genes and common sites of rearrangement for 15 genes. Results: 5/8 biopsies and 62/65 (overall 93%) of prostate foci yielded sufficient DNA (>50 ng) for analysis from 40m of FFPE tissue. Average median sequence coverage was 934x. TMPRSS2-ERG fusion was present in 28% of NEPC. Recurrent homozygous deletions involving PTEN and RB1 were seen, and 1 tumor with BRCA2 loss. Several high confidence non-synonymous mutations were identified including TP53 (40%), CTNNB1 (12%), and less frequently mutations involving PTEN, PIK3CA, AR, as well as other novel mutations/fusions. There was high concordance between NEPC and PCA foci in mixed tumors, as well as between primary tumor and metastases. High confidence lesions were validated with exome sequencing and FISH. Conclusions: This study shows feasibility of an in-depth DNA analysis using FFPE tissue, and even biopsy material. Comprehensive genome sequencing has nominated novel biologic pathways and provides insight into disease progression from PCA to NEPC, as well as potential new drug targets for a tumor that is currently lethal. This is a useful and comprehensive sequencing tool to evaluate tumors such as NEPC and other metastatic tumors, where obtaining tissue is challenging.

scholarly journals Laboratory Verification of a BRCA1 and BRCA2 Massively Parallel Sequencing Assay from Wet Bench to Bioinformatics for Germline DNA Analysis

2021 ◽  
Author(s):  
Kok-Siong Poon ◽  
Lily Chiu ◽  
Karen Mei-Ling Tan
Keyword(s):  
Genetic Testing ◽  
Sanger Sequencing ◽  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Massively Parallel ◽  
Brca1 And Brca2 ◽  
Clinical Diagnostic Laboratory ◽  
Parallel Sequencing ◽  
Miseq Platform ◽  
Brca Genetic Testing

Abstract Introduction A robust genetic test for BRCA1 and BRCA2 genes is necessary for the diagnosis, prognosis, and treatment of patients with hereditary breast and ovarian cancer. We evaluated a commercial amplicon-based massively parallel sequencing (MPS) assay, BRCA MASTR Plus on the MiSeq platform, for germline BRCA genetic testing. Methods This study was performed on 31 DNA from cell lines and proficiency testing samples to establish the accuracy of the assay. A reference cell line DNA, NA12878 was used to determine the reproducibility of the assay. Discordant MPS result was resolved orthogonally by the current gold-standard Sanger sequencing method. Results The analytical accuracy, sensitivity, and specificity for variant detection were 93.55, 92.86, and 100.00%, respectively. Both sequencing depth and variant allele frequencies were highly reproducible by comparing the NA12878 DNA tested in three separate runs. The single discordant result, later confirmed by Sanger sequencing was due to the inability of the MASTR Reporter software to identify a 40-bp deletion in BRCA1. Conclusion The BRCA MASTR Plus assay on the MiSeq platform is accurate and reproducible for germline BRCA genetic testing, making it suitable for use in a clinical diagnostic laboratory. However, Sanger sequencing may still serve as a confirmatory method to improve diagnostic capability of the MPS assay.

scholarly journals High-Resolution Profiling of Fetal DNA Clearance from Maternal Plasma by Massively Parallel Sequencing

2013 ◽  
Vol 59 (8) ◽  
pp. 1228-1237 ◽  
Author(s):  
Stephanie CY Yu ◽  
Shara WY Lee ◽  
Peiyong Jiang ◽  
Tak Y Leung ◽  
KC Allen Chan ◽  
...  
Keyword(s):  
High Resolution ◽  
Half Life ◽  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Maternal Plasma ◽  
Massively Parallel ◽  
Slow Phase ◽  
Parallel Sequencing ◽  
Fetal Dna ◽  
Circulating Fetal Dna

BACKGROUND With the advent of massively parallel sequencing (MPS), DNA analysis can now be performed in a genomewide manner. Recent studies have demonstrated the high precision of MPS for quantifying fetal DNA in maternal plasma. In addition, paired-end sequencing can be used to determine the size of each sequenced DNA fragment. We applied MPS in a high-resolution investigation of the clearance profile of circulating fetal DNA. METHODS Using paired-end MPS, we analyzed serial samples of maternal plasma collected from 13 women after cesarean delivery. We also studied the transrenal excretion of circulating fetal DNA in 3 of these individuals by analyzing serial urine samples collected after delivery. RESULTS The clearance of circulating fetal DNA occurred in 2 phases, with different kinetics. The initial rapid phase had a mean half-life of approximately 1 h, whereas the subsequent slow phase had a mean half-life of approximately 13 h. The final disappearance of circulating fetal DNA occurred at about 1 to 2 days postpartum. Although transrenal excretion was involved in the clearance of circulating fetal DNA, it was not the major route. Furthermore, we observed significant changes in the size profiles of circulating maternal DNA after delivery, but we did not observe such changes in circulating fetal DNA. CONCLUSIONS MPS of maternal plasma and urinary DNA permits high-resolution study of the clearance profile of circulating fetal DNA.

scholarly journals Sequence Artifacts in DNA from Formalin-Fixed Tissues: Causes and Strategies for Minimization

2015 ◽  
Vol 61 (1) ◽  
pp. 64-71 ◽  
Author(s):  
Hongdo Do ◽  
Alexander Dobrovic
Keyword(s):  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Amplicon Sequencing ◽  
Careful Consideration ◽  
Dna Lesions ◽  
Massively Parallel ◽  
Cytosine Deamination ◽  
Accurate Detection ◽  
Ffpe Tissues ◽  
Formalin Fixed

Abstract BACKGROUND Precision medicine is dependent on identifying actionable mutations in tumors. Accurate detection of mutations is often problematic in formalin-fixed paraffin-embedded (FFPE) tissues. DNA extracted from formalin-fixed tissues is fragmented and also contains DNA lesions that are the sources of sequence artifacts. Sequence artifacts can be difficult to distinguish from true mutations, especially in the context of tumor heterogeneity, and are an increasing interpretive problem in this era of massively parallel sequencing. Understanding of the sources of sequence artifacts in FFPE tissues and implementation of preventative strategies are critical to improve the accurate detection of actionable mutations. CONTENT This mini-review focuses on DNA template lesions in FFPE tissues as the source of sequence artifacts in molecular analysis. In particular, fragmentation, base modification (including uracil and thymine deriving from cytosine deamination), and abasic sites are discussed as indirect or direct sources of sequence artifacts. We discuss strategies that can be implemented to minimize sequence artifacts and to distinguish true mutations from sequence artifacts. These strategies are applicable for the detection of actionable mutations in both single amplicon and massively parallel amplicon sequencing approaches. SUMMARY Because FFPE tissues are usually the only available material for DNA analysis, it is important to maximize the accurate informational content from FFPE DNA. Careful consideration of each step in the work flow is needed to minimize sequence artifacts. In addition, validation of actionable mutations either by appropriate experimental design or by orthogonal methods should be considered.

scholarly journals Progress in the implementation of massively parallel sequencing for forensic genetics: results of a European-wide survey among professional users

2021 ◽  
Author(s):  
Theresa E. Gross ◽  
Jan Fleckhaus ◽  
Peter M. Schneider
Keyword(s):  
Online Survey ◽  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Forensic Genetics ◽  
Massively Parallel ◽  
Forensic Dna ◽  
Educational Training ◽  
Forensic Dna Analysis ◽  
Parallel Sequencing ◽  
Horizon 2020

AbstractA European-wide online survey was conducted to generate an overview on the state-of-the-art using massively parallel sequencing (MPS) platforms for forensic DNA analysis and DNA phenotyping among forensic practitioners in Europe. The survey was part of the dissemination activities of the “VISible Attributes through GEnomics – VISAGE” Horizon 2020 funded European research project [30], in preparation of a series of educational training activities. A total of 105 replies from 32 European countries representing participants from police, governmental, academic, and private laboratories providing professional services in the field of forensic genetics were included in the final analysis. Of these, 73% already own an MPS platform or are planning to acquire one within the next 1–2 years. One-third of the participants have already carried out MPS-based STR sequencing, identity, or ancestry SNP typing. A total of 23–40% of participants are planning to explore all FDP applications showing the overall very high interest in using MPS for the whole range of forensic MPS markers and applications. About 50% of the participants have previously gathered experience using forensic DNA phenotyping (FDP) markers based on conventional (i.e., not MPS-based) DNA typing methods. A total of 55% of the participants have attended training on the general use of MPS technology, but 36% have received no training whatsoever. Accordingly, 90% have expressed high or medium interest to attend training on the analysis and interpretation of DNA phenotyping data for predicting appearance, ancestry, and age. The results of our survey will provide valuable information for organizing relevant training workshops on all aspects of MPS-based DNA phenotyping for the forensic genetics scientific community.

Recent Developments in Forensic DNA Technology

2020 ◽  
pp. 105-127
Author(s):  
Henry Erlich ◽  
Cassandra Calloway ◽  
Steven B. Lee
Keyword(s):  
Mitochondrial Dna ◽  
Dna Analysis ◽  
Massively Parallel Sequencing ◽  
Massively Parallel ◽  
Degraded Dna ◽  
Forensic Dna ◽  
Current Standard ◽  
Forensic Dna Analysis ◽  
Str Loci ◽  
Recent Developments

The current standard of forensic DNA analysis is genotyping the length polymorphism of STR loci by capillary electrophoresis and analyzing the polymorphism of mitochondrial DNA by Sanger sequencing. However, the trend of dramatic technological developments begun in the mid 1980s has continued, with the most consequential recent innovations being (1) the development of next generation sequencing (NGS) or massively parallel sequencing (MPS) and (2) the implementation of commercial Rapid DNA instruments that automate genotyping of all CODIS core STR loci from sample to profile in 90 minutes. This chapter reviews the principles, benefits, and applications of NGS or MPS technology, with a focus on the critical features of massively parallel and clonal sequencing and the ability to perform quantitative analysis of mixtures. The potential to analyze degraded DNA by using NGS/MPS to sequence mitochondrial DNA and SNPs is discussed, as are the benefits and limitations of Rapid DNA instruments.

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