Applicability of DNA Analysis on Adhesive Tape in Forensic Casework

Adhesive tape is commonly used in crimes and is often the subject of forensic evaluation. DNA analysis of adhesive tape can provide DNA profiles of suspects. The object of this study was to evaluate the applicability of DNA analysis on adhesive tape samples in forensic casework. We retrospectively reviewed all cases involving adhesive tape or similar items received by our institute for DNA analysis during the past 11 years. From 100 forensic cases reviewed, 150 adhesive tape samples were examined. A total of 98 DNA profiles were obtained from these samples. Sixty-two of the profiles provided feasible case-relevant information. In conclusion, DNA profiling of adhesive tape samples can be useful in a variety of forensic cases.

Simultaneous analysis of nuclear and mitochondrial DNA, mRNA and miRNA from backspatter from inside parts of firearms generated by shots at "triple contrast" doped ballistic models

When a firearm projectile hits a biological target a spray of biological material (e.g., blood and tissue fragments) can be propelled from the entrance wound back towards the firearm. This phenomenon has become known as "backspatter" and if caused by contact shots or shots from short distances traces of backspatter may reach, consolidate on, and be recovered from, the inside surfaces of the firearm. Thus, a comprehensive investigation of firearm-related crimes must not only comprise of wound ballistic assessment but also backspatter analysis, and may even take into account potential correlations between these emergences. The aim of the present study was to evaluate and expand the applicability of the "triple contrast" method by probing its compatibility with forensic analysis of nuclear and mitochondrial DNA and the simultaneous investigation of co-extracted mRNA and miRNA from backspatter collected from internal components of different types of firearms after experimental shootings. We demonstrate that "triple contrast" stained biological samples collected from the inside surfaces of firearms are amenable to forensic co-analysis of DNA and RNA and permit sequence analysis of the entire mtDNA displacement-loop, even for "low template" DNA amounts that preclude standard short tandem repeat DNA analysis. Our findings underscore the "triple contrast" method's usefulness as a research tool in experimental forensic ballistics.

Serological and DNA-based evaluation of Chlamydia pneumoniae infection in inflammatory bowel disease

OBJECTIVES: Chlamydia has been associated with autoimmune diseases, but a link between chlamydial infection and the aetiopathogenesis of inflammatory bowel disease (IBD) remains controversial. In this study we assessed the relationship between chlamydial infection and IBD, as evidenced by serological measurement and DNA analysis of mucosal biopsy specimens. PATIENTS AND METHODS: The sera of 78 patients with Crohn's disease (CD), 24 patients with ulcerative colitis (UC), 73 healthy family members, and 20 healthy controls were tested for anti-C. pneumoniae IgG titres. A subgroup consisting of 13 UC and 39 CD patients was screened for the presence of chlamydial DNA on 42 inflamed versus 30 non-inflamed biopsy specimens and for mutations of their NOD2/CARD15 gene. RESULTS: Anti-C. pneumoniae IgG antibodies were found in the sera of 32 (41%) patients with CD, 11 (46%) patients with UC, 35 (48%) of unaffected family members, and nine (45%) unrelated healthy controls. Thirty-five percent of the control, 18% CD and 24% UC biopsy specimens contained C. pneumoniae DNA. In CD, however, C. pneumoniae DNA was significantly more frequently found in inflamed (27%) versus non-inflamed (8%) biopsy specimens (P < 0.05, Fisher's exact test). The frequencies of NOD2/CARD15 mutations were 33% for CD patients with C. pneumoniae DNA compared to 47% for CD patients without C. pneumoniae DNA. CONCLUSION: We found no marked differences in respect to anti-C. pneumoniae serum IgG or C. pneumoniae DNA between healthy controls and patients with IBD. However, in CD patients, inflamed tissue specimens contained significantly more likely C. pneumoniae DNA compared with biopsies from unaffected areas. Thus C. pneumoniae is unlikely to be of pathogenic importance in IBD while it may still influence local clinical manifestations.

Analysis of mRNA transcripts improves the success rate of molecular genetic testing in OTC deficiency

BACKGROUND: Ornithine transcarbamylase (OTC) deficiency is the most common inborn error of urea metabolism that can lead to hyperammonemic crises and orotic aciduria. To date, a total of 341 causative mutations within the OTC gene have been described. However, in about 20% of the patients with enzymatically confirmed OTC deficiency no mutation can be detected when sequencing of genomic DNA analyzing exons and adjacent intronic segments of the OTC gene is performed. METHODS: Standard genomic DNA analysis of the OTC gene in five consecutive patients from five families revealed no mutation. Hence, liver tissue was obtained by needle sampling or open biopsy and RNA extracted from liver was analyzed. RESULTS: Complex rearrangements of the OTC transcript (three insertions and two deletions) were found in all five patients. CONCLUSION: In patients with a strong suspicion of OTC deficiency despite normal results of sequencing exonic regions of the OTC gene, characterization of liver OTC mRNA is highly effective in resolving the genotype. Liver tissue sampling by needle aspiration allows for both enzymatic analysis and RNA based diagnostics of OTC deficiency.

Analysis of coprolites from the extinct mountain goat Myotragus balearicus

Humans colonized the Balearic Islands 5-4 ka ago. They arrived in a uniquely adapted ecosystem with the Balearic mountain goat Myotragus balearicus (Bovidae, Antilopinae, Caprini) as the only large mammal. This mammal went extinct rapidly after human arrival. Several hypotheses have been proposed to explain the extinction of M. balearicus. For the present study ancient DNA analysis (Sanger sequencing, Roche-454, Ion Torrent), and pollen and macrofossil analyses were performed on preserved coprolites from M. balearicus, providing information on its diet and paleo-environment. The information retrieved shows that M. balearicus was heavily dependent on the Balearic box species Buxus balearica during at least part of the year, and that it was most probably a browser. Hindcast ecological niche modelling of B. balearica shows that local distribution of this plant species was affected by climate changes. This suggests that the extinction of M. balearicus can be related to the decline and regional extinction of a plant species that formed a major component of its diet. The vegetation change is thought to be caused by increased aridity occurring throughout the Mediterranean. Previous hypotheses relating the extinction of M. balearicus directly to the arrival of humans on the islands must therefore be adjusted. (C) 2013 University of Washington. Published by Elsevier Inn All rights reserved.

Congenital hypotrichosis and partial anodontia in a crossbred beef calf

Clinical examination, skin biopsies, skull radiographs, and DNA analysis of a 2-day-old Red Angus-Charolais-Simmental cross bull calf confirmed the diagnosis of congenital hypotrichosis and anodontia defect (HAD), also called anhidrotic ectodermal dysplasia, which is a rare anomaly caused by a deletion in the bovine EDA gene on the X chromosome.

[Nocardia cyriacigeorgici: First report of invasive human infection]

BACKGROUND Diagnostic laboratories increasingly offer bacterial identification to the species level. The 17 nocardia species known to date differ in their clinical presentation, antibiotic resistance patterns and geographic distribution. The discovery of a new species with pathogenicity for humans calls for the characterization of its clinical and epidemiological properties. PATIENTS AND METHODS Nocardia isolated from multifocal brain abscesses of an immunocompromised patient were further identified by the analysis of their cellular fatty acids and sequencing of the 16S ribosomal DNA. Quantitative antibiotic resistance testing was performed with E-tests. RESULTS The 16S ribosomal DNA analysis showed a 99 % homology to Nocardia cyriacigeorgici. This is the first report of this species as an invasive human pathogen. N. cyriacigeorgici was found susceptible for meropenem, amikacin, ceftriaxon and cotrimoxazole. The combination of surgical drainage and antibiotic treatment for 13 months was curative. CONCLUSIONS N. cyriacigeorgici has the potential to cause invasive infections at least in immunocompromised patients. Comparing clinical and in vitro characteristics with N. asteroides, the main causative agent of nocardial infections in Europe, we found no clinically relevant differences.

When the prey gets too big: an uncommon road accident involving a motorcyclist, a car and a bird

We present the postmortem findings of a fatal road accident involving a motorcyclist, a car, and a common buzzard. Both the motorcyclist and the bird died on the scene of the accident and were examined by postmortem full-body CT and autopsy. In addition, a facial injury of the motorcyclist was compared with the dimensions of the buzzard’s beak and claws by 3D scan technologies. Blood splatters collected on the bird’s beak, feet, and tail were examined by DNA analysis. The overall findings suggested a collision of a common buzzard with a motorcyclist in full speed, causing the motorcyclist to lose control of his vehicle and crash with an approaching car on the oncoming lane.

Models of hybridization during range expansions and their application to recent human evolution

Several lines of genetic, archeological and paleontological evidence suggest that anatomically modern humans (Homo sapiens) colonized the world in the last 60,000 years by a series of migrations originating from Africa (e.g. Liu et al., 2006; Handley et al., 2007; Prugnolle, Manica, and Balloux, 2005; Ramachandran et al. 2005; Li et al. 2008; Deshpande et al. 2009; Mellars, 2006a, b; Lahr and Foley, 1998; Gravel et al., 2011; Rasmussen et al., 2011). With the progress of ancient DNA analysis, it has been shown that archaic humans hybridized with modern humans outside Africa. Recent direct analyses of fossil nuclear DNA have revealed that 1–4 percent of the genome of Eurasian has been likely introgressed by Neanderthal genes (Green et al., 2010; Reich et al., 2010; Vernot and Akey, 2014; Sankararaman et al., 2014; Prufer et al., 2014; Wall et al., 2013), with Papua New Guineans and Australians showing even larger levels of admixture with Denisovans (Reich et al., 2010; Skoglund and Jakobsson, 2011; Reich et al., 2011; Rasmussen et al., 2011). It thus appears that the past history of our species has been more complex than previously anticipated (Alves et al., 2012), and that modern humans hybridized several times with local hominins during their expansion out of Africa, but the exact mode, time and location of these hybridizations remain to be clarifi ed (Ibid.; Wall et al., 2013). In this context, we review here a general model of admixture during range expansion, which lead to some predictions about expected patterns of introgression that are relevant to modern human evolution.

Quantitative 1-Step DNA Methylation Analysis with Native Genomic DNA as Template

DNA methylation analysis currently requires complex multistep procedures based on bisulfite conversion of unmethylated cytosines or on methylation-sensitive endonucleases. To facilitate DNA methylation analysis, we have developed a quantitative 1-step assay for DNA methylation analysis.

Quantitative analysis of O(6)-methylguanine DNA methyltransferase (MGMT) promoter methylation in patients with low-grade gliomas

Methylation of the MGMT promoter is supposed to be a predictive and prognostic factor in glioblastoma. Whether MGMT promoter methylation correlates with tumor response to temozolomide in low-grade gliomas is less clear. Therefore, we analyzed MGMT promoter methylation by a quantitative methylation-specific PCR in 22 patients with histologically verified low-grade gliomas (WHO grade II) who were treated with temozolomide (TMZ) for tumor progression. Objective tumor response, toxicity, and LOH of microsatellite markers on chromosomes 1p and 19q were analyzed. Histological classification revealed ten oligodendrogliomas, seven oligoastrocytomas, and five astrocytomas. All patients were treated with TMZ 200 mg/m2 on days 1-5 in a 4 week cycle. The median progression-free survival was 32 months. Combined LOH 1p and 19q was found in 14 patients; one patient had LOH 1p alone and one patient LOH 19q alone. The LOH status could not be determined in two patients and was normal in the remaining four. LOH 1p and/or 19q correlated with longer time to progression but not with radiological response to TMZ. MGMT promoter methylation was detectable in 20 patients by conventional PCR and quantitative analysis revealed the methylation status was between 12 and 100%. The volumetric response to chemotherapy analyzed by MRI and time to progression correlated with the level of MGMT promoter methylation. Therefore, our retrospective case series suggests that quantitative methylation-specific PCR of the MGMT promoter predicts radiological response to chemotherapy with TMZ in WHO grade II gliomas.

Prediction of whole-genome DNA-DNA similarity, determination of G+C content and phylogenetic analysis within the family Pasteurellaceae by multilocus sequence analysis (MLSA)

Genome predictions based on selected genes would be a very welcome approach for taxonomic studies, including DNA-DNA similarity, G+C content and representative phylogeny of bacteria. At present, DNA-DNA hybridizations are still considered the gold standard in species descriptions. However, this method is time-consuming and troublesome, and datasets can vary significantly between experiments as well as between laboratories. For the same reasons, full matrix hybridizations are rarely performed, weakening the significance of the results obtained. The authors established a universal sequencing approach for the three genes recN, rpoA and thdF for the Pasteurellaceae, and determined if the sequences could be used for predicting DNA-DNA relatedness within the family. The sequence-based similarity values calculated using a previously published formula proved most useful for species and genus separation, indicating that this method provides better resolution and no experimental variation compared to hybridization. By this method, cross-comparisons within the family over species and genus borders easily become possible. The three genes also serve as an indicator of the genome G+C content of a species. A mean divergence of around 1 % was observed from the classical method, which in itself has poor reproducibility. Finally, the three genes can be used alone or in combination with already-established 16S rRNA, rpoB and infB gene-sequencing strategies in a multisequence-based phylogeny for the family Pasteurellaceae. It is proposed to use the three sequences as a taxonomic tool, replacing DNA-DNA hybridization.