Evaluation of the IrisPlex DNA-based eye color prediction assay in a United States population

DNA phenotyping is a rapidly developing area of research in forensic biology. Externally visible characteristics (EVCs) can be determined based on genotype data, specifically based on single nucleotide polymorphisms (SNPs). These SNPs are chosen based on their association with genes related to the phenotypic expression of interest, with known examples in eye, hair, and skin color traits. DNA phenotyping has forensic importance when unknown biological samples at a crime scene do not result in a criminal database hit; a phenotypic profile of the sample can therefore be used to develop investigational leads. IrisPlex, an eye color prediction assay, has previously shown high prediction rates for blue and brown eye color in a Dutch European population. The objective of this work was to evaluate its utility in a North American population. We evaluated six SNPs included in the IrisPlex assay in population sample collected from a USA college campus. We used a quantitative method of eye color classification based on (RGB) color components of digital photographs of the eye taken from each study volunteer so that each eye was placed in one of three eye color categories: brown, intermediate, or blue. Objective color classification was shown to correlate with basic human visual determination making it a feasible option for use in future prediction assay development. Using these samples and various models, the maximum prediction accuracies of the IrisPlex system after allele frequency adjustment was 58% and 95% brown and blue eye color predictions, respectively, and 11% for intermediate eye colors. Future developments should include incorporation of additional informative SNPs, specifically related to the intermediate eye color, and we recommend the use of a Bayesian approach as a prediction model as likelihood ratios can be determined for reporting purposes.     

Forensic validation of the SNPforID 52-plex assay

The advantages of single nucleotide polymorphism (SNP) typing in forensic genetics are well known and include a wider choice of high-throughput typing platforms, lower mutation rates, and improved analysis of degraded samples. However, if SNPs are to become a realistic supplement to current short tandem repeat (STR) typing methods, they must be shown to successfully and reliably analyse the challenging samples commonly encountered in casework situations.The European SNPforID consortium, supported by the EU GROWTH programme, has developed a multiplex of 52 SNPs for forensic analysis, with the amplification of all 52 loci in a single reaction followed by two single base extension (SBE) reactions which are detected with capillary electrophoresis.In order to validate this assay, a variety of DNA extracts were chosen to represent problems such as low copy number and degradation that are commonly seen in forensic casework. A total of 40 extracts were used in the study, each of which was sent to two of the five participating laboratories for typing in duplicate or triplicate. Laboratories were instructed to carry out their analyses as if they were dealing with normal casework samples. Results were reported back to the coordinating laboratory and compared with those obtained from traditional STR typing of the same extracts using Powerplex^® 16 (Promega). These results indicate that, although the ability to successfully type good quality, low copy number extracts is lower, the 52-plex SNP assay performed better than STR typing on degraded samples, and also on samples that were both degraded and of limited quantity, suggesting that SNP analysis can provide advantages over STR analysis in forensically relevant circumstances. However, there were also additional problems arising from contamination and primer quality issues and these are discussed.     

Mutations in the SCN5A gene: Evidence for a link between long QT syndrome and sudden death?

Mutations in cardiac ion channel genes leading to channel dysfunctions or changes in the gene expression may cause inherited arrhythmogenic diseases. These genetic diseases are important causes of sudden unexplained death (SUD). Ten cases of SUD, including six cases of sudden infant death syndrome (SIDS) and four cases of SUD from people in the age of 14–40 years were examined by postmortem molecular analysis. Genomic DNA was extracted from blood cells and two long QT syndrome relevant genes, SCN5A encoding the α-subunit of the voltage-gated sodium channel Na_v1.5 and KCNH2 encoding the α-subunit of the voltage-gated potassium channel HERG were selected for mutation analysis by complete gene sequencing. Various silent mutations in the KCNH2 and SCN5A genes as well as the known H558R polymorphism in SCN5A were detected. Moreover, sequence variations in the 3′ untranslated region (3′UTR) and 5′ untranslated region (5′UTR) of the SCN5A gene were observed. This study suggests that these areas are important regions to investigate the impact of changes in cardiac ion channel function on the risk of sudden unexpected death.     

Apports nutritionnels précoces,hypotrophie acquise et devenir du prématuré

Early nutrition is an important determinant of growth, morbidity and psychomotor development in very low birth weight infants. Recent data suggest that “aggressive nutrition” providing early and high protein supply from the first hours of life improves nitrogen balance, reduces postnatal growth retardation, decreases morbidity and is positively related to psychomotor development. This paper will summarize the metabolic bases of this optimal nutrition for preterm infants.     

Genetic analysis of 27 Y-chromosomal STR loci in a Zimbabwean Shona ethnic group

Buccal swabs from 200 unrelated Zimbabwean males were collected from voluntary participants located in Harare province. The 5-dye SureID® 27Y Human STR Identification Kit was used to perform multiplex polymerase chain reactions (PCR) and generate Y-chromosomal DNA profiles. This kit targets markers DYS456, DYS576, DYS570, DYS481, DYF387S1, DYS627, DYS393, DYS391, DYS390, DYS635, DYS449, DYS533, DYS438, DYS389I, DYS448, DYS389II, DYS19, GATA_H4, DYS518, DYS458, DYS460, DYS437, DYS439, DYS392, and DYS385, similar to the Yfiler® Plus Amplification Kit. A total of 161 haplotypes were generated with the PowerPlex® Y system, whereas 159 complete haplotypes were generated for the Yfiler® Plus system. Haplotype Discrimination Capacity (DC) with the Yfiler® Plus system was determined to be 0.9686, while the Genetic Diversity (GD) of the targeted loci ranged from 0.03748 at DYS392 to 0.867239 at DYS449. One haplotype contained the triallelic pattern 37, 38, and 39 at DYS387S1. In addition, marker DYS387S1 and marker DYS385 had 13 counts of microvariant alleles overall, while 9 null allele counts were noted at marker DYS448. Genetic distances between our population data and 22 other data sets from African countries and people of African descent were estimated and results showed significant genetic variation.     

The discussion on pathomechanical changes of heart-spirit disease pattern from the vacuity detriment of heart

The heart tends to suffer from detriment (sun) and from diseases that belong to vacuity patterns. Due to the heart's governance of the spirit-mind, the depletion of heart yı̄n, yáng, qı̀ or blood will give rise to a variety of pathological changes of the heart-spirit. Heart qı̀ vacuity may result in insufficiency of spirit-qı̀, mental confusion, heart-spirit dissipation and desertion. Heart yáng vacuity can lead to devitalized essence-spirit, or fulminant desertion of heart yáng. Heart blood vacuity probably deprives heart-spirit of nourishment, even leading to the spirit's failure to abide. Heart yı̄n vacuity may lead to the heart-spirit becoming deprived of nourishment, giving rise to fulminant desertion of heart yı̄n. It thereby shows that heart diseases, due to either yáng qı̀ depletion or heart yı̄n blood insufficiency, can result in disease patterns of heart-spirit.     

Multiple recurrent mutations at four human Y-chromosomal single nucleotide polymorphism sites in a 37 bp sequence tract on the ARSDP1 pseudogene

The male-specific region of the human Y chromosome (MSY) is passed down clonally from father to son and mutation is the single driving force for Y-chromosomal diversification. The geographical distribution of MSY variation is non-random. Therefore, Y-chromosomal single nucleotide polymorphisms (Y-SNPs) are of forensic interest, as they can be utilized, e.g. for deducing the bio-geographical origin of biological evidence. This extra information can complement short tandem repeat data in criminal investigations. For forensic applications, however, any targeted marker has to be unequivocally interpretable.Here, we report findings for 17 samples from a population study comprising specimens from ∼3700 men living in Tyrol (Austria), indicating apparent homoplasic mutations at four Y-SNP loci on haplogroup R-M412/L51/S167, R-U152/S28, and L-M20 Y chromosomes. The affected Y-SNPs P41, P37, L202, and L203 mapped to a 37 bp region on Yq11.21. Observing in multiple phylogenetic contexts up to four homoplasic mutations within such a short sequence tract is unlikely to result from a series of independent parallel mutations. Hence, we rather propose X-to-Y gene conversion as a more likely scenario.Practical implications arising from markers exhibiting paralogues on the Y chromosome or sites with a high propensity to recurrent mutation for database searches are addressed.     

Development and validation of a new STR 25-plex typing system

In this study, a new STR 25-plex typing system, including 23 autosomal STRs (D1S1656, D2S1338, D2S441, D3S1358, D5S818, D6S1043, D7S820, D8S1179, D10S1248, D12S391, D13S317, D16S539, D18S51, D19S433, D21S11, D22S1045, CSF1PO, FGA, Penta D, Penta E, TH01, TPOX, vWA) and a Y-STR locus of DYS391 and amelogenin, was developed. The included 24 STRs belonged to the main international DNA databases (CODIS, ISSL, ESS-extended, UCL, GCL and NCIDD) except D6S1043 (specially chosen for Chinese population). Developmental validation indicated that the STR 25-plex typing system was reproducible, accurate, sensitive and robust. The sensitivity testing of the system was such that a full profile was obtainable even with 125 pg of human DNA. Specificity testing was demonstrated by the lack of cross-reactivity with a variety of commonly encountered animal species and microbial pool. For the stability testing, full profiles can been obtained with humic acid concentration ≤ 60 ng/μL and hematin < 500 μM. Also, this multiplex system is suitable for mixture study. All of the minor alleles were called for ratios of 1:1, 1:3 and 3:1 of the mixture with the system. In addition, the whole PCR amplification can finish within 1 h, making the system suitable for fast-detection. For the forensic evaluation of the multiplex system, 23 autosomal STRs included followed the Hardy–Weinberg equilibrium. A total of 268 alleles were detected for the 23 autosomal STR loci among 200 individuals. Since 23 autosomal STRs were independent from each other, CMEC_duo was 0.99999916563607 and CMEC_tri was 0.99999999986525. All the forensic efficiency parameters demonstrated that this multiplex system is highly polymorphic and informative in the Han population of China.     

Demonstration of DSI-semen—A novel DNA methylation-based forensic semen identification assay

Determining whether the source tissue of biological material is semen is important in confirming sexual assaults, which account for a considerable percentage of crime cases. The gold standard for confirming the presence of semen is microscopic identification of sperm cells, however, this method is labor intensive and operator-dependent. Protein-based immunologic assays, such as PSA, are highly sensitive and relatively fast, but suffer from low specificity in some situations. In addition, proteins are less stable than DNA under most environmental insults. Recently, forensic tissue identification advanced with the development of several approaches based on mRNA and miRNA for identification of various body fluids. Herein is described DNA source identifier (DSI)-semen, a DNA-based assay that determines whether the source tissue of a sample is semen based on detection of semen-specific methylation patterns in five genomic loci. The assay is comprised of a simple single tube biochemical procedure, similar to DNA profiling, followed by automatic software analysis, yielding the identification (semen/non-semen) accompanied by a statistical confidence level. Three additional internal control loci are used to ascertain the reliability of the results. The assay, which aims to replace microscopic examination, can easily be integrated by forensic laboratories and is automatable. The kit was tested on 135 samples of semen, saliva, venous blood, menstrual blood, urine, and vaginal swabs and the identification of semen vs. non-semen was correct in all cases. In order to test the assay's applicability in “real-life” situations, 33 actual casework samples from the forensic biological lab of the Israeli police were analyzed, and the results were compared with microscopic examination performed by Israeli police personnel. There was complete concordance between both analyses except for one sample, in which the assay identified semen whereas no sperm was seen in the microscope. This sample likely represents true semen because sperm cells were detected from an adjacent sample from the same garment, therefore in this case the assay appears to be more sensitive than the microscopic examination. These results demonstrate that this assay is a bona fide confirmatory test for semen.