Quantitative behavioral genetic and molecular genetic foundations of the approach and avoidance strategies.

Two studies examined genetic and environmental influences on traits proposed by the revised Reinforcement Sensitivity Theory (rRST) of personality. Both quantitative and molecular behavioral genetic methods were applied considering the effects of COMT, DRD2, HTR1A and TPH2 single nucleotide polymorphisms (SNPs). Study one included 274 monozygotic and 154 dizygotic twins for the quantitative behavioral study; and in study two there were 431 twins for the molecular genetic study. The Reinforcement Sensitivity Questionnaire was used to assess basic personality traits defined by the rRST. Univariate biometric modeling suggested that genetic influences accounted for 34-44% of variance of Behavioral Approach System (BAS), Behavioral Inhibition System (BIS) and Fight-Fligh-Freeze System. Molecular genetic analyses proposed the significant main effect of COMT SNP on the BAS and TPH2 SNP on the BIS, and pointed out epistatic effects of COMT x DRD2 on BAS and HTR1A x TPH2 on Fight. Results demonstrated substantial heritability for all rRST constructs, as well as for differences in the molecular genetic basis of both approach-related and avoidance-related dimensions.

Twin study of laboratory-induced aggression.

The aim of this study was to explore genetic and environmental contributions to laboratory-induced aggressive behavior. On a sample of 478 adult twins (316 monozygotic), the Competitive Reaction Time Task was used for aggression induction. The results showed that the initial, basic level of aggression could be explained by both shared (45%) and nonshared environmental factors (55%), while only nonshared environmental factors (100%) had a significant influence on changes in aggression as provocation increased. Genetic factors had no influence on laboratory-induced aggression. The results highlight the importance of environmental factors in shaping situation-specific aggressive responses to provocation.

Proteomic analysis of menstrual blood.

Menstruation is the expulsion of the endometrial lining of the uterus following a nearly month long preparation for embryo implantation and pregnancy. Increasingly, the health of the endometrium is being recognized as a critical factor in female fertility, and proteomes and transcriptomes from endometrial biopsies at different stages of the menstrual cycle have been studied for both diagnostic and therapeutic purposes (1 Kao, L. C., et al. 2003 Endocrinology 144, 2870-2881; Strowitzki, Tet al. 2006 Hum. Reprod. Update 12, 617-630; DeSouza, L., et al. 2005 Proteomics 5, 270-281). Disorders of the uterus ranging from benign to malignant tumors, as well as endometriosis, can cause abnormal menstrual bleeding and are frequently diagnosed through endometrial biopsy (Strowitzki, Tet al. 2006 Hum. Reprod. Update 12, 617-630; Ferenczy, A. 2003 Maturitas 45, 1-14). Yet the proteome of menstrual blood, an easily available noninvasive source of endometrial tissue, has yet to be examined for possible causes or diagnoses of infertility or endometrial pathology. This study employed five different methods to define the menstrual blood proteome. A total of 1061 proteins were identified, 361 were found by at least two methods and 678 were identified by at least two peptides. When the menstrual blood proteome was compared with those of circulating blood (1774 proteins) and vaginal fluid (823 proteins), 385 proteins were found unique to menstrual blood. Gene ontology analysis and evaluation of these specific menstrual blood proteins identified pathways consistent with the processes of the normal endometrial cycle. Several of the proteins unique to menstrual blood suggest that extramedullary uterine hematopoiesis or parenchymal hemoglobin synthesis may be occurring in late endometrial tissue. The establishment of a normal menstrual blood proteome is necessary for the evaluation of its usefulness as a diagnostic tool for infertility and uterine pathologies. Identification of unique menstrual blood proteins should aid the forensic community in distinguishing menstrual blood from circulating blood.

Global survey on evaluative reporting on DNA evidence with regard to activity-level propositions.

For many criminal cases, the source of who deposited the DNA is not what the prosecutor and the defense are trying to dispute. In court, the question may be how the DNA was deposited at the crime scene rather than who the DNA came from. Although laboratories in many countries have begun to evaluate DNA evidence given formal activity-level propositions (ALPs), it is unknown how much other forensic practitioners know and what they think about activity-level evaluative reporting (ALR). To collect this information, a survey with 21 questions was submitted to international forensic science organizations across Europe, Australia, South America, Canada, Asia, and Africa. The survey combined open-ended and multiple-choice questions and received 162 responses. Responses revealed a wide range of knowledge on the topic. Overall, most respondents were somewhat knowledgeable about ALR, ALP, and current practices in court and expressed their support of the concept. A majority of participants identified gaps and obstacles regarding ALR they would like to see addressed. Examples include (1) need for more education/training at all stakeholder levels, (2) need for more DNA evidence-related data under realistic case scenarios, (3) need to internally implement and validate a formalized and objective approach for reporting, and (4) in some countries the need to achieve court admissibility. This global survey gathered the current concerns of forensic DNA practitioners and outlined several operational concerns. The information can be used to advance the implementation of ALR in laboratories and court testimony worldwide.

Body fluid identification by mass spectrometry.

Standard methods for body fluid identification typically rely on detection of the functional proteins specific to or enriched in them, such as hemoglobin in blood, alkaline phosphatase and PSA in semen, or α-amylase in saliva. While these markers can be relatively specific, the multiple methods used to identify them frequently rely on nonspecific chemical, enzymatic, or antibody reactions that usually require the structural integrity of the markers and are not confirmatory because other proteins or substances can also give positive test results. Recent advances in proteomics and mass spectrometry offer the ability to simultaneously detect multiple body fluid protein markers in a single, confirmatory test. Here, multiple markers for blood, saliva, and semen are identified by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS). Data demonstrate the ability to detect these body fluids at nanoliter to subnanoliter levels and to distinguish mixtures. Protein stability of mock samples assayed after 16 months showed no diminution of signal. Because multiple peptides from multiple protein markers are detected and effectively sequenced by MALDI MS/MS, the assay is confirmatory. As mass spectrometry detects whatever peptides are present in a sample, no a priori knowledge of an unknown stain is necessary to perform the test.

Improved eye- and skin-color prediction based on 8 SNPs.

AIM: To improve the 7-plex system to predict eye and skin color by increasing precision and detailed phenotypic descriptions. METHODS: Analysis of an eighth single nucleotide polymorphism (SNP), rs12896399 (SLC24A4), showed a statistically significant association with human eye color (P=0.007) but a rather poor strength of agreement (κ=0.063). This SNP was added to the 7-plex system (rs12913832 at HERC2, rs1545397 at OCA2, rs16891982 at SLC45A2, rs1426654 at SLC24A5, rs885479 at MC1R, rs6119471 at ASIP, and rs12203592 at IRF4). Further, the instruction guidelines on the interpretation of genotypes were changed to create a new 8-plex system. This was based on the analysis of an 803-sample training set of various populations. The newly developed 8-plex system can predict the eye colors brown, green, and blue, and skin colors light, not dark, and not light. It is superior to the 7-plex system with its additional ability to predict blue eye and light skin color. RESULTS: The 8-plex system was tested on an additional 212 samples, the test set. Analysis showed that the number of positive descriptions for eye colors as being brown, green, or blue increased significantly (P=6.98e-15, z-score: -7.786). The error rate for eye-color prediction was low, at approximately 5%, while the skin color prediction showed no error in the test set (1% in training set). CONCLUSIONS: We can conclude that the new 8-plex system for the prediction of eye and skin color substantially enhances its former version.

Development of Improved DNA Collection and Extraction Methods for Handled Documents.

Handwritten documents may contain probative DNA, but most crime laboratories do not process this evidence. DNA recovery should not impair other evidence processing such as latent prints or indented writing. In this study, single fingermarks on paper were sampled with flocked swabs, cutting, and dry vacuuming. In addition, two extraction methods were compared for the sample type. DNA yields were low across all methods; however, this work confirms the ability to recover DNA from paper and the usefulness of the vacuum sampling method combined with the Chelex-Tween method. Stability of touch DNA deposits were compared over an 11-month period to better understand degradation that may occur over time. No significant difference in DNA recovery was observed, suggesting DNA deposits on paper are stable over an 11-month span.

Ethnic differences in out-of-hospital fatal pulmonary embolism.

BACKGROUND: In-hospital pulmonary embolism (PE) has been extensively studied in large populations; however, out-of-hospital fatal PE studies are rare. Here, we systematically evaluated a large number of decedents who suffered fatal PE outside of hospitals and were subsequently investigated by the New York City Office of Chief Medical Examiner. METHODS AND RESULTS: A total of 578 consecutive out-of-hospital fatal PE cases were analyzed. All underwent autopsy, toxicology, microbiology, and genetic testing. Incidence rates and baseline characteristics were analyzed. Race-adjusted incidence rates of out-of-hospital fatal PE (per 100 000 people per year) were as follows: blacks, 3.73 (95% confidence interval, 3.31 to 4.11); whites, 1.15 (95% confidence interval, 0.96 to 1.33); and Hispanics, 0.93 (95% confidence interval, 0.72 to 1.10). Overall, obesity (body mass index ≥30 kg/m(2)) was 2.5- to 3-fold higher in fatal PE cases than in the New York City population as a whole. Carrier frequencies for prothrombin G20210A in fatal PE were 2- to 10-fold higher than reported frequencies in ethnically matched controls. Cumulative distribution curves showed that compared with whites, blacks and Hispanics died at significantly younger ages (P<0.001). Univariate and multiple linear regression analyses showed that in addition to nonwhite ethnicity, heterozygous carriers for factor V Leiden (P=0.001) and obesity (P=0.002) are significantly associated with younger age at death. CONCLUSION: There are unique epidemiological differences in out-of-hospital fatal PE between ethnic groups in New York City.

American forensic DNA practitioners' opinion on activity level evaluative reporting.

The technical advancements made in DNA profiling now allow for very low DNA amounts to be analyzed. Accordingly, the argument often made in criminal courts is not who the DNA belongs to but rather how it was deposited. Despite the complexity of the relevant DNA transfer, persistence, prevalence, and recovery issues, forensic laboratories in some European countries have used evaluative reports with activity level propositions, while this is not current practice in the United States. The purpose of this study was to gain an overview of the opinions about activity level reporting (ALR) held by forensic biologists in the United States. A seventeen-question survey was distributed to members of the American Society of Crime Laboratory Directors and U.S. members of the International Society for Forensic Genetics. The survey included multiple-choice and open-response questions and received 54 responses. The majority of responses expressed moderate support of ALR. Participants mentioned six major concerns to be addressed prior to implementing ALR in the United States: (1) effect of number of variables involved; (2) need of education for practitioners/legal system; (3) inadequate number of activity studies with realistic scenarios; (4) difficulty of achieving admissibility in court; (5) need for standardized approaches/guidelines; and (6) requisite shift in perspective as to the validity of ALR. Overall, this small segment of U.S. forensic DNA practitioners appear to be willing to implement ALR once these concerns are fully addressed and resolved. As a follow-up, it would be worthwhile exploring these and other questions with a larger group and also other disciplines.

High-throughput seminal fluid identification by automated immunoaffinity mass spectrometry.

The identification of semen during a criminal investigation may be a critical component in the prosecution of a sexual assault. Commonly employed enzymatic and affinity-based methods for detection lack specificity, are time-consuming, and only provide a presumptive indication that semen is present where microscopic visualization is unable to meet the throughput demands. Contrary to traditional approaches, protein mass spectrometry provides true confirmatory results, but multiday sample preparation and nanoflow sample separation requirements have limited the practical applicability of these approaches. Aiming at streamlining sexual assault screening by mass spectrometry, the work here coupled a 60-minute rapid tryptic digestion, semenogelin-II peptide affinity purification on an Agilent AssayMap Bravo automation platform, and a 3-minute targeted LC-MS/MS method on an Agilent 6495 triple quadrupole mass spectrometer operating in multiple reaction monitoring mode for detecting semenogelin-II peptides in sexual assault samples. The developed assay was assessed using casework-type samples and was successful in detecting trace levels (0.0001 µl) of semen recovered from both cotton and vaginal swabs, as well as semen recovered from vaginal swabs during menses or adulterated with personal lubricants. This work represents a promising technique for high-throughput seminal fluid identification in sexual assault-type samples by mass spectrometry.

Latent, genetic, and molecular genetic structure of the Wisconsin Card Sorting Test.

OBJECTIVE: The main goal of this study was to explore the latent structure and genetic basis of cognitive processes involved in the Wisconsin Card Sorting Task (WCST) within phenotypic, behavioral genetic, and molecular genetic research paradigms. METHOD: The sample used in phenotypic and behavioral genetic analyses comprised 468 twins (154 monozygotic and 80 dizygotic twin pairs), while molecular genetic analyses were performed on 404 twins from the same sample. The zygosity of most twin pairs (96.8%) was determined via deoxyribonucleic acid (DNA) analysis of buccal swabs. Trained researchers administered the Wisconsin Card Sorting Test (WCST; Heaton et al., 1993) to the entire sample. RESULTS: A phenotypic factor analysis of WCST variables suggested a single-factor solution. Overall heritability ranged from 0.19 to 0.23 across different measures of the WCST. The presence of a single general genetic factor, which could be identified from different measures of the WCST, indicated the unity of various WCST indicators and the existence of a common basic ability. Performance on the WCST did not reveal significant differences between the three genotypes on catechol-O-methyltransferase (COMT) and dopamine receptor D2 (DRD2). Carriers of the brain-derived neurotrophic factor (BDNF) Met + genotype exhibited better performance in cognitive functions in comparison to the BDNF Met- genotype. CONCLUSIONS: This study highlighted similarities in the phenotypic and genetic structures of the WCST, suggesting one general factor underlying different cognitive functions. The BDNF Met + genotype showed significant main effects on different WCST measures. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Differences in MB-COMT DNA methylation in monozygotic twins on phenotypic indicators of impulsivity.

Epigenetic modifications of the membrane bound catechol-O-methyltransferase (MB-COMT) gene may affect the enzymatic degradation of dopamine, and consequently, human behavior. This study investigated the association between membrane bound catechol-O-methyltransferase DNA methylation (DNAm) differences in 92 monozygotic (MZ) twins with phenotypic manifestations of cognitive, behavioral, and personality indicators associated with reward-related behaviors and lack of control. We used pyrosequencing to determine DNAm of the regulatory region of membrane bound catechol-O-methyltransferase in saliva DNA. Results of intrapair differences in the percentage of membrane bound catechol-O-methyltransferase DNAm at each of five CpG sites show that there are associations between phenotypic indicators of lack of control and membrane bound catechol-O-methyltransferase DNAm differences on CpG1, CpG2 and CpG4, suggesting the common epigenetic patterns for personality traits, cognitive functions, and risk behaviors.

Non-destructive DNA recovery from handwritten documents using a dry vacuum technique.

Investigations of many crimes such as robberies, kidnappings, and terrorism are often associated with the recovery of a paper document which has been written by the perpetrator. Paper can provide a variety of forensic evidence such as DNA, latent fingermarks, and indented writing. The focus of this study was DNA recovery from handwritten notes through a vacuum suction device while preserving the other evidence types and the integrity of the document. Copy paper was used to create handwritten documents and sheets with deliberate fingerprints, and indentations. The homemade vacuum device consists of a glass pipette blocked with a moistened swab and attached to a vacuum source. The method collected sufficient DNA amounts for DNA typing analysis with 80% of the 11 copy paper samples tested giving probative DNA profiles with five being eligible for DNA database entry. DNA recovery was also tested on other commonly encountered paper types. DNA quantities would have been sufficient for STR typing for approximately 50% of manila envelopes and notebook paper samples, but not for magazine pages and bank deposit slips. Deliberate sebaceous and eccrine latent fingermarks placed onto copy paper and developed with magnetic fingerprint developer or 1,2 indanedione were not affected by the vacuum swabbing technique. Simulated robbery notes with indented writing and processed using an Electrostatic Detection Apparatus (ESDA) demonstrated no interference through the DNA collection. This vacuum-based collection method enables laboratories to reverse the current questioned document workflow and start with DNA collection.

Methylation-Based Age Prediction Using Pyrosequencing Platform from Seminal Stains in Han Chinese Males.

Various methods have been performed to predict an unknown individual's age from biological traces in forensic investigations. A considerably accurate age prediction for the semen donor can help narrow down the search in a sexual assault case. The aim of this study was to develop an assay for age prediction from seminal stains in Han Chinese males. We built a sperm-specific linear regression model using bisulfite pyrosequencing. Validations were conducted with a Mean Absolute Deviation from the chronological age (MAD) of 4.219 years in liquid semen, 4.158 years in fresh seminal stains, 4.393 years in aged seminal stains, and 3.880 years in mixed stains, respectively. Furthermore, our strategy enables accurate age prediction using a forensic casework sample. The strategy indicated that we produced an accurate and reliable age prediction tool for the semen donors in Han Chinese males for forensic purposes.

DNA commission of the International society for forensic genetics: Assessing the value of forensic biological evidence - Guidelines highlighting the importance of propositions. Part II: Evaluation of biological traces considering activity level propositions.

The value of the evidence depends critically on propositions. In the second of two papers intended to provide advice to the community on difficult aspects of evaluation and the formulation of propositions, we focus primarily on activity level propositions. This helps the court address the question of "How did an individual's cell material get there?". In order to do this, we expand the framework outlined in the first companion paper. First, it is important not to conflate results and propositions. Statements given activity level propositions aim to help address issues of indirect vs direct transfer, and the time of the activity, but it is important to avoid use of the word 'transfer' in propositions. This is because propositions are assessed by the Court, but DNA transfer is a factor that scientists need to take into account for the interpretation of their results. Suitable activity level propositions are ideally set before knowledge of the results and address issues like: X stabbed Y vs. an unknown person stabbed Y but X met Y the day before. The scientist assigns the probability of the evidence, if each of the alternate propositions is true, to derive a likelihood ratio. To do this, the scientist asks: a) "what are the expectations if each of the propositions is true?" b) "What data are available to assist in the evaluation of the results given the propositions?" When presenting evidence, scientists work within the hierarchy of propositions framework. The value of evidence calculated for a DNA profile cannot be carried over to higher levels in the hierarchy - the calculations given sub-source, source and activity level propositions are all separate. A number of examples are provided to illustrate the principles espoused, and the criteria that such assessments should meet. Ideally in order to assign probabilities, the analyst should have/collect data that are relevant to the case in question. These data must be relevant to the case at hand and we encourage further research and collection of data to form knowledge bases. Bayesian Networks are extremely useful to help us think about a problem, because they force us to consider all relevant possibilities in a logical way. An example is provided.

DNA commission of the International Society of Forensic Genetics (ISFG): Recommendations on the interpretation of Y-STR results in forensic analysis.

Forensic genetic laboratories perform a large amount of STR analyses of the Y chromosome, in particular to analyze the male part of complex DNA mixtures. However, the statistical interpretation of evidence retrieved from Y-STR haplotypes is challenging. Due to the uni-parental inheritance mode, Y-STR loci are connected to each other and thus haplotypes show patterns of relationship on the familial and population level. This precludes the treatment of Y-STR loci as independently inherited variables and the application of the product rule. Instead, the dependency structure of Y-STRs needs to be included in the haplotype frequency estimation process affecting also the current paradigm of a random match probability that is in the autosomal case approximated by the population frequency assuming unrelatedness of sampled individuals. Information on the degree of paternal relatedness in the suspect population as well as on the familial network is however needed to interpret Y-chromosomal results in the best possible way. The previous recommendations of the DNA commission of the ISFG on the use of Y-STRs in forensic analysis published more than a decade ago [1] cover the interpretation issue only marginally. The current recommendations address a number of topics (frequency estimators, databases, metapopulations, LR formulation, triage, rapidly mutating Y-STRs) with relevance for the Y-STR statistics and recommend a decision-based procedure, which takes into account legal requirements as well as availability of population data and statistical methods.

X-chromosome STR sequence variation, repeat structure, and nomenclature in humans and chimpanzees.

Qualitative information on the sequence composition of the allele and locus structure of the X-STRs DXS8378, DXS9898, DXS6789, GATA31E08, and GATA172D05 was generated in this study. Sequence data were obtained from chimpanzees (Pan troglodytes) and diverse human population groups including Africans, Caucasians, Asians, African-Americans, and Hispanics. Results revealed DXS8378 as the most stable locus. On the other hand, DXS9898 and GATA172D05 showed unstable regions identified through chimpanzee-human sequence comparison. At DXS6789, intra-allelic variation was found in all human populations, i.e., alleles with same fragment sizes showed structural differences only detected by sequencing. At the GATA31E08 locus, a previously unreported variation between humans and chimpanzees was identified in an adjacent region upstream from the repeat. This resulted in the addition of two repeat units and the proposal of a new allele nomenclature at this locus. Also, the sequence analyses did not detect ethnic differences between the studied population samples that would justify the use of these markers to help identify ethnic origin in an anthropological context.

Validation of testing and interpretation protocols for low template DNA samples using AmpFlSTR Identifiler.

AIM: To test the reliability, robustness, and reproducibility of short tandem repeat (STR) profiling of low template DNA (LT-DNA) when employing a defined set of testing and interpretation parameters. METHODS: DNA from known donors was measured with a quantitative real time polymerase chain reaction (PCR) assay that consistently detects less than 1 pg/microL of DNA within a factor of 0.3. Extracts were amplified in triplicate with AmpFlSTR Identifiler reagents under enhanced PCR conditions. Replicates were examined independently and alleles confirmed using a consensus approach. Considering observed stochastic effects inherent to LT-DNA samples, interpretation protocols were developed and their accuracy verified through examination of over 800 samples. RESULTS: Amplification of 100 pg or less of DNA generated reproducible results with anticipated stochastic effects. Down to 25 pg of DNA, 92% or more of the expected alleles were consistently detected while lower amounts yielded concordant partial profiles. Although spurious alleles were sometimes observed within sample replicates, they did not repeat. To account for allelic dropout, interpretation guidelines were made especially stringent for determining homozygous alleles. Due to increased heterozygote imbalance, stutter filters were set conservatively and minor components of mixtures could not be resolved. Applying the resultant interpretation protocols, 100% accurate allelic assignments for over 107 non-probative casework samples, and subsequently 319 forensic casework samples, were generated. CONCLUSION: Using the protocols and interpretation guidelines described here, LT-DNA testing is reliable and robust. Implementation of this method, or one that is suitably verified, in conjunction with an appropriate quality control program ensures that LT-DNA testing is suitable for forensic purposes.

Malignant tumors and forensics--dilemmas and proposals.

AIM: To evaluate the effect of genetic instability and degradation in archived histology samples from cancerous tumors and to investigate the validity of short tandem repeat (STR) typing of these samples and its potential effect on human identification. METHODS: Two hundred and twenty eight slides of archival pathology tissues from 13 different types of malignant tumors were compared with healthy tissues from the same individuals. DNA analysis was performed using standard techniques for forensic STR analysis, PowerPlex16 and Identifiler on 2 distinct sample sets. Genetic instability was assessed by comparing reference tissues with cancerous tissues derived from the same individual. Loss of heterozygosity, a > or =50% reduction in heterozygosity ratio between healthy and diseased samples, and microsatellite instability, the presence of an additional allele not present in reference tissue, were assessed. The quality of profiles obtained with respect to completeness among the archived samples and degradation using the 2 platforms were also compared. RESULTS: Profiles obtained using the Identifiler system were generally more complete, but showed 3-fold higher levels of instability (86%) than those obtained using PowerPlex 16 (27%). Instances of genetic instability were distributed throughout all loci in both multiplex STR systems. CONCLUSION: After having compared 2 widely used forensic chemistries, we suggest individual validation of each kit for use with samples likely to exhibit instability combined with fixation induced degradation or artifact. A "one size fits all" approach for interpretation of these samples among commercially available multiplexes is not recommended.

Combined DNA Typing and Protein Identification from Unfired Brass Cartridges,,.

Biological evidence analysis from contact traces is adversely affected by low quantity and quality of DNA. Proteins in these samples contain potentially individualizing information and may be particularly important for difficult surfaces such as brass, where DNA may yield incomplete profiles. In this study, touched unfired brass cartridges were sampled using dry tape or wet swabs and analyzed by separating DNA and protein from the same collected material, thus producing both genomic and proteomic information. DNA recovery was similar for both collection methods, with tape yielding an average of 1.36 ± 1.87 ng and swabs, 1.34 ± 3.04 ng. Analysis by mass spectrometry identified 95 proteins, with the two collection methods showing no significant difference (p = 0.76) in the average number of collected proteins: 44.5 ± 10.9, (tape) versus 47.9 ± 20.4 (swabs). Proteins can be collected from fingerprints at levels necessary to provide identifying information, thus expanding information obtained from challenging evidence.